V I EWS AR T I C L E

Intermediate filament protein evolution and protists

Harald Preisner | J€orn Habicht | Sriram G. Garg | Sven B. Gould

Institute for Molecular Evolution, Heinrich-

Heine-University, D€usseldorf, Germany

Correspondence

Sven B. Gould, Institute for Molecular

Evolution, Heinrich-Heine-University,

D€usseldorf, Germany

Email: gould@hhu.de

Funding information

Deutsche Forschungsgemeinschaft, Grant

Number: CRC1208/A04

AbstractMetazoans evolved from a single protist lineage. While all eukaryotes share a conserved actin and

tubulin-based cytoskeleton, it is commonly perceived that intermediate filaments (IFs), including

lamin, vimentin or keratin among many others, are restricted to metazoans. Actin and tubulin pro-

teins are conserved enough to be detectable across all eukaryotic genomes using standard

phylogenetic methods, but IF proteins, in contrast, are notoriously difficult to identify by such

means. Since the 1950s, dozens of cytoskeletal proteins in protists have been identified that seem-

ingly do not belong to any of the IF families described for metazoans, yet, from a structural and

functional perspective fit criteria that define metazoan IF proteins. Here, we briefly review IF pro-

tein discovery in metazoans and the implications this had for the definition of this protein family.

We argue that the many cytoskeletal and filament-forming proteins of protists should be incorpo-

rated into a more comprehensive picture of IF evolution by aligning it with the recent

identification of lamins across the phylogenetic diversity of eukaryotic supergroups. This then

brings forth the question of how the diversity of IF proteins has unfolded. The evolution of IF pro-

teins likely represents an example of convergent evolution, which, in combination with the speed

with which these cytoskeletal proteins are evolving, generated their current diversity. IF proteins

did not first emerge in metazoa, but in protists. Only the emergence of cytosolic IF proteins that

appear to stem from a nuclear lamin is unique to animals and coincided with the emergence of

true animal multicellularity.

K E YWORD S

cytoskeleton, eukaryote evolution, intermediate filaments, lamins, multicellularity

1 | INTRODUCTION

The eukaryotic cell is characterized by a list of different traits. Among

these traits are elaborate compartmentalization (nucleus, mitochond-

rion, vacuole, peroxisomes, endoplasmic reticulum, and intracellular

vesicle trafficking, etc.), recombination, meiosis and sex, and also an

intricate cytoskeleton. Among prokaryotic lineages we can observe

individual structures and features analogous to those traits, but not in

combination (Lane & Martin, 2010) and with regard to the cytoskele-

ton, its complexity or dynamics are never tantamount to that of eukar-

yotes. Three main pillars of filament-forming structures have been

characterized, which together configure the cytoskeleton of a eukaryo-

tic cell. These are the actin filaments (AFs; also known as microfila-

ments), microtubules (MTs) and intermediate filaments (IFs).

Briefly, AFs are depicted as polymers of 5–9 nm in diameter that

are composed of two actin strands wrapping around each other form-

ing a helix. The formation of such an actin-based double helix is a

property unique to eukaryotes (Erickson, 2017; Ghoshdastider, Jiang,

Popp, & Robinson, 2015). MTs appear as hollow cylindrical tubes that

are assembled from globular tubulin heterodimers that form a ring of

typically 14—variations from 8 to 20 have been reported—parallel pro-

tofilaments with an outer diameter of approximately 25 nm (Huber

et al., 2013). Both AFs and MTs fulfill a vast number of different duties

in eukaryotic cells (Erickson, 2017; Wickstead & Gull, 2011). They are

associated with fission, vesicle, and organelle trafficking (Akhmanova &

Steinmetz, 2015; Skau et al., 2011) and are required for the two types

of forces that eukaryotic cells can translate into locomotion: actin-

based gliding (through amoeboid pseudopodia or apicomplexan gliding)

and tubulin-based flagella-driven swimming (Fr�enal, Dubremetz, Leb-

run, & Soldati-Favre, 2017; Fritz-Laylin et al., 2010; Kusdian, Woehle,

Martin, & Gould, 2013).

Monomeric actin and tubulin, as well as their polymerized

filaments, interact with a large set of different accessory proteins

(motor-, nucleating-, depolymerization-, and crosslinking proteins) that

Cytoskeleton. 2018;75:231–243. wileyonlinelibrary.com/journal/cm VC 2018Wiley Periodicals, Inc. | 231

Received: 24 January 2018 | Revised: 2 March 2018 | Accepted: 12 March 2018DOI: 10.1002/cm.21443

http://orcid.org/0000-0002-2038-8474

are mostly unique to each of the two cytoskeletal components (Akhma-

nova & Steinmetz, 2015; Huber et al., 2013). The large number of

accessory proteins also pays tribute to the complex machineries that

underlie the mechanisms that orchestrate the flexible states of the

cytoskeleton, which ranges from rigid structural scaffolds to highly

dynamic machineries that engage in the transport of cargo. Cytoskeletal

dynamics are very pronounced in some protists, in which cell shape-

changing processes can reach speeds ten-times those recorded for the

fastest measured muscle contraction (Marshall, 2008). Accessory pro-

teins, but in particular the actin and tubulin proteins themselves, are

conserved across all eukaryotic supergroups and many can be traced

back to the root of eukaryotes (Fritz-Laylin et al., 2010; Kusdian et al.,

2013; Wickstead & Gull, 2011) (Figure 1). For the third major compo-

nent of the eukaryotic cytoskeleton, IF proteins, the situation is more

involved.

IF proteins are part of a far more diverse family. In mammals

alone, about 70 different IF protein-encoding genes have been identi-

fied (Goldman, Cleland, Murthy, Mahammad, & Kuczmarski, 2012;

Snider & Omary, 2014) that can be assigned to one of the six IF pro-

tein classes that are currently recognized (Coulombe & Wong, 2004;

Guerette, Khan, Savard, & Vincent, 2007; Herrmann, Strelkov, Bur-

khard, & Aebi, 2009). Based on criteria such as sequence homology,

assembly behavior, overall structure, and solubility properties, these

classes are: type I keratins (acidic), type II keratins (basic), type III that

for instance include vimentin and desmin, type IV which are neuro-

and muscle-associated filaments made of NF-L, -M, -H, and nestin,

type V that unites the lamins (of the nuclear lamina) and type VI

whose members are often described as orphans (Coulombe & Wong,

2004; Guerette et al., 2007; Herrmann & Aebi, 2004). It remains

uncertain whether nestin (Fig. 2B), syncoilin and synemin, among

others, should be classified as type IV or type VI proteins, as they

deviate in their structure from the more canonical IF proteins (Guer-

ette et al., 2007). Each type is rather specific regarding the tissue in

which it is found and the partners its members interact with, indicating

a specialization regarding the functional spectrum of individual IF pro-

tein families in multicellular eukaryotes. In general, however, the unify-

ing duty of intermediate filaments and their few associated proteins

lies mainly in buffering shearing forces and the support of membranes

(Lazarides, 1980; Goldman et al., 2012; Snider & Omary, 2014). IF pro-

teins are first and foremost stress absorbers.

Whereas actin and tubulin have been identified in all five eukaryo-

tic supergroups (Opisthokonta, Amoebozoa, Archaeplastida, Excavata,

and the SAR lineage; Figure 1) due to their sequence conservation, the

same cannot be said about IF proteins (Eriksson et al., 2009; Herrmann

FIGURE 1 Presence–absence pattern of cytoskeletal proteins across major eukaryotic lineages. Black dots show the identification of ahomolog through a BLAST search (30% sequence identity, e value of 1.0 e210) using human sequences as the query when available, and incase of the protist IF proteins from the species in which it was originally identified (e.g., tetrin of Tetrahymena, giardin of Giardia, etc; seeSupporting Information Table 1 for details). For those cases where BLAST did not retrieve a hit, an additional HMMER search was run toidentify remote homologs (red dots). Note that in some cases neither BLAST nor HMMER identifies homologs although they are known tobe present (pink dots; no claim to completeness). Note that such identified homologs only refer to a top hit retrieved, but they canrepresent false-positives and not necessearily functional orthologs. Bioinformatic approaches alone do not reflect the true nature ofcytoskeletal diversity and are only partly able to uncover the evolution of the individual components [Color figure can be viewed atwileyonlinelibrary.com]

232 | PREISNER ET AL.

http://wileyonlinelibrary.com

et al., 2009; Fuchs & Weber, 1994). Scientific literature (Coulombe,

Bousquet, Ma, Yamada, & Wirtz, 2000; Gruenbaum & Foisner, 2015;

Herrmann & Strelkov, 2011; Kim & Coulombe, 2007; Kornreich, Avi-

nery, Malka-Gibor, Laser-Azogui, & Beck, 2015; Melcer, Gruenbaum, &

Krohne, 2007) as well as current teaching material (Alberts et al., 2015;

Lodish et al., 2016), tell us that bona fide IF proteins are restricted to

metazoans. For protists, which are about a billion years older than met-

azoans (Shu, Isozaki, Zhang, Han, & Maruyama, 2014; Suzuki & Oba,

2015) and make up the remaining phylogenetic diversity of eukaryotes,

no apparent homologs to any of the metazoan IF protein families—

except for lamin (Kollmar, 2015; Koreny & Field, 2016), but more on

that later—have been identified. Why?

Virtually all of the knowledge we gained about IF proteins through-

out the last 5–6 decades stems from studies on metazoans. The prop-

erties that have been recognized and are used to identify a potential IF

protein, in particular the sequence pattern and order of the alpha-

helical rod domain, is inferred solely from multicellular organisms,

moreover almost entirely from higher vertebrates. The majority of

eukaryotic life, however, is single-celled, and protists unite the vast

majority of the genetic and, we would argue, also cytoskeletal diversity

that has been explored (Dawson & Paredez, 2013). The underlying pro-

teins of this diversity are largely undiscovered due to an imbalance

regarding the availability of eukaryotic genomes (Sibbald & Archibald,

2017) and our focus on model species. One is prone to question

whether this bias has influenced the recognition of potential IF protein

families in protists. Only recently, genes encoding homologs of lamin

(including their characteristic domains such a CDK1 phosphorylation

consensus, a CaaX prenylation motif, an Ig-like LTD domain, and a

nuclear localization signal) were identified across a broad range of

eukaryotic lineages (Kollmar, 2015; Koreny & Field, 2016; Kruger et al.,

2012). This is a good time to revisit our perception about IF proteins,

keeping eukaryotic diversity in mind.

Here we briefly review the history of metazoan IF proteins and in

light of recent findings revisit the many protein families identified in

protists throughout the last few decades that are often decribed as

‘filament-forming’ or to be ‘IF-like’. There are plenty of proteins with

such labels and they share many similarities with metazoan IF proteins

in terms of their structure and function, while sharing no recognizable

FIGURE 2 Diversity of repetitive motifs and domain architecture among exemplary intermediate filament proteins. (a) Canonicalintermediate filament proteins of humans, including, for example, vimentin and desmin, contain repetitive motifs with an amino acid bias(low complexity regions) that can also be part of the regions forming the coiled-coils. These proteins are thought to share a common origin(tracing back to a duplication of a lamin gene), but evolve so rapidly that they are not alignable. Similar motifs with a similar amino acid biasand partly also predicted to form coiled-coils, are found among a vast number of filament-forming cytoskeletal proteins identified in variousprotists. Sometimes these repetitive motifs are highly conserved like in articulin, sometimes they are more diverse like in alveolin (a cytos-keletal protein supporting the membranous alveolar sacs of alveolate organisms such as the apicomplexan Toxoplasma and the ciliate Tetra-hymena). (b) Domain distribution, as predicted by SMART (which can be imcomplete, as e.g. NE81 is known to contain an NLS motif aswell), among different cytoskeletal proteins. For all proteins shown, evidence has been presented for them to be part of the cytoskeletonand support cells or cellular compartments structurally. SF-assemblin of Chlamydomonas, for example, forms striated fibers that supportmicrotubules. The SF assembling domain is also retrieved for giardin (which form filaments with a diameter of 2.5 nm), but the primarysequence of the two proteins shares no homology. It highlights the diversity of these filament-forming proteins and their complex, likelynonlinear origins [Color figure can be viewed at wileyonlinelibrary.com]

PREISNER ET AL. | 233

http://wileyonlinelibrary.com

primary sequence identity with any one of them. We conclude that IF

proteins evolved convergently many times throughout eukaryotic evo-

lution, which furthermore questions the restrictive use of the term IF

protein for metazoans.

2 | F IRST THINGS FIRST: THEPROKARYOTIC CYTOSKELETON

Prokaryotes were long thought to lack a cytoskeleton altogether and

the components and structures of the cytoskeleton to be, by and large,

restricted to eukaryotes. Throughout the last two decades, cytoskeletal

proteins have now been identified in many different prokaryotes, but

their number and complexity still remains manageable. Despite this

reduced complexity, prokaryotes still have access to eukaryote-like

‘superstructures’ such as individual rods, rings, twisted filament pairs,

spirals or sheets to name a few (Erickson, 2017; Pilhofer & Jensen,

2013). These structures are often associated with the different mor-

phologies of some bacteria (Ausmees, Kuhn, & Jacobs-Wagner, 2003;

Kysela, Randich, Caccamo, & Brun, 2016). Among the first proteins of

the eukaryotic cytoskeleton identified in prokaryotes were a few dis-

tant homologs of actin and tubulin, but genes encoding IF-like proteins

are only sporadically found among the many thousand prokaryotic

genomes sequenced and their origin is not as clear cut as that of actin

and tubulin.

MreB, crenactin, FtsA, MamK, AlfA, and ParM are prokaryotic

homologs of actin. MreB, the most frequently found prokaryotic homo-

logue of actin, forms protofilaments reminiscent of eukaryotic actin fila-

ments in vitro (van den Ent et al., 2001) and builds a higher ordered

structure subtending the plasma membrane of rod-shaped bacteria

(Jones, Carballido-Lopez, & Errington, 2001). The formation of an obli-

gate right-handed and double-stranded helical, however, is so far

unique to eukaryotes (Ghoshdastider et al., 2015). Actin homologs with

the highest sequence identity to eukaryotic genes were recently identi-

fied among the ASGARD archaea (Zaremba-Niedzwiedzka et al., 2017),

which, in contrast, encode only distantly related homologs of eukaryo-

tic tubulin, that is, the FtsZ superfamily. In the absence of any enriched

culture that one could study through the microscope, their morphology

and the potential manifestation of a more complex cytoskeleton

remains obscure.

FtsZ, not to be confused with FtsA which is a homolog of actin,

shares a high structural similarity with tubulin and likely represents the

prokaryotic homolog of eukaryotic tubulin (Wickstead & Gull, 2011).

FtsZ and proteins with FtsZ/tubulin superfamily folds are found mainly

in bacteria and to a lesser degree in archea (Duggin et al., 2015; Wick-

stead & Gull, 2011). Prokaryotic FtsZ proteins are less well conserved

across prokaryotes than tubulin is conserved across eukaryotes, and

prokaryotic FtsZ-based protofilaments do not form microtubules and

interact with each other only laterally (Lowe & Amos, 2000). Next to

the proteins TubZ and RepX, BtubA and BtubB (of the gram-negative

Prosthecobacter) are the two homologs with some of the highest

sequence identity to eukaryotic a- and b-tubulin (Larsen et al., 2007;

Tinsley & Khan, 2006). It has been suggested that BtubA and BtubB

might have been acquired horizontally from eukaryotes through gene

transfer (horizontal gene transfer, HGT), also because of their isolated

phylogenetic position (Jenkins et al., 2002; Sontag, Sage, & Erickson,

2009). Noticeable is the lack of specialized motor proteins such as kine-

sin, dynein and myosin in all prokaryotes, underscoring the more rigid

nature of the prokaryotic cytoskeleton in comparison to the eukaryotic

one. Motor proteins likely evolved in the last eukaryotic common ances-

tor (LECA) and before the radition of the eukaryotic supergroups

(Wickstead, Gull, & Richards, 2010; Wickstead & Gull, 2011).

The first intermediate filament-like protein of a prokaryote identi-

fied was crescentin (CreS) of Caulobacter crescentus (Ausmees et al.,

2003). Its unique presence in Caulobacter, together with a noticeably

similar coiled-coil organization to the human lamin A and keratin19

(and even some sequence identity), suggests the acquisition of this IF

protein from a eukaryotic source through HGT (Wickstead & Gull,

2011). Other prokaryotic genes with homology to eukaryotic IF pro-

teins include FilP of streptomyces (Fuchino et al., 2013), the cytoplas-

mic CfpA and Scc of spirochetes and AglZ of Myxococcus xanthus

(Izard, Samsonoff, Kinoshita, & Limberger, 1999; Mazouni et al., 2006;

Yang et al., 2004). The coiled-coil rich proteins (Ccrp’s) of Helicobacter

pylori have been suggested to represent a novel class of bacterial cyto-

skeleton proteins (Specht, Schatzle, Graumann, & Waidner, 2011;

Waidner et al., 2009). The sporadic distribution of such filament-

forming (IF) proteins across prokaryotic genomes speaks in favor of

an independent origin in each lineage, either through convergent

evolution of orphan genes or the acquisition through eukaryote-to-

prokaryote HGT.

Prokaryotes also encode one additional class of filament-forming

proteins, which are conserved across many phylogenetically diverse

prokaryotes: the Walker A Cytoskeletal ATPases or in short WACAs

(Michie & Lowe, 2006). One well-known example is the bacterial

MinD, which forms a filament-based scaffold at the cell’s periphery and

inhibits Z-ring formations during division in E. coli (Pichoff & Lutken-

haus, 2001; Shih, Le, & Rothfield, 2003). MinD is also found in some

eukaryotes as part of the plastid division apparatus (Colletti et al.,

2000; de Vries & Gould, 2017; Miyagishima, 2011; Pyke, 2010). Like

FtsZ that is found in such (mostly phototrophic) eukaryotes, MinD is a

legacy of the cyanobacterial origin of the plastid (Nakanishi, Suzuki,

Kabeya, & Miyagishima, 2009; Zimorski, Ku, Martin, & Gould, 2014).

Whether MinD, and its partner MinE, are plastid or nuclear-encoded

might influence the number of plastids present in each cell (de Vries &

Gould, 2017), which highlights the role of cytoskeletal scaffolds in reg-

ulating organelle and cell division, both in prokaryotes and eukaryotes.

3 | THE DEFINITION OF IF PROTEINSIS FOUNDED UPON METAZOA,MAINLY VERTEBRATES

The earliest records of intermediate filaments are likely of those identi-

fied in muscle cells of chick embryos. Electron-microscopy revealed

fibrous material with a diameter of approximately 10 nm in the cytosol,

a size ranging between that of thick and thin myofibrils of muscle tis-

sue, hence their name ‘intermediate’ filaments (Ishikawa, Bischoff, &

Holtzer, 1968). Only later that eponymous term became more

234 | PREISNER ET AL.

commonly used to indicate their intermediate width ranging between

that of AFs and MTs (Fuchs & Weber, 1994). Soon after, it was realized

that these new filament types were not restricted to muscle cells, but a

common component of all types of cells from different metazoan tis-

sues (Herrmann & Strelkov, 2011).

The first information on the structure of IFs had been collected

many decades earlier through X-ray diffraction examinations of hair

and wool fibers harboring keratins (Astbury & Woods, 1934; Pauling &

Corey, 1953). The implementation of cloning and cDNA sequencing

techniques, later combined with computer-based model building,

allowed the characterization of the primary and secondary structure of

keratin and that of other IF proteins downstream (Dowling, Parry, &

Sparrow, 1983; Hanukoglu & Fuchs, 1982; McLachlan, 1978; Parry &

Fraser, 1985; Steinert, Rice, Roop, Trus, & Steven, 1983). The hierarchi-

cal packing order of metazoan IFs was proposed based on scanning-

transmission electron microscopy (Aebi, H˚Ner, Troncoso, Eichner, &

Engel, 1988; Alberts et al., 2015; Engel, Eichner, & Aebi, 1985;

Goldman et al., 2012; Heins et al., 1993; Herrmann et al., 1996; Steven,

Hainfeld, Trus, Wall, & Steinert, 1983). Through these analyses, the bio-

chemical and structural properties of IF proteins from different meta-

zoan—and only metazoan—tissues were determined.

Several individual criteria for the identification of IF proteins have

been worked out. First, they are resistant against the extraction

with buffers based on a high salt- and nonionic detergent content

(Herrmann & Strelkov, 2011). Second, in principal, they have the intrinsic

ability to form filaments autonomously (Fuchs & Weber, 1994). Third,

their primary sequence visualizes a tripartite structure: a central

a-helical rod domain (made up from repetitive motifs) that takes up

most of the protein sequence, which is flanked by non-a-helical seg-

ments: a head- (N-terminus) and tail-domain (C-terminus). The rod-

domain is described as being conserved in length and in the arrangement

of repetitive segments (coil 1A, coil 1B, coil 2A, coil 2B) containing hep-

tad repeats, which are essential for the formation of superhelical coiled-

coil dimers. The segments are interrupted by different linker domains

(L1, L12, and L2) and a stutter region that is an irregularity in the heptad

repeat pattern (Fuchs & Weber, 1994; Herrmann et al., 2009). More-

over, conserved consensus sequences in coil 1A and coil 2B are often

mentioned and viewed as crucial for higher ordered IF protein assembly

(Herrmann & Strelkov, 2011). In contrast, the head- and the tail-domain

vary in size, sequence and substructure, presumably due to their interac-

tions with different proteins in the context of individual and specific

functions (Fuchs & Weber, 1994). The discovery of IF proteins in verte-

brates continues to influence their definition and has set the benchmark

for the identification and classification of IF proteins ever since. Recent

developments, however, demonstrate they have their flaws.

4 | WHEN THE DEFINITIONS FAILAND LAMINS ARE NO LONGER AMETAZOAN-SPECIFIC PROTEIN FAMILY

Nestin, a stem cell marker protein, differs strongly from other IF pro-

teins (Neradil & Veselska, 2015). It contains an unusual short amino-

terminal head- and a very long carboxy-terminal tail-domain (Figure 2b)

(Guerette et al., 2007). Thus, the usually central located a-helical rod-

domain is not centrally positioned and nestin can furthermore not

assemble into bona fide IF by itself but needs an interaction partner

such as a-internexin or vimentin to eventually be incorporated into an

existing filament (Herrmann et al., 2009; Steinert et al., 1999). There is

an ambiguity, whether nestin should be assigned to IF protein class IV

or to VI together with the so-called orphan and unconventional IF pro-

teins. The same is true for the proteins syncoilin, synemin, tanabin (spe-

cific to amphibians), transitin, and paranemin (specific to birds), all of

which indicate similar deviations (Guerette et al., 2007; Herrmann

et al., 2009).

It was once thought that insects do not require IF proteins due to

their stabilizing chitin-based exoskeleton (Herrmann & Strelkov, 2011)

and that the metazoan fresh-water polyp Hydra attenuata represents

the most basal branching eukaryote to possess IF proteins (Erber et al.,

1999; Peter & Stick, 2015). Both views changed with the discovery of

isomin, a cytoplasmic IF protein in the hexapod Isotomurus maculatus

(Mencarelli, Ciolfi, Caroti, Lupetti, & Dallai, 2011) and the identification

of the lamin homolog NE81 in the amoeba Dictyostelium discoideum

(Kruger et al., 2012). Lamins build up the nuclear lamina, a dense mesh-

work subtending the inner nuclear envelope membrane. Crucial for

their integration into the nucleus is a nucleus localization signal (NLS)

and a CaaX-motif within the carboxy-terminal tail-domain. The pres-

ence of these two domains are one of the major structural differences

that distinguish lamins from cytoplasmic IF proteins (Peter & Stick,

2015). Lamin is considered to represent the most primordial IF protein

(Dodemont, Riemer, & Weber, 1990; Erber et al., 1999), which is why

the identification of lamin homologs throughout eukaryotic diversity

(Kollmar, 2015; Koreny & Field, 2016) was a game changer, as it dem-

onstrated this nuclear IF protein not to be a metazoan invention.

When sequences of nuclear lamins became available, it was ob-

vious that they share homologies to cytoplasmic IF proteins (McKeon,

Kirschner, & Caput, 1986). Theory has it that a duplication event of a

‘ur-lamin’ and the subsequent loss of the NLS and the CaaX-motif,

resulted in the first ‘ur-cytoplasmic IF protein’ early in metazoan evolu-

tion (Peter & Stick, 2015). Due to the presence of lamin in eukaryotic

supergroups other than metazoan (Kollmar, 2015; Koreny & Field,

2016), one must now conclude that an ‘ur-lamin’ was part of the pro-

teins encoded by the LECA. Whether lamin was lost in the eukaryotic

groups that apparently lack it (and if so why) or whether it has evolved

beyond a point that it is recognizable remains an open question. Lamins

from different nonopisthokont species, however, localize to the nucleus

of human cells and here form filaments (Koreny & Field, 2016), sug-

gesting the protein’s function is ancient and has remained conserved.

The likely presence of lamin in the LECA requires a reevaluation of IF

protein evolution and questions the restrictive use of the term for met-

azoan proteins.

5 | THE CHALLENGES OF INTERMEDIATEFILAMENT PROTEIN IDENTIFICATION

The identification of IF proteins beyond metazoans is an onerous task.

Metazoan IF proteins are primarily defined and identified through the

PREISNER ET AL. | 235

tertiary structure they are predicted to form and their biochemical

properties such as detergent-resistance, but not primary amino acid

sequence (Herrmann & Strelkov, 2011; Peter & Stick, 2015). IF protein

families exhibit a strong heterogeneity, leading to uncertain assign-

ments among the individual classes. Coiled-coil motifs of non-

homologous protein families can display a similar amino acid composi-

tion, due to their heptad repeats being essential for the formation of

helices, but again not primary amino acid sequence. The sequences

that are predicted to form the coiled-coil motifs are rapidly evolving. In

some cases only the non-repetitive parts of homologous genes from

phylogenetically closely-related species are conserved, while the repeti-

tive middle domains forming the coiled-coils share no sequence iden-

tity whatsoever (Gould et al., 2011). Hence, almost always, BLAST-

based searches that rely on primary sequence conservation are inept

regarding the identification of IF proteins among emerging genome

data, especially from evolutionary distant eukaryotes.

Hidden Markov model-based searches such as HMMER can

uncover distant homologs (Figure 1), but a ‘positive’ hit provides no

certainty regarding the function of the identified protein, whether

through BLAST or HMMER. Giardia is thought to lack all canonical

actin-associated proteins (Paredez et al., 2011), yet standard searches

using common cutoff values identify, for example, Arp2/3 and plastin

in the excavate parasite (Figure 1). This can occur due to only a partial

overlap of the sequences analyzed (e.g. that of a single functional

domain) or even a true evolutionary connection of genes, but with their

products carrying out different functions (Arp2/3 for instance is part of

the actin family). In some cases, a phylogenetic tree dedicated to a sin-

gle gene family can help in making a more reliable call, but eventually

only ‘wet lab’ work will fully resolve such issues. Simultaneously, such

computer-based identifications can miss proteins that are known to be

present and functionally identical to their homolog, too: neither BLAST

nor HMMER identified profilin in the Malaria-agent Plasmodium falcipa-

rum or alveolin in the ciliate Tetrahymena thermophila, although they

encode the respective proteins (El-Haddad et al., 2013; Sch€uler &

Matuschewski, 2006). It demonstrates the imperfection of a plain bioin-

formatic approach and serves as an example regarding the limits when

inferring phenotype from genotype.

Coiled-coil regions are not restricted to proteins of the IF family.

They can be found among a variety of proteins including transcription

factors in which they mediate dimerization (Lee et al., 2017) or viral

proteins in which they serve the interaction with membranes (Buzon

et al., 2010). ARABI-COIL identifies 1,500 coiled-coil containing pro-

teins in Arabidopsis (Rose & Meier, 2004) and almost 10% of the genes

in yeast are predicted to encode a stretch of protein that forms a

coiled-coil (Newman, Wolf, & Kim, 2000). The numbers are likely simi-

lar for all eukaryotes, which in comparison to prokaryotes generally

encode about twice as many coiled-coil containing proteins (Liu & Rost,

2001). However, comparing common IF proteins with those that have

functions other than forming filamentous structures of the cytoskele-

ton, shows there is a tendency regarding the length of the predicted

coiled-coiled regions they harbor. Coiled-coil forming sequences of IF

proteins are usually longer than those found, for example, in transcrip-

tion factors, viral proteins, or the interacting domains of receptor

kinases (Rose & Meier, 2004; Rose, Schraegle, Stahlberg, & Meier,

2005). Still, the presence of a coiled-coil domain alone, which is also

not necessarily always correctly predicted, is no reliable marker for the

identification of IF proteins. Considering in addition the amino acid

composition of repetitive motifs, of which some also form coiled-coil

domains, might help.

The proteome analysis of the detergent-resistant membrane skele-

ton including the epiplasm, a filamentous mesh that subtends the alve-

olar sacs of alveolates such as Tetrahymena thermophila, uncovered an

abundance of proteins that were united by repetitive motifs consisting

of charged (K, E, D) and hydrophobic (L, I, V, P, A) amino acids that out-

numbered the others (Gould et al., 2011). Making use of a search algo-

rithm that took into account this amino acid bias and their presence in

repetitive motifs, identified a significant number of proteins with a simi-

lar tendency among proteins of the cytoskeleton of the excavate para-

site Trichomonas vaginalis that was confirmed by proteome and

localization studies (Preisner et al., 2016). Moreover, a synthetic protein

consisting of a motif (DEVINEQERIKQVIKINGQDLQERKE) that was

based on the identified amino acid bias and arranged in a repetitive

manner, was sufficient to anchor GFP to the cytoskeleton of the ciliate

T. thermophila (El-Haddad et al., 2013). Such features of (protozoan) IF

proteins (Fig. 2), complemented with the simultaneous prediction of

coiled-coil regions using algorithms such as MultiCoil (Wolf, Kim, &

Berger, 1997) or Paircoil2 (McDonnell, Jiang, Keating, & Berger, 2006),

are one way to approach the identification of filament-forming proteins

when BLAST searches fail.

The concept of a functional module that is a “discrete entity whose

function is separable from those of other modules” (Hartwell, Hopfield,

Leibler, & Murray, 1999), can provide one explanation for the diversity

and rapid evolution (Figure 2) of IF proteins (Fleury-Aubusson, 2003).

The ‘actin module’ and the ‘tubulin module’ each fulfill varying duties in

the cell and each interacts with several other separate modules along

the way. This is not the case for the IF module, first and foremost

because no single IF module exists. There is the ‘lamin module’, the

desmin module’ or the ‘keratin module’, but each carries out a rather

specific duty, interacting, if at all, with a very small number of other

proteins. Any mutation in actin or tubulin immediately affects a broad

range of, mostly essential, processes (intracellular transport, locomo-

tion, cytokinesis, etc.). This is not the case for a member of the IF pro-

tein family to the same degree. It is easier for a mutation in an IF

protein to manifest than in proteins of the actin and tubulin family.

Either way, both the origin of metazoan IF proteins (which mainly

appear to stem from an ancient lamin duplication) and the more general

rapid diversification of IF protein sequences is what impairs their iden-

tification across the protozoan to metazoan divide.

6 | PROTISTS INTERMEDIATE FILAMENTPROTEINS ARE PLENTY

A protist’s cortex can be soft or rigid (Cavalier-Smith, 2002) and ulti-

mately fulfills the same functions as that of a metazoan cell. Because a

protist cell is not protected by the multicellular context of metazoan

tissue (Tilney & Tilney, 1996), in paticular those protists that are not

236 | PREISNER ET AL.

TABLE1

Cytoskeleton-associated

proteinsofdive

rseprotists

Protein

Organ

ism

Localization

Referen

ces

Alveo

lins/IM

Cproteins

a Alveo

lin1,bAlveo

lin2,c IMC1,dIM

C3,eIM

C4,f IM

C5,

gIM

C6,hIM

C7,iIM

C8,j IM

C9,kIM

C10,l IM

C11,

mIM

C12,nIM

C13,oIM

C14,pIM

C15

a,bTe

trah

ymenathermop

hila,

c–pTo

xoplasmagond

ii

a alveo

li,bbe

twee

nlong

itud

inal

microtubu

les,

c,d,e,g,h,k,m

,oco

rtical,IM

C,f,i,jba

salco

mplex

,l apicalcap,

basalen

doftheIM

C,

nco

rtical

IMC,ba

salco

mplex

,pco

rtical

IMC,ap

ical

cap,

basalco

mplex

,centrosomes

a Gould

etal.(2008)

bEl-Had

dad

etal.(2013)

c Man

n,Gaskins,an

dBecke

rs(2002)

dGubbels,W

ieffer,an

dStriep

en(2004)

eHuet

al.(2006)

f–pAnderson-W

hiteet

al.(2011)

Apicalpo

larring

proteins

a RNG1,bRNG2

Toxoplasmagond

iiap

ical

polarring

a Tranet

al.(2010)

bKatriset

al.(2014)

Articulins

a 80kD

aArticulin,a 86kD

aArticulin,

bArticulin

1,bArticulin

4,c A

rticulin

p60

a,bEu

glenagracilis,

c Pseud

omicrothorax

dubius

epiplasm

a Marrs

andBouck

(1992)

bHutten

lauch

,Peck,

andStick(1998)

c Hutten

lauch

etal.(1995)

Basal

appa

ratusproteins

a BAp9

0,bBAp9

5Sp

ermatozop

sissimilis

proximal

plates

oftheba

sal

appa

ratus

a Geimer,Le

chtreck,

andMelko

nian(1998b)

bGeimer,Clees,Melko

nian,an

dLe

chtreck(1998a)

Epiplasmins

a EPI11,aEPI20,a EPI30,aEPI38,

a EPI40,aEPI41,bEpiplasmin

C

a Param

ecium

tetrau

relia

bTe

trah

ymenapyriform

isep

iplasm

a Fleury-A

ubussonet

al.(2013)

bBouch

ard,C

homilier,R

avet,M

ornon,andVigues

(2001)

Giardins

a a-1

giardin,

ba-2

giardin,

c ß-giardin,dß-giardin,eDAPs

Giardia

lamblia

a plasm

amem

bran

eoftroph

ozo

ites,

bmicroribb

ons,

c,d,eve

ntraldisk

a,bFelizianiet

al.(2011)

bAlonso

andPea

ttie

(1992),Pea

ttie,Alonso,Hein,a

nd

Cau

lfield

(1989)

c Nohria,

Alonso,an

dPea

ttie

(1992)

eHagen

etal.(2011)

Glid

ing-asso

ciated

proteins

GAP45,GAP50

Plasmod

ium

falciparum

IMC

Bau

m,Pap

enfuss,Bau

m,Sp

eed,&

Cowman

(2006)

Hea

d–stalkproteins

a 183kD

ahe

ad-stalk

protein,

bGASP

-180,c G

HSP

115

Giardia

lamblia

a unk

nown,

baxone

mofthean

teriorflagella,

c plasm

amem

bran

eoftroph

ozo

ites,cytoplasmab

a Marshallan

dHolberton(1995)

bElm

endorf,R

ohrer,Khoury,B

outten

ot,an

dNash(2005)

c Bae

,Kim

,Kim

,Yong,

andPark(2009)

Plateins

alph

a-1Platein,alph

a-2Platein,beta-/gam

maPlatein

Euplotes

aediculatus

alve

olarplates

Kloetzelet

al.(2003)

Septins

a Cdc

10,aCdc

11,a C

dc12,a C

dc3,bSe

p1,

bSe

p2,bSe

p3

a Saccharom

yces

cerevisisae,

bTe

trah

ymenathermop

hile

a cellco

rtex

,bmitoch

ond

ria

a Hartw

ell(1971)

bW

loga,S

trzyzewska-Jowko

,Gae

rtig,an

dJerka-Dziad

osz

(2008)

Tetrins

TetrinA,T

etrinB,T

etrinC

Tetrah

ymena

cytostom

Brimmer

andW

eber

(2000)

(Continues)

PREISNER ET AL. | 237

protected by an additional cell wall or silica shell, have evolved complex

scaffolding structures to support their plasma membrane; their

genomes encode more proteins that potentially form filaments and are

part of a the cytoskeleton (Gould et al., 2011). Their structural cytoskel-

eton can be so well developed that even when detergent-treated (i.e.,

stripped of their plasma membranes), they maintain their shape and the

flagella remain functional as long as they are provided with ATP (Good-

enough, 1983). It has been argued before that ‘cytoskeletal polymers’

of protists find little, if any, recognition when the eukaryotic cytoskel-

ton is discussed (Fleury-Aubusson, 2003). Acknowledging the diversity

of filament forming proteins other than actin and tubluin among eukar-

yotes, will help to identify additional IF proteins in both protists and

metazoans.

Filament-forming proteins with structural properties identical to

those of metazoans IFs are plenty in protists (Table 1) and the proto-

cols used to isolate them are based on those used for vertebrate IF

proteins (Herrmann & Aebi, 2004; Palm et al., 2005; Williams, Honts, &

Jaeckel-Williams, 1987). To name a few, there are (1) the articulins of

euglenoids (Huttenlauch et al., 1998; Marrs & Bouck, 1992) and the (2)

plateins of ciliates (Kloetzel et al., 2003) that build up the membrane

skeleton and exhibit a tripartite structure including a centrally located

and highly repetitive region (Huttenlauch & Stick, 2003), (3) the giardins

of Giardia which are associated with the plasma membrane and the

ventral disk of the excavate parasite (Feliziani et al., 2011; Holberton,

Baker, & Marshall, 1988), (4) MDM1 that is part of an extended net-

work throughout the cytoplasm of Saccharomyces (McConnell & Yaffe,

1993), (5) FAZ1, a filament associated protein essential for a stable

flagellum attachment zone of trypanosomes (Vaughan, Kohl, Ngai,

Wheeler, & Gull, 2008), (6) epiplasmins, major components of the epi-

plasm of ciliates, (7) tetrins of the cytostome of ciliates (Brimmer &

Weber, 2000; Coffe, Le Caer, Lima, & Adoutte, 1996), or (8) alveolins,

membrane skeleton proteins characteristic for all alveolates (Gould,

Tham, Cowman, McFadden, & Waller, 2008). There is a severe bias in

the availability of sequenced eukaryotic genomes (Sibbald & Archibald,

2017) and even less attention has been paid on the analysis of proto-

zoan cytoskeletal components with regard to a connection—even if it is

only a functional one—to metazoan IF proteins.

7 | CONCLUSION

All eukaryotic life evolved from a single lineage and all extant eukaryo-

tic groups share a list of features, both genetic and morphological, that

unite them. Many different lines of evidence suggest that the last euk-

aryotic common ancestor, LECA, was no less complex than your extant

garden variety protist. The LECA was characterized by a versatile (most

likely facultative anaerobic) metabolism, meiosis and sex, both actin-

and tubulin-based motility, and a complex endomembrane system

including vesicle flux and the eponymous nucleus. Therefore, the

recent identification of nuclear lamins in representatives of all major

eukaryotic supergroups comes as no real surprise, yet requests a shift

of perspective. It demonstrates that intermediate filaments evolved

early in eukaryotic evolution and that they are not limited toTABLE1

(Continue

d)

Protein

Organ

ism

Localization

Referen

ces

Other

proteins

FAZ1

Fen

estrin

Fin1

Lamin-likeprotein

MDM1

Med

ianbo

dyprotein

NE81

p477

SALP

-1SF

-assem

blin

SPM1

TgM

E49_0

44470

TgM

E49_0

52880

TTHERM_0

0188980

TTHERM_0

0128280DFB1)

TTHERM_0

0388620

TTHERM_0

0474830

WCB

Trypan

osom

abrucei

Tetrah

ymena

Saccha

romyces

cerevisiae

Amph

idinium

carterae

Saccha

romyces

cerevisiae

Giardia

lamblia

Dictyostelium

Tricho

mon

asvagina

lisGiardia

lamblia

Chlorop

hyceae

Toxoplasma

Toxoplasmagond

iiTo

xoplasmagond

iiTe

trah

ymenathermop

hila

Tetrah

ymenathermop

hila

Tetrah

ymenathermop

hila

Tetrah

ymenathermop

hila

Trypan

osom

abrucei

flagellum

attach

emen

tzo

nebe

neaththeep

iplasm

betw

eenthespindlepo

les

nuclea

rmatrix

cytoplasm

ventraldisk

nuclea

ren

velope

atractoph

or

ventraldisc

basalap

paratus

subp

ellicular

microtubu

les

cellap

excellap

exbe

neaththeplasmamem

bran

ealong

the

long

itud

inal

cilia

withina

periodicarrang

emen

tcytostom

plasmamem

bran

e

Vau

ghan

etal.(2008)

Nelsen,W

illiams,Yi,Knaak,

andFranke

l(1994,2011),

vanHem

ertet

al.(2002)

Mingu

ez,Franca,an

dDelae

spina(1994,2011)

McC

onnellan

dYaffe

(1993,2011)

Woessner

andDaw

son(2012)

Kruge

ret

al.(2012)

Brich

eux,

Coffe,

andBruge

rolle

(2007)

Palm,W

eiland,Griffiths,Ljungstrom,an

dSv

ard(2003)

Lech

treckan

dMelko

nian(1991)

Tran,Li,C

hyan,Chung,

andMorrissette(2012)

Gould

etal.(2011)

Gould

etal.(2011)

Gould

etal.(2011)

Gould

etal.(2011)

Gould

etal.(2011)

Gould

etal.(2011)

Baines

andGull(2008)

Allsharech

aracteristics(form

filamen

ts,co

ntainrepe

titive

motifs,arepred

ictedto

form

long

coiled-co

ils)withmetazoan

IFproteins.

238 | PREISNER ET AL.

metazoans, which is corroborated by the presence of the many sur-

veyed filament-forming proteins of protists that share characteristics

first described for canonical metazoan IF proteins. Protozoan IF pro-

teins share neither sequence homology nor necessarily domain archi-

tecture with cytosolic IF proteins of metazoans, because only the

majority of the latter originate from an ancient lamin-like ancestor. The

LECA potentially possessed intermediate filament proteins other than

lamin, but in different lineages they experienced different fates. Due to

the evidently rapid and likely also convergent evolution, tracking their

exact phylogenetic history remains tedious, if not impossible. Despite

the lack in phylogenetic resolution, one can conclude that intermediate

filament proteins are a universal component of eukaryotic biology and

that many more await their discovery. Like those already characterized,

most of them will likely be unique to the individual group in which they

are identified.

ACKNOWLEDGMENTS

We would like to thank Gary Kusdian for early discussions and

gratefully acknowledge the funding through the DFG (CRC 1208)

and the VolkswagenStiftung to SBG.

ORCID

Sven B. Gould http://orcid.org/0000-0002-2038-8474

REFERENCES

Aebi, U., Häner, M., Troncoso, J., Eichner, R., & Engel, A. (1988). Unifying

principles in intermediate filament (IF) structure and assembly. Proto-

plasma, 145(2–3), 73–81.

Akhmanova, A., & Steinmetz, M. O. (2015). Control of microtubule orga-

nization and dynamics: Two ends in the limelight. Nature Reviews:

Molecular Cell Biology, 16(12), 711–726.

Alberts, B., Johnson, A., Lewis, J., Morgan, D., Raff, M., Roberts, K., &

Walter, P. (2015). Molecular biology of the cell (pp. 889–962). NewYork: Garland Science, Taylor & Francis Group, LLC.

Alonso, R. A., & Peattie, D. A. (1992). Nucleotide sequence of a second

alpha giardin gene and molecular analysis of the alpha giardin genes

and transcripts in Giardia lamblia. Molecular & Biochemical Parasitol-

ogy, 50(1), 95–104.

Anderson-White, B. R., Ivey, F. D., Cheng, K., Szatanek, T., Lorestani, A.,

Beckers, C. J., . . . Gubbels, M. J. (2011). A family of intermediate

filament-like proteins is sequentially assembled into the cytoskeleton

of Toxoplasma gondii. Cellular Microbiology, 13(1), 18–31.

Astbury, W. T., & Woods, H. J. (1934). X-ray studies of the structures of

hair, wool and related fibres. II. The molecular structure and elastic

properties of hair keratin. Transactions of Royal Society London, A232,

333–394.

Ausmees, N., Kuhn, J. R., & Jacobs-Wagner, C. (2003). The bacterial

cytoskeleton: An intermediate filament-like function in cell shape.

Cell, 115(6), 705–713.

Bae, S. S., Kim, J., Kim, T. S., Yong, T. S., & Park, S. J. (2009). Giardia lam-

blia: Immunogenicity and intracellular distribution of GHSP-115, a

member of the Giardia head-stalk family of proteins. Experimental

Parasitology, 122(1), 11–16.

Baines, A., & Gull, K. (2008). WCB is a C2 domain protein defining the

plasma membrane: Sub-pellicular microtubule corset of kinetoplastid

parasites. Protist, 159(1), 115–125.

Baum, J., Papenfuss, A. T., Baum, B., Speed, T. P., & Cowman, A. F.

(2006). Regulation of apicomplexan actin-based motility. Nature

Reviews: Microbiology, 4(8), 621–628.

Bouchard, P., Chomilier, J., Ravet, V., Mornon, J. P., & Vigues, B. (2001).

Molecular characterization of the major membrane skeletal protein in

the ciliate Tetrahymena pyriformis suggests n-plication of an early

evolutionary intermediate filament protein subdomain. Journal of Cell

Science, 114(Pt 1), 101–110.

Bricheux, G., Coffe, G., & Brugerolle, G. (2007). Identification of a new

protein in the centrosome-like “atractophore” of Trichomonas vagina-lis. Molecular & Biochemical Parasitology, 153(2), 133–140.

Brimmer, A., & Weber, K. (2000). The cDNA sequences of three tetrins,

the structural proteins of the Tetrahymena oral filaments, show that

they are novel cytoskeletal proteins. Protist, 151(2), 171–180.

Buzon, V., Natrajan, G., Schibli, D., Campelo, F., Kozlov, M. M., & Weis-

senhorn, W. (2010). Crystal structure of HIV-1 gp41 including both

fusion peptide and membrane proximal external regions. PLoS Patho-

gens, 6(5), e1000880.

Cavalier-Smith, T. (2002). The phagotrophic origin of eukaryotes and

phylogenetic classification of Protozoa. International Journal of Sys-

tematic & Evolutionary Microbiology, 52(Pt 2), 297–354.

Coffe, G., Le Caer, J. P., Lima, O., & Adoutte, A. (1996). Purification, in

vitro reassembly, and preliminary sequence analysis of epiplasmins,

the major constituent of the membrane skeleton of Paramecium. Cell

Motility & the Cytoskeleton, 34(2), 137–151.

Colletti, K. S., Tattersall, E. A., Pyke, K. A., Froelich, J. E., Stokes, K. D., &

Osteryoung, K. W. (2000). A homologue of the bacterial cell division

site-determining factor MinD mediates placement of the chloroplast

division apparatus. Current Biology, 10(9), 507–516.

Coulombe, P. A., Bousquet, O., Ma, L. L., Yamada, S., & Wirtz, D. (2000).

The ’ins’ and ’outs’ of intermediate filament organization. Trends inCellular Biology, 10(10), 420–428.

Coulombe, P. A., & Wong, P. (2004). Cytoplasmic intermediate filaments

revealed as dynamic and multipurpose scaffolds. Nature Cell Biology,

6(8), 699–706.

Dawson, S. C., & Paredez, A. R. (2013). Alternative cytoskeletal land-

scapes: cytoskeletal novelty and evolution in basal excavate protists.

Current Opinion in Cell Biology, 25(1), 134–141.

de Vries, J., & Gould, S. B. (2017, in press). The monoplastidic bottleneck

in algae and plant evolution. Journal of Cellular Science. https://doi.

org/10.1242/jcs.203414

Dodemont, H., Riemer, D., & Weber, K. (1990). Structure of an inverte-

brate gene encoding cytoplasmic intermediate filament (IF) proteins—Implications for the origin and the diversification of IF proteins. Embo

Journal, 9(12), 4083–4094.

Dowling, L. M., Parry, D. A., & Sparrow, L. G. (1983). Structural homology

between hard alpha-keratin and the intermediate filament proteins

desmin and vimentin. Bioscience Reports, 3(1), 73–78.

Duggin, I. G., Aylett, C. H., Walsh, J. C., Michie, K. A., Wang, Q., Turnbull,

L., & L€owe, J. (2015). CetZ tubulin-like proteins control archaeal cell

shape. Nature, 519(7543), 362–365.

El-Haddad, H., Przyborski, J. M., Kraft, L. G. K., McFadden, G. I., Waller,

R. F., & Gould, S. B. (2013). Characterization of TtALV2, an Essential

Charged Repeat Motif Protein of the Tetrahymena thermophila Mem-

brane Skeleton. Eukaryotic Cell, 12(6), 932–940.

Elmendorf, H. G., Rohrer, S. C., Khoury, R. S., Bouttenot, R. E., & Nash,

T. E. (2005). Examination of a novel head-stalk protein family in

PREISNER ET AL. | 239

http://orcid.org/0000-0002-2038-8474https://doi.org/10.1242/jcs.203414https://doi.org/10.1242/jcs.203414

Giardia lamblia characterised by the pairing of ankyrin repeats and

coiled-coil domains. International Journal of Parasitology, 35(9), 1001–1011.

Engel, A., Eichner, R., & Aebi, U. (1985). Polymorphism of reconstituted

human epidermal keratin filaments: Determination of their mass-per-

length and width by scanning transmission electron microscopy

(STEM). Journal of Ultrastructure Research, 90(3), 323–335.

Erber, A., Riemer, D., Hofemeister, H., Bovenschulte, M., Stick, R., Pano-

poulou, G., . . . Weber, K. (1999). Characterization of the Hydra lamin

and its gene: A molecular phylogeny of metazoan lamins. Journal of

Molecular Evolution, 49(2), 260–271.

Erickson, H. P. (2017). The discovery of the prokaryotic cytoskeleton:

25th anniversary. Molecular Biology of the Cell, 28(3), 357–358.

Eriksson, J. E., Dechat, T., Grin, B., Helfand, B., Mendez, M., Pallari, H. M., &

Goldman, R. D. (2009). Introducing intermediate filaments: from dis-

covery to disease. Journal of Clinical Investigation, 119(7), 1763–1771.

Feliziani, C., Merino, M. C., Rivero, M. R., Hellman, U., Pistoresi-Palencia,

M. C., & Ropolo, A. S. (2011). Immunodominant proteins alpha-1 giar-

din and beta-giardin are expressed in both assemblages A and B of

Giardia lamblia. BMC Microbiology, 11(1), 233.

Fleury-Aubusson, A. (2003). Novel cytoskeletal proteins in protists: Intro-

ductory remarks. Journal of Eukaryotic Microbiology, 50(1), 3–8.

Fleury-Aubusson, A., Bricheux, G., Damaj, R., Lemullois, M., Coffe, G.,

Donnadieu, F., . . . Bouchard, P. (2013). Epiplasmins and epiplasm in

paramecium: The building of a submembraneous cytoskeleton. Protist,

164(4), 451–469.

Fr�enal, K., Dubremetz, J. F., Lebrun, M., & Soldati-Favre, D. (2017). Glid-

ing motility powers invasion and egress in Apicomplexa. Nature

Reviews: Microbiology, 15(11), 645–660.

Fritz-Laylin, L. K., Prochnik, S. E., Ginger, M. L., Dacks, J. B., Carpenter,

M. L., Field, M. C., & Dawson, S. C. (2010). The genome of Naegleria

gruberi illuminates early eukaryotic versatility. Cell, 140(5), 631–642.

Fuchino, K., Bagchi, S., Cantlay, S., Sandblad, L., Wu, D., Bergman, J., . . . Aus-

mees, N. (2013). Dynamic gradients of an intermediate filament-like

cytoskeleton are recruited by a polarity landmark during apical growth.

Proceedings of National Academic Sciences USA, 110(21), E1889–E1897.

Fuchs, E., & Weber, K. (1994). Intermediate filaments—Structure, dynamics,function, and disease. Annual Review of Biochemistry, 63, 345–382.

Geimer, S., Clees, J., Melkonian, M., & Lechtreck, K. F. (1998a). A novel

95-kD protein is located in a linker between cytoplasmic microtu-

bules and basal bodies in a green flagellate and forms striated fila-

ments in vitro. Journal of Cellular Biology, 140, 1149–1158.

Geimer, S., Lechtreck, K. F., & Melkonian, M. (1998b). A novel basal

apparatus protein of 90 kD (BAp90) from the flagellate green alga

Spermatozopsis similis is a component of the proximal plates and iden-

tifies the D-(dexter) surface of the basal body. Protist, 149, 173–184.

Ghoshdastider, U., Jiang, S. M., Popp, D., & Robinson, R. C. (2015). In search

of the primordial actin filament. Proceedings of the National Academy of

Sciences of the United States of America, 112(30), 9150–9151.

Goldman, R. D., Cleland, M. M., Murthy, S. N. P., Mahammad, S., & Kucz-

marski, E. R. (2012). Inroads into the structure and function of interme-

diate filament networks. Journal of Structural Biology, 177(1), 14–23.

Goodenough, U. W. (1983). Motile detergent-extracted cells of Tetrahy-

mena and Chlamydomonas. Journal of Cell Biology, 96(6), 1610–1621.

Gould, S. B., Kraft, L. G. K., van Dooren, G. G., Goodman, C. D., Ford, K. L.,

Cassin, A. M., . . . Waller, R. F. (2011). Ciliate pellicular proteome identifies

novel protein families with characteristic repeat motifs that are common

to alveolates.Molecular Biology & Evolution, 28(3), 1319–1331.

Gould, S. B., Tham, W. H., Cowman, A. F., McFadden, G. I., & Waller, R.

F. (2008). Alveolins, a new family of cortical proteins that define the

protist infrakingdom Alveolata. Molecular Biology & Evolution, 25(6),

1219–1230.

Gruenbaum, Y., & Foisner, R. (2015). Lamins: nuclear intermediate fila-

ment proteins with fundamental functions in nuclear nechanics and

genome regulation. Annual Review of Biochemistry, 84, 131–164.

Gubbels, M. J., Wieffer, M., & Striepen, B. (2004). Fluorescent protein

tagging in Toxoplasma gondii: Identification of a novel inner mem-

brane complex component conserved among Apicomplexa. Molecular

& Biochemical Parasitology, 137(1), 99–110.

Guerette, D., Khan, P. A., Savard, P. E., & Vincent, M. (2007). Molecular

evolution of type VI intermediate filament proteins. BMC Evolutionary

Biology, 7, 164.

Hagen, K. D., Hirakawa, M. P., House, S. A., Schwartz, C. L., Pham, J. K.,

Cipriano, M. J., & Dawson, S. C. (2011). Novel structural components

of the ventral disc and lateral crest in Giardia intestinalis. PLoS

Neglected Tropical Diseases, 5(12), e1442.

Hanukoglu, I., & Fuchs, E. (1982). The cDNA sequence of a human epi-

dermal keratin: Divergence of sequence but conservation of structure

among intermediate filament proteins. Cell, 31(1), 243–252.

Hartwell, L. H. (1971). Genetic control of the cell division cycle in yeast.

IV. Genes controlling bud emergence and cytokinesis. Experimental

Cell Research, 69(2), 265–276.

Hartwell, L. H., Hopfield, J. J., Leibler, S., & Murray, A. W. (1999).

From molecular to modular cell biology. Nature, 402(6761 Supp),

C47–C52.

Heins, S., Wong, P. C., Muller, S., Goldie, K., Cleveland, D. W., & Aebi, U.

(1993). The rod domain of NF-L determines neurofilament architec-

ture, whereas the end domains specify filament assembly and net-

work formation. Journal of Cell Biology, 123(6 Pt 1), 1517–1533.

Herrmann, H., & Aebi, U. (2004). Intermediate filaments: molecular struc-

ture, assembly mechanism, and integration into functionally distinct

intracellular scaffolds. Annual Review of Biochemistry, 73, 749–789.

Herrmann, H., Haner, M., Brettel, M., Muller, S. A., Goldie, K. N., Fedtke,

B., . . . Aebi, U. (1996). Structure and assembly properties of the

intermediate filament protein vimentin: the role of its head, rod and

tail domains. Journal of Molecular Biology, 264(5), 933–953.

Herrmann, H., & Strelkov, S. V. (2011). History and phylogeny of inter-

mediate filaments: Now in insects. BMC Biology, 9, 16.

Herrmann, H., Strelkov, S. V., Burkhard, P., & Aebi, U. (2009). Intermedi-

ate filaments: Primary determinants of cell architecture and plasticity.

Journal of Clinical Investigation, 119(7), 1772–1783.

Holberton, D., Baker, D. A., & Marshall, J. (1988). Segmented alpha-

helical coiled-coil structure of the protein giardin from the Giardia

cytoskeleton. Journal of Molecular Biology, 204(3), 789–795.

Hu, K., Johnson, J., Florens, L., Fraunholz, M., Suravajjala, S., DiLullo, C.,

. . . Murray, J. M. (2006). Cytoskeletal components of an invasion

machine: The apical complex of Toxoplasma gondii. PLoS Pathogens, 2

(2), e13–138.

Huber, F., Schnauß, J., R€onicke, S., Rauch, P., M€uller, K., F€utterer, C., &

Käs, J. (2013). Emergent complexity of the cytoskeleton: From single

filaments to tissue. Advances in Physics, 62(1), 1–112.

Huttenlauch, I., Geisler, N., Plessmann, U., Peck, R. K., Weber, K., & Stick,

R. (1995). Major epiplasmic proteins of ciliates are articulins—Cloning,recombinant expression, and structural characterization. Journal of

Cell Biology, 130(6), 1401–1412.

Huttenlauch, I., Peck, R. K., & Stick, R. (1998). Articulins and epiplasmins:

Two distinct classes of cytoskeletal proteins of the membrane skele-

ton in protists. Journal of Cellular Science, 111, 3367–3378.

Huttenlauch, I., & Stick, R. (2003). Occurrence of articulins and epiplas-

mins in protists. Journal of Eukaryotic Microbiology, 50(1), 15–18.

240 | PREISNER ET AL.

Ishikawa, H., Bischoff, R., & Holtzer, H. (1968). Mitosis and intermediate-

sized filaments in developing skeletal muscle. Journal of Cell Biology,

38(3), 538–555.

Izard, J., Samsonoff, W. A., Kinoshita, M. B., & Limberger, R. J. (1999).

Genetic and structural analyses of cytoplasmic filaments of wild-type

Treponema phagedenis and a flagellar filament-deficient mutant. Jour-

nal of Bacteriology, 181, 6739–6746.

Jenkins, C., Samudrala, R., Anderson, I., Hedlund, B. P., Petroni, G.,

Michailova, N., . . . Staley, J. T. (2002). Genes for the cytoskeletal pro-

tein tubulin in the bacterial genus Prosthecobacter. Proceedings of the

National Academy of Sciences of the United States of America, 99(26),

17049–17054.

Jones, L. J. F., Carballido-Lopez, R., & Errington, J. (2001). Control of cell

shape in bacteria: Helical, actin-like filaments in Bacillus subtilis. Cell,

104(6), 913–922.

Katris, N. J., van Dooren, G. G., McMillan, P. J., Hanssen, E., Tilley, L., &

Waller, R. F. (2014). The apical complex provides a regulated gateway

for secretion of invasion factors in Toxoplasma. PLoS Pathogens, 10(4),

e1004074.

Kim, S., & Coulombe, P. A. (2007). Intermediate filament scaffolds fulfill

mechanical, organizational, and signaling functions in the cytoplasm.

Genes & Development, 21(13), 1581–1597.

Kloetzel, J. A., Baroin-Tourancheau, A., Miceli, C., Barchetta, S., Farmar,

J., Banerjee, D., & Fleury-Aubusson, A. (2003). Plateins: A novel fam-

ily of signal peptide-containing articulins in euplotid ciliates. Journal

of Eukaryotic Microbiology, 50(1), 19–33.

Kollmar, M. (2015). Polyphyly of nuclear lamin genes indicates an early

eukaryotic origin of the metazoan-type intermediate filament pro-

teins. Scientific Reports, 5, 10652.

Koreny, L., & Field, M. C. (2016). Ancient eukaryotic origin and evolu-

tionary plasticity of nuclear lamina. Genome Biology & Evolution, 8(9),

2663–2671.

Kornreich, M., Avinery, R., Malka-Gibor, E., Laser-Azogui, A., & Beck, R.

(2015). Order and disorder in intermediate filament proteins. FEBS

Letters, 589(19 Pt A), 2464–2476.

Kruger, A., Batsios, P., Baumann, O., Luckert, E., Schwarz, H., Stick, R., . . .

Graf, R. (2012). Characterization of NE81, the first lamin-like nucleos-

keleton protein in a unicellular organism. Molecular Biology & Cell, 23

(2), 360–370.

Kusdian, G., Woehle, C., Martin, W. F., & Gould, S. B. (2013). The actin-

based machinery of Trichomonas vaginalis mediates flagellate-

amoeboid transition and migration across host tissue. Cellular Microbi-

ology, 15(10), 1707–1721.

Kysela, D. T., Randich, A. M., Caccamo, P. D., & Brun, Y. V. (2016). Diver-

sity takes shape: understanding the mechanistic and adaptive basis of

bacterial morphology. PLoS Biology, 14(10), e1002565.

Lane, N., & Martin, W. (2010). The energetics of genome complexity.

Nature, 467(7318), 929–934.

Larsen, R. A., Cusumano, C., Fujioka, A., Lim-Fong, G., Patterson, P., &

Pogliano, J. (2007). Treadmilling of a prokaryotic tubulin-like protein,

TubZ, required for plasmid stability in Bacillus thuringiensis. Genes &

Development, 21(11), 1340–1352.

Lazarides, E. (1980). Intermediate filaments as mechanical integrators of

cellular space. Nature, 283(5744), 249–256.

Lechtreck, K. F., & Melkonian, M. (1991). Striated microtubule-associated

fibers: Identification of assemblin, a novel 34-kD protein that forms

paracrystals of 2 nm filaments in vitro. Journal of Cellular Biology, 115

(3), 705–716.

Lee, H. W., Kang, N. Y., Pandey, S. K., Cho, C., Lee, S. H., & Kim, J.

(2017). Dimerization in LBD16 and LBD18 transcription factors is

critical for lateral root formation. Plant Physiology, 174(1), 301–311.

Liu, J. F., & Rost, B. (2001). Comparing function and structure between

entire proteomes. Protein Science, 10(10), 1970–1979.

Lodish, H. F., Berk, A., Kaiser, C. A., Krieger, M., Bretscher, A., Ploegh, H.,

. . . Martin, K. C. (2016). Molecular cell biology (pp. 821–871). NewYork: W.H. Freeman and Co.

Lowe, J., & Amos, L. A. (2000). Helical tubes of FtsZ from Methanococcus

jannaschii. Biological Chemistry, 381(9–10), 993–999.

Mann, T., Gaskins, E., & Beckers, C. (2002). Proteolytic processing of

TgIMC1 during maturation of the membrane skeleton Toxoplasma

gondii. Journal of Biological Chemistry, 277(43), 41240–41246.

Marrs, J. A., & Bouck, G. B. (1992). The two major membrane skeletal

proteins (articulins) of Euglena gracilis define a novel class of cytos-

keletal proteins. Journal of Cell Biology, 118(6), 1465–1475.

Marshall, J., & Holberton, D. V. (1995). Giardia gene predicts a 183 kDa

nucleotide-binding head-stalk protein. Journal of Cellular Science, 108,

2683–2692.

Marshall, W. F. (2008). Don’t blink: Observing the ultra-fast contractionof spasmonemes. Biophysical Journal, 94(1), 4–5.

Mazouni, K., Pehau-Arnaudet, G., England, P., Bourhy, P., Saint Girons, I.,

& Picardeau, M. (2006). The SCC spirochetal coiled-coil protein forms

helix-like filaments and binds to nucleic acids generating nucleopro-

tein structures. Journal of Bacteriology, 188, 469–476.

McConnell, S. J., & Yaffe, M. P. (1993). Intermediate filament formation

by a yeast protein essential for organelle inheritance. Science (New

York, NY), 260, 687–689.

McDonnell, A. V., Jiang, T., Keating, A. E., & Berger, B. (2006). Paircoil2:

Improved prediction of coiled coils from sequence. Bioinformatics

(Oxford, England), 22, 356–358.

McKeon, F. D., Kirschner, M. W., & Caput, D. (1986). Homologies in

both primary and secondary structure between nuclear envelope and

intermediate filament proteins. Nature, 319, 463–468.

McLachlan, A. D. (1978). Coiled coil formation and sequence regularities

in the helical regions of alpha-keratin. Journal of Molecular Biology,

124, 297–304.

Melcer, S., Gruenbaum, Y., & Krohne, G. (2007). Invertebrate lamins.

Experimental Cell Research, 313, 2157–2166.

Mencarelli, C., Ciolfi, S., Caroti, D., Lupetti, P., & Dallai, R. (2011). Isomin:

A novel cytoplasmic intermediate filament protein from an arthropod

species. BMC Biology, 9, 17.

Michie, K. A., & Lowe, J. (2006). Dynamic filaments of the bacterial cyto-

skeleton. Annual Review of Biochemistry, 75, 467–492.

Minguez, A., Franca, S., & Delaespina, S. M. D. (1994). Dinoflagellates

have a eukaryotic nuclear matrix with lamin-like proteins and topo-

isomerase-II. Journal of Cellular Science, 107, 2861–2873.

Miyagishima, S. (2011). Mechanism of plastid division: From a bacterium

to an organelle. Plant Physiology, 155, 1533–1544.

Nakanishi, H., Suzuki, K., Kabeya, Y., & Miyagishima, S. (2009). Plant-

specific protein MCD1 determines the site of chloroplast division

in concert with bacteria-derived MinD. Current Biology, 19, 151–156.

Nelsen, E. M., Williams, N. E., Yi, H., Knaak, J., & Frankel, J. (1994). Fen-

estrin and Conjugation in Tetrahymena thermophila. Journal of Eukary-

otic Microbiology, 41, 483–495.

Neradil, J., & Veselska, R. (2015). Nestin as a marker of cancer stem cells.

Cancer Science, 106, 803–811.

Newman, J. R., Wolf, E., & Kim, P. S. (2000). A computationally directed

screen identifying interacting coiled coils from Saccharomyces

PREISNER ET AL. | 241

cerevisiae. Proceedings of the National Academy of Sciences of the

United States of America, 97, 13203–13208.

Nohria, A., Alonso, R. A., & Peattie, D. A. (1992). Identification and char-

acterization of gamma-giardin and the gamma-giardin gene from Giar-

dia lamblia. Molecular & Biochemical Parasitology, 56, 27–38.

Palm, D., Weiland, M., McArthur, A. G., Winiecka-Krusnell, J., Cipriano,

M. J., Birkeland, S. R., & Svärd, S. (2005). Developmental changes in

the adhesive disk during Giardia differentiation. Molecular & Biochemi-

cal Parasitology, 141, 199–207.

Palm, J. E. D., Weiland, M. E. L., Griffiths, W. J., Ljungstrom, I., & Svard,

S. G. (2003). Identification of immunoreactive proteins during acute

human giardiasis. Journal of Infectious Diseases, 187, 1849–1859.

Paredez, A. R., Assaf, Z. J., Sept, D., Timofejeva, L., Dawson, S. C., Wang,

C.-J. R., & Cande, W. Z. (2011). An actin cytoskeleton with evolutio-

narily conserved functions in the absence of canonical actin-binding

proteins. Proceedings of National Academic Science, 108, 6151–6156.

Parry, D. A. D., & Fraser, R. D. B. (1985). Intermediate filament structure.

1. Analysis of IF protein sequence data. International Journal of Biolog-

ical Macromolecules, 7, 203–213.

Pauling, L., & Corey, R. B. (1953). Compound helical configurations of

polypeptide chains: structure of proteins of the alpha-keratin type.

Nature, 171, 59–61.

Peattie, D. A., Alonso, R. A., Hein, A., & Caulfield, J. P. (1989). Ultrastruc-

tural-localization of giardins to the edges of disk microribbons of

Giarida lamblia and the nucleotide and deduced protein-sequence of

alpha-giardin. Journal of Cell Biology, 109, 2323–2335.

Peter, A., & Stick, R. (2015). Evolutionary aspects in intermediate fila-

ment proteins. Current Opinion in Cell Biology, 32, 48–55.

Pichoff, S., & Lutkenhaus, J. (2001). Escherichia coli division inhibitor

MinCD blocks septation by preventing Z-ring formation. Journal of

Bacteriology, 183, 6630–6635.

Pilhofer, M., & Jensen, G. J. (2013). The bacterial cytoskeleton: More

than twisted filaments. Current Opinion in Cell Biology, 25, 125–133.

Preisner, H., Karin, E. L., Poschmann, G., Stuhler, K., Pupko, T., & Gould,

S. B. (2016). The cytoskeleton of parabasalian parasites comprises

proteins that share properties common to intermediate filament pro-

teins. Protist, 167, 526–543.

Pyke, K. A. (2010). Plastid division. AoB Plant, plq016.

Rose, A., & Meier, I. (2004). Scaffolds, levers, rods and springs: Diverse

cellular functions of long coiled-coil proteins. Cellular & Molecular Life

Sciences, 61, 1996–2009.

Rose, A., Schraegle, S. J., Stahlberg, E. A., & Meier, I. (2005). Coiled-coil

protein composition of 22 proteomes: Differences and common

themes in subcellular infrastructure and traffic control. BMC Evolu-

tionary Biology, 5, 66.

Sch€uler, H., & Matuschewski, K. (2006). Regulation of apicomplexan

microfilament dynamics by a minimal set of actin-binding proteins.

Traffic (Copenhagen, Denmark), 7, 1433–1439.

Shih, Y. L., Le, T., & Rothfield, L. (2003). Division site selection in Esche-

richia coli involves dynamic redistribution of Min proteins within

coiled structures that extend between the two cell poles. Proceedings

of the National Academy of Sciences of the United States of America,

100, 7865–7870.

Shu, D. G., Isozaki, Y., Zhang, X. L., Han, J., & Maruyama, S. (2014). Birth

and early evolution of metazoans. Gondwana Research, 25, 884–895.

Sibbald, S. J., & Archibald, J. M. (2017). More protist genomes needed.

Nature Ecology & Evolution, 1, 145.

Skau, C. T., Courson, D. S., Bestul, A. J., Winkelman, J. D., Rock, R. S.,

Sirotkin, V., & Kovar, D. R. (2011). Actin filament bundling by fimbrin

is important for endocytosis, cytokinesis, and polarization in fission

yeast. Journal of Biological Chemistry, 286, 26964–26977.

Snider, N. T., & Omary, M. B. (2014). Post-translational modifications of

intermediate filament proteins: Mechanisms and functions. Nature

Reviews: Molecular Cell Biology, 15, 163–177.

Sontag, C. A., Sage, H., & Erickson, H. P. (2009). BtubA-BtubB heterodimer is

an essential intermediate in protofilament assembly. PLoS One, 4, e7253.

Specht, M., Schatzle, S., Graumann, P. L., & Waidner, B. (2011). Helico-

bacter pylori possesses four coiled-coil-rich proteins that form

extended filamentous structures and control cell shape and motility.

Journal of Bacteriology, 193, 4523–4530.

Steinert, P. M., Chou, Y. H., Prahlad, V., Parry, D. A. D., Marekov, L. N.,

Wu, K. C., . . . Goldman, R. D. (1999). A high molecular weight inter-

mediate filament-associated protein in BHK-21 cells is nestin, a type

VI intermediate filament protein—Limited co-assembly in vitro toform heteropolymers with type III vimentin and type IV alpha-inter-

nexin. Journal of Biological Chemistry, 274, 9881–9890.

Steinert, P. M., Rice, R. H., Roop, D. R., Trus, B. L., & Steven, A. C.

(1983). Complete amino acid sequence of a mouse epidermal keratin

subunit and implications for the structure of intermediate filaments.

Nature, 302, 794–800.

Steven, A. C., Hainfeld, J. F., Trus, B. L., Wall, J. S., & Steinert, P. M. (1983).

The distribution of mass in heteropolymer intermediate filaments

assembled in vitro. Stem analysis of vimentin/desmin and bovine epider-

mal keratin. Journal of Biological Chemistry, 258, 8323–8329.

Suzuki, N., & Oba, M. (2015). Oldest fossil records of marine protists and

the geologic history toward the establishment of the modern-type

marine protist world. In S.T. Ohtsuka, T. Suzaki, T. Horiguchi, N.

Suzuki, and F. Not (Eds.), Marine protists: Diversity and dynamics (pp.

359–396). Tokyo, Japan: Springer.

Tilney, L. G., & Tilney, M. S. (1996). The cytoskeleton of protozoan para-

sites. Current Opinion in Cell Biology, 8(1), 43–48.

Tinsley, E., & Khan, S. A. (2006). A novel FtsZ-like protein is involved in

replication of the anthrax toxin-encoding pXO1 plasmid in Bacillus

anthracis. Journal of Bacteriology, 188(8), 2829–2835.

Tran, J. Q., de Leon, J. C., Li, C., Huynh, M. H., Beatty, W., & Morrissette,

N. S. (2010). RNG1 is a late marker of the apical polar ring in Toxo-

plasma gondii. Cytoskeleton (Hoboken, NJ), 67(9), 586–598.

Tran, J. Q., Li, C., Chyan, A., Chung, L., & Morrissette, N. S. (2012).

SPM1 stabilizes subpellicular microtubules in Toxoplasma gondii.

Eukaryotic Cell, 11(2), 206–216.

van den Ent, F., Amos, L. A., & Lowe, J. (2001). Prokaryotic origin of the

actin cytoskeleton. Nature, 413(6851), 39–44.

van Hemert, M. J., Lamers, G. E. M., Klein, D. C. G., Oosterkamp, T. H.,

Steensma, H. Y., & van Heusden, G. P. H. (2002). The Saccharomyces

cerevisiae Fin1 protein forms cell cycle-specific filaments between

spindle pole bodies. Proceedings of National Academic Science USA, 99

(8), 5390–5393.

Vaughan, S., Kohl, L., Ngai, I., Wheeler, R. J., & Gull, K. (2008). A repetitive

protein essential for the flagellum attachment zone filament structure

and function in Trypanosoma brucei. Protist, 159(1), 127–136.

Waidner, B., Specht, M., Dempwolff, F., Haeberer, K., Schaetzle, S.,

Speth, V., . . . Graumann, P. L. (2009). A novel system of cytoskeletal

elements in the human pathogen Helicobacter pylori. PLoS Pathogens,

5(11), e1000669.

Wickstead, B., & Gull, K. (2011). The evolution of the cytoskeleton. Jour-

nal of Cell Biology, 194(4), 513–525.

Wickstead, B., Gull, K., & Richards, T. A. (2010). Patterns of kinesin evo-

lution reveal a complex ancestral eukaryote with a multifunctional

cytoskeleton. BMC Evolutionary Biology, 10, 110.

242 | PREISNER ET AL.

Williams, N. E., Honts, J. E., & Jaeckel-Williams, R. F. (1987). Regional

differentiation of the membrane skeleton in Tetrahymena. Journal of

Cellular Science, 87, 457–463.

Wloga, D., Strzyzewska-Jowko, I., Gaertig, J., & Jerka-Dziadosz, M.

(2008). Septins stabilize mitochondria in Tetrahymena thermophila.

Eukaryotic Cell, 7(8), 1373–1386.

Woessner, D. J., & Dawson, S. C. (2012). The Giardia median body

protein is a ventral disc protein that is critical for maintaining a

domed disc conformation during attachment. Eukaryotic Cell, 11(3),

292–301.

Wolf, E., Kim, P. S., & Berger, B. (1997). MultiCoil: A program for predict-

ing two- and three-stranded coiled coils. Protein Science: A Publication

of the Protein Society, 6(6), 1179–1189.

Yang, R., Bartle, S., Otto, R., Stassinopoulos, A., Rogers, M., Plamann, L.,

& Hartzell, P. (2004). AglZ is a filament-forming coiled-coil protein

required for adventurous gliding motility of Myxococcus xanthus. Jour-

nal of Bacteriology, 186(18), 6168–6178.

Zaremba-Niedzwiedzka, K., Caceres, E. F., Saw, J. H., Backstrom, D., Juzo-

kaite, L., Vancaester, E., & Ettema, T. (2017). Asgard archaea illuminate

the origin of eukaryotic cellular complexity. Nature, 541(7637), 353–358.

Zimorski, V., Ku, C., Martin, W. F., & Gould, S. B. (2014). Endosymbiotic

theory for organelle origins. Current Opinion in Microbiology, 22, 38–48.

SUPPORTING INFORMATION

Additional Supporting Information may be found online in the sup-

porting information tab for this article.

How to cite this article: Preisner H, Habicht J, Garg SG, Gould

SB. Intermediate filament protein evolution and protists. Cyto-

skeleton. 2018;75:231–243. https://doi.org/10.1002/cm.21443

PREISNER ET AL. | 243

https://doi.org/10.1002/cm.21443