the ribosome as a hub for protein quality control - mol cell 2013

7/28/2019 The Ribosome as a Hub for Protein Quality Control - Mol Cell 2013

1/11

Molecular Cell

Review

The Ribosome as a Hub for Protein Quality Control

Sebastian Pechmann,1,2 Felix Willmund,1,2 and Judith Frydman1,*1Department of Biology, Stanford University, Stanford, CA 94305-5020, USA2These authors contributed equally to this work*Correspondence: [email protected]://dx.doi.org/10.1016/j.molcel.2013.01.020

Cells face a constant challenge as they produce new proteins. The newly synthesized polypeptides must be

folded properly to avoid aggregation. If proteins do misfold, they must be cleared to maintain a functional and

healthy proteome. Recent work is revealing the complex mechanisms that work cotranslationally to ensure

protein quality control during biogenesis at the ribosome. Indeed, the ribosome is emerging as a central hub

in coordinating these processes, particularly in sensing the nature of the nascent protein chain, recruiting

protein folding and translocation components, and integrating mRNA and nascent chain quality control.

The tiered and complementary nature of these decision-making processes confers robustness and fidelity

to protein homeostasis during protein synthesis.

Introduction

Accurate protein folding is a prerequisite for all cellular

processes, and protein homeostasis disfunction is linked to an

expanding list of human diseases (Powers et al., 2009). This is

challenging for cells because proteins are continuously made

by the approximately 106107 ribosomes in a eukaryotic cell.

Accurately folding newly made polypeptides is a complex task,

which must be achieved in the face of the vectorial nature of

translation, whereby the N-terminal end is available for folding

before the other end has been synthesized, as well as the

crowded cellular environment awaiting the unfolded emerging

chains (Gershenson and Gierasch, 2011; Hartl et al., 2011).

The cell must not only promote accurate folding but also

must prevent the accumulation of misfolded species that may

arise from inefficient folding, errors in translation, and aberrant

mRNAs. Misfolded proteins can overwhelm the quality control

machinery, leading to the accumulation of aggregated species

that are often cytotoxic and linked to severe misfolding

diseases that include Alzheimers and Parkinsons (Chiti and

Dobson, 2006).

To ensure proper folding and quality control of the nascent

proteome, the cell depends on the concerted action of sophisti-

cated pathways to both guide translated proteins into their

functional conformation and prevent the accumulation of defec-

tive polypeptides. Molecular chaperones are central elements

of these quality control systems, as they facilitate protein

biogenesis by assisting polypeptide folding, translocation, andassembly of newly made proteins in the crowded cell (Gershen-

son and Gierasch, 2011; Hartl et al., 2011). The cellular quality

control machinery also responds to errors during protein syn-

thesis by either buffering destabilizing mutations (Fares et al.,

2002; Jarosz et al., 2010) or targeting polypeptides for degrada-

tion (Kaganovich et al., 2008; Tyedmers et al., 2010).

Interestingly, the ribosome itself is emerging as a central actor

in protein folding and quality control processes, acting as a

proofreader during protein biogenesis and initiating downstream

events that regulate the fate of nascent chains. Here, we review

recent insights into the quality control of newly made proteins,

and sketch an emerging integrated picture of tightly coevolved,

dynamic, and hierarchical safety mechanisms that guarantee

high fidelity in nascent chain protein biogenesis.

Biophysical Constraints on Nascent Chain Folding

Emerging from the ribosome, nascent chains have to avoid off-

pathway misfolding and aggregation (Jahn and Radford, 2008).

Because proteins vary substantially in their propensities to fold

and aggregate, the intrinsic properties of the protein sequences

will dramatically influence their ability to fold upon translation.

Studies of the in vitro folding kinetics of small proteins have es-

tablishedthat thefoldingrate increaseswith thenumber of native

contacts in the folded structure (Eaton et al., 2000), as well as

with the number of potential nonnative contacts (Bowman

et al., 2011). These analyses implicate sequence length as the

strongest determinant of folding kinetics for small single-domain

proteins (Ouyang and Liang, 2008). With this in mind, the vecto-

rial nature of polypeptide synthesis at the ribosome can both

assist and complicate successful folding. The number of

possible structural conformations is initially constrained, thus

reducing the chance of misfolding of N-terminal domains

through interaction with C-terminal sequences that have not

yet emerged from the ribosome (Cabrita et al., 2010; Komar,

2009; Zhang and Ignatova, 2011). On the other hand, folding of

long multidomain proteins may require the formation of contacts

distant in polypeptide sequence, which must leave the ribosome

before completing folding (OBrien et al., 2012; Zhang and Igna-

tova, 2011). Early experiments demonstrated that N-terminaldomains can fold cotranslationally, thus enhancing the rate

and yield of folding in vivo compared to the in vitro folding

kinetics (Frydman et al., 1999). The constraints of cotranslational

folding likely influence protein evolution; individual protein

domains are often maintained as independent folding units

(Han et al., 2007), and the overall distribution of their lengths in

proteomes reflects a length limit for folding on a biologically rele-

vant time scale (Lin and Zewail, 2012).

The amino acid sequences of proteins also determine

their intrinsic propensity to aggregate (Goldschmidt et al.,

2010; Sawaya et al., 2007; Tartaglia et al., 2007). Nonspecific

interactions between aggregation-prone and promiscuous

Molecular Cell 49, February 7, 2013 2013 Elsevier Inc. 411
mailto:[email protected]://dx.doi.org/10.1016/j.molcel.2013.01.020http://crossmark.dyndns.org/dialog/?doi=10.1016/j.molcel.2013.01.020&domain=pdfhttp://dx.doi.org/10.1016/j.molcel.2013.01.020mailto:[email protected]


2/11

proteins can be deleterious (Vavouri et al., 2009), and aggrega-

tion is often cytotoxic (Chiti and Dobson, 2006). However, aggre-

gation-prone sequences are also clearly important for protein

structure and function, and are therefore ubiquitously present

in the proteome (Fowler et al., 2007; Goldschmidt et al., 2010).For instance, all cellular signaling pathways involve interactions

between proteins (Levy and Pereira-Leal, 2008), and central

hubs in protein networks often rely on large proteins with

complex folds, exposed hydrophobic, aggregation-prone

surfaces, and long unstructured regions (Tsai et al., 2009). As

discussed below, chaperones are an important cellular response

to the inevitable presence of aggregation-prone sequences in

the proteome.

Cellular Strategies to Optimize De Novo Folding

How are the biophysical constraints on successful cotransla-

tional folding balanced with the cellular need for long, more

complex, and slowly folding proteins? It appears that the chal-

lenge is met through a 2-fold strategy, first through the evolu-tionary optimization of protein sequences to prevent aggregation

and maximize solubility, and second through theaction of a ribo-

some-anchored chaperone machinery.

Several studies suggest that protein sequences have evolved

to minimize the folding burden of the cell. Most proteins are

short, and longer aggregation-prone proteins tend to be on

average less abundant (Tartaglia et al., 2007). In turn, highly ex-

pressed proteins have a lower propensity to aggregate(Tartaglia

et al., 2007) and are generally more conserved (Drummond et al.,

2005); these features help to maintain solubility and reduce the

risk of erroneous mutations that can promote aggregation

(Gsponer and Babu, 2012; Pechmann and Vendruscolo,

2010). Aggregation-prone regions are often flanked by charged

amino acids or proline residues that act as gatekeeper resi-

dues, also to maintain solubility and minimize aggregation

(Buell et al., 2009). Since aggregation can be strongly promoted

by specific interaction of complementary sequence motifs (Sa-

waya et al., 2007), multidomain proteins with homologous

domains generally maintain low sequence identities to prevent

coaggregation of their domains (Wright et al., 2005).

As discussed below, recent studies of cotranslational chap-

erone function suggest a direct relationship between biophysical

constraints on cellular folding and the need for energetically

costly quality control mechanisms (Calloni et al., 2012; Willmund

et al., 2013). Thus, maintaining protein homeostasis relies on

a tiered risk management strategy. The organization and evolu-

tionary dynamics of theproteomeprovide a first line of defense inprotein quality control, where sequences have evolved to avoid

aggregation, and most proteins tend to be short and thus

fold efficiently. This allows the energy-dependent chaperone

systems to preferentially protect long, aggregation-prone but

functionally important proteins.

Role of the mRNA in Translational Fidelity and

Cotranslational Folding

Natively folded proteins are only marginally stable in the cell

(Baldwin et al., 2011; DePristo et al., 2005). The canonical error

rate during protein synthesis results in approximately one per

104 misincorporated amino acids (Zaher and Green, 2009a)

that could tip the balance away from correct folding. Recent

work indicates that the mRNA sequences, and not just the

encoded amino acid sequences, also influence the cellular

synthesis and folding of their corresponding proteins (Figure 1).

Almost all amino acids can be encoded by several synonymous

codons that are under selection for translational speed (dos Reis

et al., 2004) and accuracy (Akashi, 1994). Translationally optimal

codons are recognized by tRNAs with higher availability, and

thus are translated more rapidly; they also confer higher transla-

tional accuracy because their corresponding tRNAs deliver the

encoded amino acid more efficiently to the peptidyltransferase

center (PTC) (Figure 1). Optimal codons have been found to

associate with evolutionarily conserved and thus likely function-

ally important sites in the corresponding proteins (Drummond

and Wilke, 2008). Moreover, optimal codons could be found

preferentially at structurally sensitive and aggregation prone

sites in proteins (Lee et al., 2010), i.e., positions that are particu-

larly susceptible to translation errors disrupting folding and func-

tion. This suggests that selection for optimal codons to enhance

translational fidelity is an important determinant of coding

sequence evolution (Figure 1).

Nonoptimal codons are recognized by lower availability tRNAs

and thus slow down translation elongation; their positioning

along the mRNA coding sequence could in principle allow the

emerging nascent chain more time to begin to fold cotranslation-

ally (Figure 1) (OBrien et al., 2012; Zhang and Ignatova, 2011).Because proteins start folding as they emerge from the ribo-

some, native contacts in secondary or tertiary structures

between distant sites in the polypeptide chain, such as in

b-sheets or between protein domains, can only form once they

have completely left the ribosome exit tunnel (Cabrita et al.,

2010; Fedyukina and Cavagnero, 2011). Indeed, clusters of

nonoptimal codons are maintained under direct selection (Clarke

and Clark, 2008), and their substitution has been found to affect

protein stability (Zhang et al., 2010) and function (Kimchi-Sarfaty

et al., 2007). The role of optimal and nonoptimal codons in co-

translational protein folding is supported by recent findings

that clusters of optimal and nonoptimal codons are evolutionarily

3

5

tRNA(high availability)

mRNA

Coord

inating

foldin

g

Preventingaggregation

Nonoptimal codons

Optimal codons

PTC

tRNA(low availability)

Figure 1. Translational Fidelity and Coordinated CotranslationalFolding at the RibosomeThe ribosomechecksmany aspectsof protein synthesis: proofreading of tRNA

binding and peptide bond formation, and sensing conformationsinsidethe exit

tunnel. Optimal codons recognized by highly available tRNAs are translated

fast and are under selection for higher translational accuracy. They are found

preferentially at sites where high fidelity is important to prevent aggregation.

Nonoptimal codons are recognized by less abundant tRNAs and thus slow

down translation. Clusters of nonoptimal codons are evolutionarily conserved

and enriched in secondary-structure elements that can fold cotranslationally,

suggesting a general role in coordinating cotranslational folding. (Adapted

from Pechmann and Frydman, 2012.)

412 Molecular Cell 49, February 7, 2013 2013 Elsevier Inc.

Molecular Cell

Review


3/11

conserved across eukaryotic organisms in a site-specific

manner (Pechmann and Frydman, 2012). Of note, both optimal

and nonoptimal codons are conserved in equal measure, high-

lighting the importance of maintaining nonoptimal codons for

correct protein biogenesis (Pechmann and Frydman, 2012).

The evolutionarily conserved patterns of optimal and nonoptimal

codons directly correlate with the secondary structure of the en-

coded polypeptides. Structural elements that can fold cotransla-

tionally, such as a helices, contain conserved clusters of nonop-

timal codons, which likely slow down elongation to enhance

cotranslational folding. These findings indicate that the rhythm

of elongation is directly attuned to the needs of cotranslational

folding (Figure 1). An even stronger attenuation of translation is

achieved by specific stalling sequences recognized inside the

ribosome (Tanner et al., 2009). In bacteria, ribosome profiling

found that the anti-Shine-Delgarno sequence, known for its

role in regulating translation initiation, is strategically placedthroughout coding sequences to shape the translation of bacte-

rial genomes (Li et al., 2012). In-depth analysis of conserved

mRNA sequence signatures together with powerful ribosome

profiling experiments are primed to further decipher the layer

of information contained within the coding mRNA sequences

that coordinates protein synthesis and cotranslational folding.

The Ribosome as a Platform for Folding and Quality

Control

The ribosome is emerging as an active and dynamic hub in

protein and mRNA quality control (Kramer et al., 2009; Wilson

and Beckmann, 2011). Low stability of mRNA secondary struc-

tures at the very beginning of coding sequences facilitates the

threading of mRNAs during translation initiation (Gu et al.,

2010), but, if needed, the ribosome itself can act as helicase to

unravel mRNA structures (Takyar et al., 2005). During translation

elongation, the binding energy of cognate tRNAs induces subtleconformational changes in the ribosome itself that facilitate tRNA

recognition and proofreading (Johansson et al., 2012; Ogle and

Ramakrishnan, 2005). An additional quality control mechanism

is implemented after peptidyl transfer and during peptide bond

formation to avoid theincorporation of an amino acid from a non-

cognate tRNA through termination of nascent chain synthesis

(Zaher and Green, 2009b).

All ribosomes have an exit tunnel spanning the distance from

the peptidyl transferase site (PTC) at the subunit interface to the

exitsiteat the other end ofthelarge subunit (Figure2; forreviews,

see Melnikovet al., 2012; Klingeet al., 2012; Schmeing and Ram-

akrishnan, 2009). The nature of the tunnel, and the structural

conformations of proteins and RNAs surrounding the exit site,

differ between eukaryotic and prokaryotic ribosomes (Melnikovet al., 2012; Klinge et al., 2012; Schmeing and Ramakrishnan,

2009). Nascent chains start to fold at the ribosomal exit tunnel

port (Bhushan et al., 2010a; OBrien et al., 2010), and they can

even form helices while still deep inside the ribosome (Lu and

Deutsch, 2005; Woolhead et al., 2004). There is a constriction

site in the exit tunnel, approximately 10 amino acids from the

PTC. In S. cerevisiae, this site consists of the ribosomal proteins

L4, L17, and L39 (Figure 2, in yellow). The constriction site can

sense nascent chain conformations and has been implicated in

communication pathways from the inside to the outside of the

ribosome (Wilson and Beckmann, 2011). The ability of the ribo-

somal tunnel to sense polypeptides during synthesis was initially

identified in bacteria (Gong and Yanofsky, 2002; Nakatogawa

and Ito, 2002). Specific sequence motifs, present, for instance,

in SecM and TnaC, cause the translating polypeptide to stall at

this constriction point in the tunnel in response to SecA or Tryp-

tophan levels, respectively (Cruz-Vera et al., 2006; Nakatogawa

and Ito, 2001). Similar stalling sequences have been identified

in eukaryotes (Bhushan et al., 2010b; Spevak et al., 2010).

However, the constriction site within the ribosomal tunnel is

likely attentive to even more subtle cues. Nascent helix interac-

tions with the tunnel have been shown to induce structural

rearrangements in the ribosome itself, triggering downstream

translocationfactorsthat control subsequenttrafficking (Figure 2)

(Bornemann et al., 2008; Lin et al., 2011). An evolutionarily

conserved regionof lowtranslational efficiency at thevery begin-

ning of mRNA coding sequences helps to prevent ribosometraffic jams during translation elongation (Tulleret al., 2010). Inter-

estingly, this region codes for a protein sequence long enough

to span the distance from the PTC to the constriction site

(Pechmann and Frydman, 2012), perhaps regulating how quickly

and in which conformation the new polypeptide reaches the

constriction site. An important and exciting question for future

research is the extent to which the ribosome tunnel communi-

cates with factorsat the ribosomeexit site,which include nascent

chain-modifying enzymes such as methionyl aminopeptidases

and N-acetyl transferases, molecular chaperones, translocation

factors, and quality control components (Kramer et al., 2009) to

influence nascent chain folding and cellular fate (Figure 2).

40S60S

PTC

L25L31

L39

L17

Ribosome-bound

factors

tRNA

L4

Chaperones

(e.g. CLIPS)

Degradation

(e.g. Ltn1)

Translocation

(e.g. SRP)

Nascent chainmodifyingenzymes (e.g. MAP, NAT)

Figure 2. The Ribosome as Control Center of Nascent Chain Fate

The ribosome exit tunnel interacts with new polypeptides. A constriction site,

in S. cerevisiae comprised of ribosomal proteins L4, L17, and L39, recognizes

nascent chain conformations and communicates both with the peptidyl

transferase site and with the exit site (blue arrows). Ribosome-associated

chaperones and nascentchain-modifying enzymes bind theribosomenear the

tunnel exit. The nascent chain fate can be predetermined from inside the

ribosome. Chaperones promote structural maturation, E3 ligases facilitate

degradation, and the SRP can be recruited to the ribosome before the poly-

peptide has exited.


Molecular Cell

Review


4/11

The Network of Ribosome-Bound Chaperones

Early studies indicated that chaperones cotranslationally bind to

nascent chains as they emerge from the ribosome (Frydman

et al., 1994). The extent of cotranslational chaperone action is

very different in eukaryotic and prokaryotic cells (Figure 3)

(Albane` se et al., 2006). The bacterial trigger factor (TF) is a single

ATP-independent ribosome-bound chaperone, associating with

many nascent chains (Oh et al., 2011; Preissler and Deuerling,

2012). Subsequent de novo folding is mostly posttranslational

(Agashe et al., 2004) and supported by the same ATP-dependent

chaperone systems that protect the proteome from stress.These include the Hsp70 system composed of DnaJ, DnaK,

and GrpE, and the chaperonin system composed of GroEL-

GroES (Figure 3) (Agashe et al., 2004; Hartl et al., 2011). In

eukaryotes, a significant amount of folding occurs cotranslation-

ally (Duncan and Mata, 2011; Frydman et al., 1999; Khushoo

et al., 2011). The eukaryotic chaperone machinery comprises

two independently regulated networks with distinct functions

(Albane` se et al., 2006). A set of chaperones linked to protein

synthesis (CLIPS) associates specifically with ribosomes to facil-

itate de novo folding and is transcriptionally coregulated with the

translation apparatus. In contrast, heat shock chaperones are

induced under stress conditions to protect the proteome and

mediate either refolding or quality control (Albane` se et al.,

2006). CLIPS include structurally and mechanistically diverse

chaperones, such as the ATP-independent nascent polypep-

tide-associated complex (NAC) and GimC/Prefoldin, as well as

ATP-dependent chaperones, including Hsp70 family members

such as SSB in yeast and the eukaryotic chaperonin TRiC/CCT

(Figure 3) (Albane` se et al., 2006). Why eukaryotes require such

a complex folding machinery, and the specificity and interplay

between these different chaperones at the ribosome are ques-

tions that remain to be addressed.

The best-studied ribosome-bound chaperone is the bacterialTF, which provides a paradigm for how chaperones may influ-

ence nascent chains as they exit the ribosome. TF binds

ribosomes close to the tunnel exit site (Ferbitz et al., 2004;

Lakshmipathy et al., 2007). Global profiling of TF-bound ribo-

somes indicated that TF interacts with cytoplasmic and outer

membrane-bound nascent polypeptides (Oh et al., 2011).

Despite its broad specificity, biophysical studies showed that

the residence time of TF on translating ribosomes correlates

with sequence hydrophobicity of the emerging nascent chains

(Kaiser et al., 2006). Nascent polypeptides are accommodated

in the interior cleft of TF where multiple hydrophobic and

hydrophilic binding sites support nascent chain interactions

40S60S

PTC

L31

40S60S

PTC

L25L31

NAC

30S50S

PTC

L23

TF

GroES

GroEL

DnaK/DnaJ

SRP

ER/Membrane/Extracellular

RAC

(Ssz1/Zuo1)Ssb/Hsp70

CCT/TRiC

Hsp70/

Hsp40

Hsp90

GimC/Prefoldin

mature protein

mature protein

mature protein

Prokaryotic

chaperone-network

Eukaryotic

chaperone-network

mature protein

Figure 3. Co- and Posttranslationally Acting Chaperone NetworksProkaryotic and eukaryotic chaperone networks differ in their complexity and division of labor. In prokayotes, trigger factor is the main ribosome-associated

chaperone,foldingnew proteinsor passing themto theHsp70 system of DnaK/DnaJor thechaperonin GroEL/ESto facilitate de novoand stress-induced folding.

In eukaryotes, ribosome-bound chaperones and cochaperones like SSB, RAC, SRP, and NAC compete for overlapping ribosomal binding sites. More diverse

and specialized networks of downstream chaperones split the task of mediating de novo and stress-induced folding. Prefoldin and TRiC are directly coupled toprotein synthesis, while Hsp90 is the primary heat shock chaperone.


Molecular Cell

Review


5/11

(Martinez-Hackert and Hendrickson, 2009). Biochemical experi-

ments indicate that TF binding to nascent chains prevents their

aggregation, protects them from proteolytic cleavage, delays

premature folding of translated nascent polypeptides, and

reduces the formation of early misfolding intermediates ofnascent polypeptide (Agashe et al., 2004; Kaiser et al., 2006;

Hoffmann et al., 2012).

The chaperones in the more complex eukaryotic CLIPS

network show neither sequence nor structural homology to

prokaryotic TF (Kramer et al., 2009; Preissler and Deuerling,

2012). Their increased complexity likely responds to the larger

and more complex nature of eukaryotic proteomes (Albane` se

et al., 2006). Recent work has begun to illuminate mechanistic

and functional aspects of two early-acting CLIPS, NAC and the

Hsp70 SSB. These two chaperones appear to have overlapping

but distinct functions, as highlighted by their strong negative

genetic interactions (Koplin et al., 2010).

The ribosome-associated NAC is a highly conserved dimeric

complex that can be crosslinked to very short nascent chains(Raue et al., 2007). Yeast cells contain a and b isoforms that

can form hetero- and homo-oligomers (Beatrix et al., 2000).

Ribosome binding appears primarily mediated through the

N terminus of b NAC to a site proximal to ribosomal protein

L31 close to the exit tunnel (Pech et al., 2010; Zhang et al.,

2012). While NAC is dispensable for viability in yeast, it is

essential in metazoans (Markesich et al., 2000). The exact

role of NAC is still under debate, but recent studies showed

it is a cotranslational binding factor for most nascent chains

(del Alamo et al., 2011). Interestingly, while NAC interacts

very broadly with translating ribosomes, including those trans-

lating precursors of secretory and mitochondrial proteins, the

different isoforms of NAC interact with different subsets of

nascent chains, suggesting they may differ in their binding

specificity and function (del Alamo et al., 2011). These experi-

ments also began to reveal the complex interplay between

different nascent chain binding factors at the ribosome. Thus,

deletion of NAC causes a relaxation in the specificity of another

cotranslationally acting factor, the signal recognition particle

(SRP), which recognizes nascent chains carrying a signal

sequence for translocation to the endoplasmic reticulum (ER)

(del Alamo et al., 2011). Since NAC and SRP can both bind

to ribosomes carrying ER-bound nascent chains, the relaxation

in specificity likely arises from the interplay of these two factors

at the ribosome. Consistent with this idea, biochemical studies

indicated that the presence of a signal sequence in the nascent

chain is sensed from within the ribosomal exit tunnel to pre-emptively recruit SRP to those ribosomes (Berndt et al.,

2009) in a manner modulated by NAC (Zhang et al., 2012).

Future studies should further determine how NAC modulates

the fate of cytoplasmic proteins, as well as mitochondrial

precursors (George et al., 1998).

Hsp70s were observed to bind cotranslationally to nascent

chains over 20 years ago (Beckmann et al., 1990; Frydman

et al., 1994), but the function of this interaction is only now

beginning to emerge. The cotranslational Hsp70 cycle is best

characterized for the S. cerevisiae Hsp70 SSB, comprising the

functionally redundant Ssb1 and Ssb2 isoforms. SSB associates

with ribosomes and directly binds a large fraction of newly trans-

lated polypeptides (Willmund et al., 2013). ATP hydrolysis

by SSB is stimulated by the ribosome-associated complex

(RAC), which is composed of the atypical Hsp70 Ssz1 and the

J domain protein Zuo1 (Huang et al., 2005). RAC binds directly

to ribosomes but does not interact with nascent chains itself(Yam et al., 2005). Instead, RAC regulates the cotranslational

substrate binding and specificity of SSB (Willmund et al.,

2013). A recent characterization of the interaction of RAC with

ribosomes showed that RAC contacts the ribosome close to

L31, bending in a crouched conformation over the exit tunnel

(Leidig et al., 2013). Structural analysis suggests that the atypical

Hsp70 componentof RAC, Ssz1, is inactive dueto a permanently

locked conformation, which prevents cycling through ATP and

thus substratebinding and release (Leidiget al., 2013). The other

cofactor completing the regulation of the ATPase cycle of SSB is

the nucleotide exchange factor Sse1/Hsp110, a large Hsp70-like

chaperone, which may itself bind and fold substrates (Raviol

et al., 2006; Shaner et al., 2005; Yam et al., 2005). Interestingly,

Sse1, which promotes substrate release from SSB (Yam et al.,2005), associates with SSB off the ribosome (Willmund et al.,

2013), suggesting that the cycle of SSB substrate binding and

ATP hydrolysis is spatially regulated by the differential interaction

between its cofactors and the ribosome. Importantly, RAC and

Sse1 are conserved in mammalian cells, indicating their role is

conserved across eukaryotes (Jaiswal et al., 2011).

A global analysis of the function and specificity of SSB illumi-

nates the role of cotranslational Hsp70 in eukaryotic protein

biogenesis (Willmund et al., 2013). SSB binds preferentially to

a subset of nascent chains encoding nuclear and cytosolic

proteins. Importantly, SSB, unlike NAC, does not bind SRP

substrates, which indicates an early sorting mechanism at the

ribosome (Willmund et al., 2013). Nascent polypeptides that

associate with SSB tend to be longer than those that do not

bind Hsp70; furthermore, they have properties that pose a signif-

icant challenge to cotranslational folding, such as enhanced

aggregation propensity, regions of intrinsic disorder, and

complex domain architectures (Willmund et al., 2013). Interest-

ingly, deletion of SSB, while viable in yeast, causes widespread

aggregation of newly made proteins with exactly those folding-

challenging characteristics. Thus, Hsp70 plays a critical role

in preventing cotranslational misfolding and downstream

aggregation.

These recent studies reveal an intricate network of early-

acting ribosome-bound chaperones choreographing the early

sorting of nascent chains (Figure 3). The generalist NAC both

protects newly made cytosolic proteins and promotes thespecificity of SRP recognition of ER-bound nascent polypep-

tides. Hsp70 in turn appears to more specifically associate

with folding-challenged cytoplasmic and nuclear proteins to

prevent their aggregation and promote folding. Interestingly,

the specificity of Hsp70 is modulated by RAC, whose binding

site on the ribosome is close to that of NAC, thus raising the

question regarding the competition for ribosomal binding sites

of cochaperones. Taken together, the highly dynamic and

partially redundant nature of this network of ribosome-

bound chaperones increases both the plasticity as well as

robustness of the quality control of polypeptides emerging

from the ribosome.


Molecular Cell

Review


6/11

Co- and Posttranslational Folding Pathways

Many cellular proteins require additional chaperone assistance

to fold, beyond the action offered by early-acting ribosome-

bound chaperones. In prokaryotes, the Hsp70/DnaK system

and the GroEL/GroES chaperonin cooperate to promote folding.They interact with substrates primarily posttranslationally,

although DnaK can also associate cotranslationally (Deuerling

et al., 1999; Teter et al., 1999). DnaK and GroEL have distinct

but overlapping substrates (Houry et al., 1999), as do DnaK

and TF(Deuerling et al., 1999; Teter et al., 1999). GroEL is essen-

tial in most prokaryotes and is an obligate chaperone for the

folding of topologically complex and aggregation-prone proteins

(Kerner et al., 2005; Tartaglia et al., 2010). The functional overlap

between TF and DnaK confers robustness to the network,

and deletions of either component are viable (Deuerling et al.,

1999; Teter et al., 1999). However, loss of both TF and DnaK

severely affects growth and disrupts access of newly translated

proteins to GroEL, thus resulting in widespread aggregation

(Calloni et al., 2012).In eukaryotes, several chaperones act co- and posttransla-

tionally to facilitate protein folding and assembly, including the

Gim complex/prefoldin (GimC/PFD), the ring-shaped chapero-

nin TRiC/CCT, the Hsp70 SSA, and Hsp90 and its cofactors

(McClellan et al., 2007; Taipale et al., 2012; Yam et al., 2008).

These chaperones interact with smaller subsets of nascent poly-

peptides compared to ribosome-bound SSB/Hsp70 and NAC

(McClellan et al., 2007; Taipale et al., 2012; Yam et al., 2008).

While the organization and specificity of these downstream

chaperones remain relatively unexplored, it is clear that TRiC/

CCT and Hsp90 have distinct and obligate sets of substrates.

TRiC/CCT associates co- and postranslationally with approxi-

mately 5%10% of newly made proteins (Yam et al., 2008),

predominantly b sheet-rich and topologically complex proteins

that likely require the protected environment of its central cavity

to fold (Douglas et al., 2011). Substrates appear to reach TRiC

with the help of upstream-acting chaperones such as Hsp70

and GimC/PFD (Melville et al., 2003; Vainberg et al., 1998).

Hsp90 is specialized in assisting the structural maturation and

conformational regulation of numerous oligomeric complexes,

structurally labile proteins, as well as signal-transduction com-

ponents (McClellan et al., 2007; Taipale et al., 2012). The soluble

SSA isoforms of Hsp70 are predominantly responsible for cyto-

plasmic folding (Kim et al., 1998; Melville et al., 2003) and assist-

ing translocation to the ER and mitochondria (McClellan and

Brodsky, 2000; Young et al., 2003). This eukaryotic chaperone

network for de novo folding is very robust, as deletions of singlecomponents like GimC/PFD, NAC, or SSB are not lethal (Alba-

ne` se et al., 2006, 2010; Koplin et al., 2010). The network appears

to be organized hierarchically, where Hsp70 and GimC act

upstream of TRiC and Hsp90 (Hartl et al., 2011). This functional

organization of hierarchical chaperone systems and dynamically

regulated cofactors brings increased plasticity while maintaining

specificity in chaperone-mediated folding.

Quality Control of Nascent Polypeptides at the

Ribosome

Newly made polypeptides can be ubiquitinated during or shortly

after synthesis (Sato et al., 1998; Turner and Varshavsky, 2000).

The extent of cotranslational ubiquitination and degradation has

been intensely debated over the last 15 years (Vabulas and Hartl,

2005; Yewdell and Nicchitta, 2006). On the one hand, the mis-

folding potential of nascent, partially unfolded polypeptides

suggests that a significant fraction may be immediately de-graded (Yewdell and Nicchitta, 2006). On the other hand, protein

synthesis is energetically expensive, and the cell devotes many

resources to regulating it. Several lines of evidence suggest

that newly made proteins are largely protected from quality

control (Frydman and Hartl, 1996; Vabulas and Hartl, 2005).

Translation of damaged mRNAs, such as those lacking a stop

codon, have been useful for understanding cotranslational

protein quality control (Ito-Harashima et al., 2007). Translation

of nonstop mRNAs (NS-mRNAs) generates proteins with polyly-

sine tracts coded by the poly(A) tail (Isken and Maquat, 2007;

Klauer and van Hoof, 2012). The polylysine peptides stall the

ribosome, likely by interacting with the negatively charged ribo-

somal tunnel wall. These aberrant complexes are recognized

by a quality control pathway that dissociates the ribosomalsubunits, recruits the exosome to degrade nonstop mRNAs

(Tomecki et al., 2010; van Hoof and Parker, 2002), and targets

the nonstop polypeptides stalled in the 60S subunit for ubiquiti-

nation and degradation (Ito-Harashima et al., 2007). Ubiquitina-

tion of these nascent chain is carried out by the ribosome-asso-

ciated ubiquitin ligase Listerin/Rkr1, (Bengtson and Joazeiro,

2010). Recently, Listerin/Rkr1 has been shown to form a complex

with the AAA+ ATPase Cdc48, Tae2, and Ydr333C, called the

ribosome quality control complex (RQC) (Brandman et al.,

2012). RQC contributes to the dissociation of the ribosomal

subunits, and to coordination of clearance of the aberrant

mRNA and nascent polypeptides (Brandman et al., 2012). Inter-

estingly, RAC and SSB, while not required for polyubiquitination

and degradation of polylysine-containing chains, contribute to

the stabilization of stalled complexes (Chiabudini et al., 2012).

Because this pathway helps ensure faithful polypeptide

synthesis by eliminating aberrant mRNAs, presumably after

a single round of translation, it serves as a safeguard rather

than a general mechanism of nascent chain quality control.

This function is underscored by theexceedinglylow levelsof Lis-

terin in the cell, which is only present at approximately 200

copies per yeast cell. Disfunction of mRNA quality control is

clearly deleterious for protein homeostasis, since Listerin

mutants lead to neurodegeneration (Chu et al., 2009) and the

RQC complex is linked to the signaling of the stress response

by HSF1 (Brandman et al., 2012). Another E3 ligase routinely re-

cruited to the ribosome is Not4 of the Ccr4/NOT complex; thisligase may play a role in either nascent chain ubiquitination or

mRNA quality control (Collart and Panasenko, 2012).

Proteome Homeostasis in Health and Disease

Nascent protein quality control will ultimately have to be under-

stood as a systems-level process encompassing all aspects of

cellular and organismal biology. As quality control capacity is

limited, its overload can lead to proteostatic collapse (Powers

et al., 2009). Excessive aggregation can directly impair protein

quality control, impacting protein function in many cellular

networks (Olzscha et al., 2011). An impairment of proteostatic

capacity is a general hallmark of aging (Morimoto, 2008; Taylor


Molecular Cell

Review


7/11

and Dillin, 2011), often accompanied by widespread aggrega-

tion, particularly of ribosomal proteins (David et al., 2010). The

immediate consequence of failed clearance of aggregated

proteins can be the onset of neurodegenerative protein misfold-

ing diseases that are especially prevalent during aging (Chiti and

Dobson, 2006). In addition, deregulation of chaperone networks

is exploited for uncontrolled cell proliferation in cancers (White-

sell and Lindquist, 2005) and helps drive aneuploidy (Oromendiaet al., 2012). Molecular chaperones also play critical roles in virus

replication (Geller et al., 2012). Advances in our understanding of

individual mechanisms and pathways are prerequisite to piece

together a more nuanced global understanding of protein

homeostasis.

Perspectives

The cell uses diverse, dynamic, and adaptive strategies to

ensure quality and fidelity of new proteins (Figure 4). For

example, genomic sequences have generally evolved to avoid

aggregation, limit misincorporation of critical amino acids, and

promote cotranslational folding. Duringtranslation, the ribosome

actively monitors all aspects of protein biosynthesis, dynamically

responding to errors and influencing the fate of the nascentprotein. The cell manages important evolutionary trade-offs

with its quality control strategies (Wagner, 2008). Stringent

quality control is more energetically expensive, and more static

than less stringent control, complicating adaptation to changing

conditions. Conversely, less-stringent quality control may be

more adaptable, but allows more damaged proteins to reach

thecellular milieu, thus increasing therisk of protein aggregation.

The combination of successive mechanisms provides an elegant

way to simultaneously maximize fidelity, specificity, and plas-

ticity of protein quality control (Figure 4).

Many exciting and pressing questions remain. How does the

ribosome sense the nascent chain inside the ribosome exit

tunnel to influence translocation, degradation, and folding

through the recruitment of ribosome-associated chaperones

and related factors? We are only beginning to understand the

signaling from within the exit tunnel and its consequences on

protein fate. Furthermore, we still dont understand how aspects

of the cellular environment, such as the metabolic state of the

cell, are signaled to folding networks. It would also be interesting

to explore the spatial control of translation and protein quality.

Ribosomes translating proteins that are destined for export

into the ER or membrane are often prelocalized near their func-

tional environment. Currently, it is not clear if and how these

specialized ribosomes are marked and targeted to specific loca-

tions. Finally, it will be useful to further examine the kinetics of

translation elongation and how it might be attuned to the co-

translational folding, as this will be important for understanding

the connections between these processes. It remains to be

seen if translation elongation directly coordinates the cotransla-

tional binding of chaperones. A better understanding of these

questions will open the door to a general integrated framework

of protein folding in the cell. The organization and execution of

the quality control of newly made proteins in the cell is a remark-

able feat waiting to be further uncovered.

ACKNOWLEDGMENTS

We thank the Frydman lab for helpful discussions. We gratefully acknowledge

support from EMBO Long-Term Fellowships to S.P. and F.W., and NIH grants

GM56433 and AI91575 to J.F.

Proteome organization

mRNA sequences &

ribosome proofreading

Ribosome-associatedchaperone network

Downstream quality

control pathways

Protein homeostasis

regulation

40SPTC

60S 3

5

tRNA

40S

60S

Folding

Degradiation

40S

60SPTC

RACSsb

Stressresponse

Enhanced fidelity, specificityand plasticity

SRPNAC

mRNAQC

Figure 4. Sequential Quality Control of Newly Made ProteinsThe cell relies on tiered quality control mechanisms. Most proteins are short

and fold readily, while long and folding-challenged but often functionallyimportant proteinscan relyon chaperone assistance.Individualsequences are

optimized for translational fidelity, thus reducing the risk of phenotypic

missense mutations, as well as to facilitate cotranslational folding. A selective

ribosome-associated chaperone network binds nascent polypeptides to

promote their folding or translocation. Cochaperones add specificity and

plasticity to the selection of substrates and downstream folding/degradation

pathways. Stresses like heat shock can temporarily rebalance the burden of

newly made proteins and chaperone capacity. This sequential organization

achieves high fidelity, specificity, and plasticity.


Molecular Cell

Review


8/11

REFERENCES

Agashe, V.R., Guha, S., Chang, H.-C., Genevaux, P., Hayer-Hartl, M., Stemp,M., Georgopoulos, C., Hartl, F.U., and Barral, J.M. (2004). Function of triggerfactor and DnaK in multidomain protein folding: increase in yield at the

expense of folding speed. Cell 117, 199209.

Akashi, H. (1994). Synonymous codon usage in Drosophila melanogaster:natural selection and translational accuracy. Genetics 136, 927935.

Albane` se,V., Yam,A.Y.-W.,Baughman, J., Parnot, C.,and Frydman,J. (2006).Systems analyses reveal two chaperone networks with distinct functions ineukaryotic cells. Cell 124, 7588.

Albane` se, V., Reissmann, S., and Frydman, J. (2010). A ribosome-anchoredchaperone network that facilitates eukaryotic ribosome biogenesis. J. CellBiol. 189, 6981.

Baldwin, A.J., Knowles, T.P.J., Tartaglia, G.G., Fitzpatrick, A.W., Devlin, G.L.,Shammas, S.L., Waudby, C.A., Mossuto, M.F., Meehan, S., Gras, S.L., et al.(2011). Metastability of native proteins and the phenomenon of amyloidformation. J. Am. Chem. Soc. 133, 1416014163.

Beatrix, B., Sakai,H., andWiedmann, M. (2000). Thealpha andbetasubunitofthe nascent polypeptide-associated complex have distinct functions. J. Biol.

Chem. 275, 3783837845.

Beckmann, R.P., Mizzen, L.E., and Welch, W.J. (1990). Interaction of Hsp 70withnewly synthesized proteins: implications for protein folding and assembly.Science 248, 850854.

Bengtson, M.H., and Joazeiro, C.A.P. (2010). Role of a ribosome-associatedE3 ubiquitin ligase in protein quality control. Nature 467, 470473.

Berndt, U., Oellerer, S., Zhang, Y., Johnson, A.E., and Rospert, S. (2009). Asignal-anchor sequence stimulates signal recognition particle binding to ribo-somes from inside the exit tunnel. Proc. Natl. Acad. Sci. USA106, 13981403.

Bhushan, S., Gartmann, M., Halic, M., Armache, J.-P., Jarasch, A., Mielke, T.,Berninghausen, O., Wilson, D.N., and Beckmann, R. (2010a).a-helical nascentpolypeptide chains visualized within distinct regions of the ribosomal exittunnel. Nat. Struct. Mol. Biol. 17, 313317.

Bhushan, S., Meyer, H., Starosta, A.L., Becker, T., Mielke, T., Berninghausen,

O., Sattler, M., Wilson, D.N., and Beckmann, R. (2010b). Structural basis fortranslational stalling by human cytomegalovirus and fungal arginine attenuatorpeptide. Mol. Cell 40, 138146.

Bornemann, T., Jockel, J., Rodnina, M.V., and Wintermeyer, W. (2008). Signalsequence-independent membrane targeting of ribosomes containing shortnascent peptides within the exit tunnel. Nat. Struct. Mol. Biol. 15, 494499.

Bowman, G.R., Voelz, V.A., and Pande, V.S. (2011). Taming the complexity ofprotein folding. Curr. Opin. Struct. Biol. 21, 411.

Brandman, O., Stewart-Ornstein, J., Wong, D., Larson, A., Williams, C.C., Li,G.-W., Zhou, S., King, D., Shen, P.S., Weibezahn, J., et al. (2012). A ribo-some-boundqualitycontrolcomplextriggers degradation of nascentpeptidesand signals translation stress. Cell 151, 104210154.

Buell, A.K., Tartaglia, G.G., Birkett, N.R., Waudby, C.A., Vendruscolo, M.,Salvatella, X., Welland, M.E., Dobson, C.M., and Knowles, T.P.J. (2009).Position-dependent electrostatic protection against protein aggregation.

ChemBioChem10

, 13091312.

Cabrita, L.D., Dobson, C.M., and Christodoulou, J. (2010). Protein folding onthe ribosome. Curr. Opin. Struct. Biol. 20, 3345.

Calloni, G., Chen, T., Schermann, S.M., Chang, H.-C., Genevaux, P., Agostini,F., Tartaglia, G.G., Hayer-Hartl, M., and Hartl, F.U. (2012). DnaK functions asa central hub in the E. coli chaperone network. Cell Rep. 1, 251264.

Chiabudini, M., Conz, C., Reckmann, F., and Rospert, S. (2012). Ribosome-associated complex and Ssb are required for translational repression inducedby polylysine segments within nascent chains. Mol. Cell. Biol. 32, 47694779.

Chiti, F., and Dobson, C.M. (2006). Protein misfolding, functional amyloid, andhuman disease. Annu. Rev. Biochem. 75, 333366.

Chu,J., Hong,N.A., Masuda,C.A., Jenkins,B.V., Nelms, K.A.,Goodnow,C.C.,Glynne, R.J., Wu, H., Masliah, E., Joazeiro, C.A.P., and Kay, S.A. (2009). A

mouse forward genetics screen identifies LISTERIN as an E3 ubiquitin ligaseinvolved in neurodegeneration. Proc. Natl. Acad. Sci. USA 106, 20972103.

Clarke, T.F., 4th, and Clark, P.L. (2008). Rare codons cluster. PLoS ONE 3,e3412. http://dx.doi.org/10.1371/journal.pone.0003412.

Collart, M.A., and Panasenko, O.O.(2012). The Ccr4notcomplex. Gene492,4253.

Cruz-Vera, L.R., Gong, M., and Yanofsky, C. (2006). Changes produced bybound tryptophan in the ribosome peptidyl transferase center in response toTnaC, a nascent leader peptide. Proc. Natl. Acad. Sci. USA103, 35983603.

David, D.C., Ollikainen, N., Trinidad, J.C., Cary, M.P., Burlingame, A.L., andKenyon, C. (2010). Widespread protein aggregation as an inherent part ofaging inC. elegans.PLoS Biol. 8, e1000450. http://dx.doi.org/10.1371/journal.pbio.1000450.

del Alamo, M., Hogan, D.J., Pechmann, S., Albanese, V., Brown, P.O., andFrydman, J. (2011). Defining the specificity of cotranslationally acting chaper-ones by systematic analysis of mRNAs associated with ribosome-nascentchain complexes. PLoS Biol. 9, e1001100. http://dx.doi.org/10.1371/journal.pbio.1001100.

DePristo, M.A., Weinreich, D.M., and Hartl, D.L.(2005).Missense meanderings

in sequencespace: a biophysical view of protein evolution. Nat.Rev. Genet.6

,678687.

Deuerling, E., Schulze-Specking, A., Tomoyasu, T., Mogk, A., and Bukau, B.(1999). Trigger factor and DnaK cooperate in folding of newly synthesizedproteins. Nature 400, 693696.

dos Reis, M., Savva, R., and Wernisch, L. (2004). Solving the riddle of codonusage preferences: a test for translational selection. Nucleic Acids Res. 32,50365044.

Douglas, N.R., Reissmann, S., Zhang, J., Chen, B., Jakana, J., Kumar, R.,Chiu, W., and Frydman, J. (2011). Dual action of ATP hydrolysis couples lidclosure to substrate release into the group II chaperonin chamber. Cell 144,240252.

Drummond, D.A., and Wilke, C.O. (2008). Mistranslation-induced proteinmisfolding as a dominant constraint on coding-sequence evolution. Cell 134,341352.

Drummond,D.A., Bloom, J.D., Adami, C., Wilke, C.O., and Arnold, F.H.(2005).Why highly expressed proteins evolve slowly. Proc. Natl. Acad. Sci. USA102,1433814343.

Duncan, C.D., and Mata, J. (2011). Widespread cotranslational formation ofprotein complexes. PLoS Genet. 7, e1002398. http://dx.doi.org/10.1371/jour-nal.pgen.1002398.

Eaton, W.A., Munoz, V., Hagen, S.J., Jas, G.S., Lapidus, L.J., Henry, E.R., andHofrichter, J. (2000). Fast kinetics and mechanisms in protein folding. Annu.Rev. Biophys. Biomol. Struct. 29, 327359.

Fares, M.A., Ruiz-Gonzalez, M.X., Moya, A., Elena, S.F., and Barrio, E. (2002).Endosymbiotic bacteria: groEL buffers against deleterious mutations. Nature417, 398.

Fedyukina, D.V., and Cavagnero, S. (2011). Protein folding at the exit tunnel.Annu. Rev. Biophys. 40, 337359.

Ferbitz, L., Maier, T., Patzelt, H., Bukau, B., Deuerling, E., and Ban, N. (2004).Trigger factor in complex with the ribosome forms a molecular cradle fornascent proteins. Nature 431, 590596.

Fowler, D.M., Koulov, A.V., Balch, W.E., and Kelly, J.W. (2007). Functionalamyloidfrom bacteria to humans. Trends Biochem. Sci. 32, 217224.

Frydman, J., and Hartl, F.U. (1996). Principles of chaperone-assisted proteinfolding: differences between in vitro and in vivo mechanisms. Science 272,14971502.

Frydman, J., Nimmesgern, E., Ohtsuka, K., and Hartl, F.U. (1994). Folding ofnascent polypeptide chains in a highmolecular mass assembly withmolecularchaperones. Nature 370, 111117.

Frydman, J., Erdjument-Bromage, H., Tempst, P., and Hartl, F.U. (1999).Co-translational domain folding as the structural basis for the rapid de novofolding of firefly luciferase. Nat. Struct. Biol. 6, 697705.


Molecular Cell

Review


9/11

Geller, R., Taguwa, S., and Frydman, J. (2012). Broad action of Hsp90 as ahost chaperone required for viral replication. Biochim. Biophys. Acta 1823,698706.

George, R., Beddoe, T., Landl, K., and Lithgow, T. (1998). The yeast nascentpolypeptide-associated complex initiates protein targeting to mitochondria

in vivo. Proc. Natl. Acad. Sci. USA 95, 22962301.

Gershenson, A., and Gierasch, L.M. (2011). Protein folding in the cell: chal-lenges and progress. Curr. Opin. Struct. Biol. 21, 3241.

Goldschmidt, L., Teng, P.K., Riek, R., and Eisenberg, D. (2010). Identifying theamylome, proteins capable of forming amyloid-like fibrils. Proc. Natl. Acad.Sci. USA107, 34873492.

Gong, F., and Yanofsky, C. (2002). Instruction of translating ribosome bynascent peptide. Science 297, 18641867.

Gsponer, J., and Babu, M.M. (2012). Cellular strategies for regulating func-tional and nonfunctional protein aggregation. Cell Rep. 2, 14251437.

Gu, W., Zhou, T., and Wilke, C.O. (2010). A universal trend of reduced mRNAstability nearthe translation-initiationsite in prokaryotesand eukaryotes. PLoSComput. Biol. 6, e1000664. http://dx.doi.org/10.1371/journal.pcbi.1000664.

Han, J.-H., Batey, S., Nickson, A.A., Teichmann, S.A., and Clarke, J. (2007).The folding and evolution of multidomain proteins. Nat. Rev. Mol. Cell Biol.8, 319330.

Hartl, F.U., Bracher, A., and Hayer-Hartl, M. (2011). Molecular chaperones inprotein folding and proteostasis. Nature 475, 324332.

Hoffmann, A., Becker, A.H., Zachmann-Brand, B., Deuerling, E., Bukau, B.,and Kramer, G. (2012). Concerted action of the ribosome and the associatedchaperone trigger factor confines nascent polypeptide folding. Mol. Cell 48,6374.

Houry, W.A., Frishman, D., Eckerskorn, C., Lottspeich, F., and Hartl, F.U.(1999). Identification of in vivo substrates of the chaperonin GroEL. Nature402, 147154.

Huang, P., Gautschi, M., Walter, W., Rospert, S., and Craig, E.A. (2005). TheHsp70 Ssz1 modulates the function of the ribosome-associated J-proteinZuo1. Nat. Struct. Mol. Biol. 12, 497504.

Isken, O., and Maquat, L.E. (2007). Quality control of eukaryotic mRNA: safe-guarding cells from abnormal mRNA function. Genes Dev. 21, 18331856.

Ito-Harashima, S., Kuroha, K., Tatematsu, T., and Inada, T. (2007). Translationof the poly(A) tail plays crucial roles in nonstop mRNA surveillance via transla-tionrepressionand protein destabilization by proteasome in yeast.Genes Dev.21, 519524.

Jahn, T.R., and Radford, S.E. (2008). Folding versus aggregation: polypep-tide conformations on competing pathways. Arch. Biochem. Biophys. 469,100117.

Jaiswal,H., Conz, C.,Otto, H.,Wolfle,T., Fitzke, E.,Mayer, M.P.,and Rospert,S. (2011). The chaperone network connected to human ribosome-associatedcomplex. Mol. Cell. Biol. 31, 11601173.

Jarosz, D.F., Taipale,M., andLindquist, S. (2010). Protein homeostasisand thephenotypic manifestation of genetic diversity: principles and mechanisms.

Annu. Rev. Genet. 44, 189216.

Johansson, M., Zhang, J., and Ehrenberg, M. (2012). Genetic code translationdisplays a linear trade-off between efficiency and accuracy of tRNA selection.Proc. Natl. Acad. Sci. USA109, 131136.

Kaganovich, D., Kopito, R., and Frydman, J. (2008). Misfolded proteinspartition between two distinct quality control compartments. Nature 454,10881095.

Kaiser, C.M., Chang, H.-C., Agashe, V.R., Lakshmipathy, S.K., Etchells, S.A.,Hayer-Hartl, M., Hartl, F.U., and Barral, J.M. (2006). Real-time observation oftrigger factor function on translating ribosomes. Nature 444, 455460.

Kerner, M.J., Naylor, D.J., Ishihama, Y., Maier, T., Chang, H.-C., Stines, A.P.,Georgopoulos, C., Frishman, D., Hayer-Hartl, M., Mann, M., and Hartl, F.U.(2005). Proteome-wide analysis of chaperonin-dependent protein folding inEscherichia coli. Cell 122, 209220.

Khushoo, A., Yang, Z., Johnson, A.E., and Skach, W.R. (2011). Ligand-driven vectorial folding of ribosome-bound human CFTR NBD1. Mol. Cell 41,682692.

Kim, S., Schilke, B., Craig, E.A., and Horwich, A.L. (1998). Folding in vivo ofa newly translated yeast cytosolic enzyme is mediated by the SSA class of

cytosolic yeast Hsp70 proteins. Proc. Natl. Acad. Sci. USA 95, 1286012865.

Kimchi-Sarfaty, C., Oh, J.M., Kim,I.-W., Sauna, Z.E., Calcagno,A.M., Ambud-kar, S.V., and Gottesman, M.M. (2007). A silent polymorphism in the MDR1gene changes substrate specificity. Science 315, 525528.

Klauer, A.A., and van Hoof, A. (2012). Degradation of mRNAs that lack a stopcodon: a decade of nonstop progress. Wiley Interdiscip.Rev. RNA3, 649660.

Klinge, S., Voigts-Hoffmann, F., Leibundgut, M., and Ban, N. (2012). Atomicstructures of the eukaryotic ribosome. Trends Biochem. Sci. 37, 189198.

Komar, A.A. (2009). A pause for thought along the co-translational foldingpathway. Trends Biochem. Sci. 34, 1624.

Koplin, A., Preissler, S., Ilina, Y., Koch, M., Scior, A., Erhardt, M., and Deuerl-ing, E. (2010). A dual function for chaperones SSB-RAC and the NAC nascentpolypeptide-associated complex on ribosomes. J. Cell Biol. 189, 5768.

Kramer, G., Boehringer, D., Ban, N., and Bukau, B. (2009). The ribosome asa platform for co-translational processing, folding and targeting of newlysynthesized proteins. Nat. Struct. Mol. Biol. 16, 589597.

Lakshmipathy, S.K., Tomic, S., Kaiser, C.M., Chang, H.-C., Genevaux, P.,Georgopoulos, C., Barral, J.M., Johnson, A.E., Hartl, F.U., and Etchells, S.A.(2007). Identification of nascent chain interaction sites on trigger factor. J.Biol. Chem. 282, 1218612193.

Lee, Y., Zhou, T., Tartaglia, G.G., Vendruscolo, M., and Wilke, C.O. (2010).Translationally optimal codons associate with aggregation-prone sites inproteins. Proteomics 10, 41634171.

Leidig, C., Bange, G., Kopp, J., Amlacher, S., Aravind, A., Wickles, S., Witte,G., Hurt, E., Beckmann, R., and Sinning, I. (2013). Structural characterizationof a eukaryotic chaperone-the ribosome-associated complex. Nat. Struct.Mol. Biol. 20, 2328.

Levy, E.D., and Pereira-Leal, J.B. (2008). Evolution and dynamics of protein

interactions and networks. Curr. Opin. Struct. Biol. 18, 349357.

Li, G.-W., Oh, E., and Weissman, J.S. (2012). The anti-Shine-Dalgarnosequence drives translational pausing and codon choice in bacteria. Nature484, 538541.

Lin, M.M., and Zewail, A.H. (2012). Hydrophobic forces and the length limit offoldable protein domains. Proc. Natl. Acad. Sci. USA109, 98519856.

Lin, P.-J., Jongsma, C.G., Pool, M.R., and Johnson, A.E. (2011). Polytopicmembrane protein folding at L17 in the ribosome tunnel initiates cyclicalchanges at the translocon. J. Cell Biol. 195, 5570.

Lu, J., and Deutsch, C. (2005). Folding zones inside the ribosomal exit tunnel.Nat. Struct. Mol. Biol. 12, 11231129.

Markesich, D.C., Gajewski, K.M., Nazimiec, M.E., and Beckingham, K. (2000).Bicaudal encodes the Drosophila beta NAC homolog, a component of theribosomal translational machinery. Development 127, 559572.

Martinez-Hackert, E., and Hendrickson, W.A. (2009). Promiscuous substraterecognition in folding and assembly activities of the trigger factor chaperone.Cell 138, 923934.

McClellan, A.J., and Brodsky, J.L. (2000). Mutation of the ATP-binding pocketof SSA1 indicates that a functional interaction between Ssa1p and Ydj1p isrequired for post-translational translocation into the yeast endoplasmic retic-ulum. Genetics 156, 501512.

McClellan, A.J., Xia, Y., Deutschbauer, A.M., Davis, R.W., Gerstein, M., andFrydman, J. (2007). Diverse cellular functions of the Hsp90 molecularchaperone uncovered using systems approaches. Cell 131, 121135.

Melnikov, S., Ben-Shem, A., Garreau de Loubresse, N., Jenner, L., Yusupova,G., and Yusupov, M. (2012). One core, two shells: bacterial and eukaryoticribosomes. Nat. Struct. Mol. Biol. 19, 560567.


Molecular Cell

Review


10/11

Melville, M.W., McClellan, A.J., Meyer, A.S., Darveau, A., and Frydman, J.(2003). The Hsp70 and TRiC/CCT chaperone systems cooperate in vivo toassemble the von Hippel-Lindau tumor suppressor complex. Mol. Cell. Biol.23, 31413151.

Morimoto, R.I. (2008). Proteotoxic stress and inducible chaperone networks in

neurodegenerative disease and aging. Genes Dev. 22, 14271438.

Nakatogawa, H., and Ito, K. (2001). Secretion monitor, SecM, undergoes self-translation arrest in the cytosol. Mol. Cell 7, 185192.

Nakatogawa, H., and Ito, K. (2002). The ribosomal exit tunnel functions asa discriminating gate. Cell 108, 629636.

OBrien, E.P., Hsu, S.-T.D., Christodoulou, J., Vendruscolo, M., and Dobson,C.M. (2010). Transient tertiary structure formation within the ribosome exitport. J. Am. Chem. Soc. 132, 1692816937.

OBrien, E.P., Vendruscolo, M., and Dobson, C.M. (2012). Prediction of vari-able translation rate effects on cotranslational protein folding. Nat. Commun.3, 868.

Ogle, J.M., and Ramakrishnan, V. (2005). Structural insights into translationalfidelity. Annu. Rev. Biochem. 74, 129177.

Oh, E., Becker, A.H., Sandikci, A., Huber, D., Chaba, R., Gloge, F., Nichols,R.J., Typas, A., Gross, C.A., Kramer, G., et al. (2011). Selective ribosomeprofiling reveals the cotranslational chaperone action of trigger factor in vivo.Cell 147, 12951308.

Olzscha, H., Schermann, S.M., Woerner, A.C., Pinkert, S., Hecht, M.H., Tarta-glia, G.G., Vendruscolo, M., Hayer-Hartl, M., Hartl, F.U., and Vabulas, R.M.(2011). Amyloid-like aggregates sequester numerous metastable proteinswith essential cellular functions. Cell 144, 6778.

Oromendia, A.B., Dodgson, S.E., and Amon, A. (2012). Aneuploidy causesproteotoxic stress in yeast. Genes Dev. 26, 26962708.

Ouyang, Z., and Liang, J. (2008). Predicting protein folding rates fromgeometric contact and amino acid sequence. Protein Sci. 17, 12561263.

Pech, M.,Spreter, T., Beckmann, R.,and Beatrix, B. (2010). Dualbindingmodeof the nascent polypeptide-associated complex reveals a novel universaladapter site on the ribosome. J. Biol. Chem. 285, 1967919687.

Pechmann, S., and Frydman, J. (2012). Evolutionary conservation of codonoptimality reveals hidden signatures of cotranslational folding. Nat. Struct.Mol. Biol. 20, 237243.

Pechmann, S., and Vendruscolo, M. (2010). Derivation of a solubility conditionfor proteins from an analysis of the competition between folding and aggrega-tion. Mol. Biosyst. 6, 24902497.

Powers, E.T., Morimoto, R.I., Dillin, A., Kelly, J.W., and Balch, W.E. (2009).Biological and chemical approaches to diseases of proteostasis deficiency.

Annu. Rev. Biochem. 78, 959991.

Preissler, S., and Deuerling, E. (2012). Ribosome-associated chaperones askey players in proteostasis. Trends Biochem. Sci. 37, 274283.

Raue, U., Oellerer, S., and Rospert, S. (2007). Association of protein biogen-esis factors at the yeast ribosomal tunnel exit is affected by the translationalstatus and nascent polypeptide sequence. J. Biol. Chem. 282, 78097816.

Raviol, H., Sadlish, H., Rodriguez, F., Mayer, M.P., and Bukau, B. (2006).Chaperone network in the yeast cytosol: Hsp110 is revealed as an Hsp70nucleotide exchange factor. EMBO J. 25, 25102518.

Sato, S., Ward, C.L., and Kopito, R.R. (1998). Cotranslational ubiquitination ofcystic fibrosis transmembrane conductance regulator in vitro. J. Biol. Chem.273, 71897192.

Sawaya, M.R., Sambashivan, S., Nelson, R., Ivanova, M.I., Sievers, S.A.,Apostol, M.I., Thompson, M.J., Balbirnie, M., Wiltzius, J.J.W., McFarlane,H.T., et al.(2007). Atomic structures of amyloid cross-beta spines reveal variedsteric zippers. Nature 447, 453457.

Schmeing, T.M., and Ramakrishnan, V. (2009). What recent ribosomestructures have revealed about the mechanism of translation. Nature 461,12341242.

Shaner, L., Wegele, H., Buchner, J., and Morano, K.A. (2005). The yeastHsp110 Sse1 functionally interacts with the Hsp70 chaperones Ssa andSsb. J. Biol. Chem. 280, 4126241269.

Spevak, C.C., Ivanov, I.P., and Sachs, M.S.(2010).Sequencerequirementsforribosome stalling by the arginine attenuator peptide. J. Biol. Chem. 285,

4093340942.

Taipale, M., Krykbaeva, I., Koeva, M., Kayatekin, C., Westover, K.D., Karras,G.I., and Lindquist, S. (2012). Quantitative analysis of HSP90-client interac-tions reveals principles of substrate recognition. Cell 150, 9871001.

Takyar, S., Hickerson, R.P., and Noller, H.F. (2005). mRNA helicase activity ofthe ribosome. Cell 120, 4958.

Tanner, D.R., Cariello, D.A., Woolstenhulme, C.J., Broadbent, M.A., andBuskirk, A.R. (2009). Genetic identification of nascent peptides that induceribosome stalling. J. Biol. Chem. 284, 3480934818.

Tartaglia, G.G.,Pechmann, S., Dobson, C.M., and Vendruscolo,M. (2007). Lifeon the edge: a link between gene expression levels and aggregation rates ofhuman proteins. Trends Biochem. Sci. 32, 204206.

Tartaglia, G.G., Dobson, C.M., Hartl, F.U., and Vendruscolo, M. (2010).Physicochemical determinants of chaperone requirements. J. Mol. Biol. 400,

579588.

Taylor, R.C., and Dillin, A. (2011). Aging as an event of proteostasis collapse.Cold Spring Harb. Perspect. Biol. 3, a004440. http://dx.doi.org/10.1101/cshperspect.a004440.

Teter, S.A., Houry, W.A., Ang, D., Tradler, T., Rockabrand, D., Fischer, G.,Blum, P., Georgopoulos, C., and Hartl, F.U. (1999). Polypeptide flux throughbacterial Hsp70: DnaK cooperates with trigger factor in chaperoning nascentchains. Cell 97, 755765.

Tomecki, R., Drazkowska, K., and Dziembowski, A. (2010). Mechanisms ofRNA degradation by the eukaryotic exosome. ChemBioChem 11, 938945.

Tsai, C.-J., Ma, B., and Nussinov, R. (2009). Protein-protein interactionnetworks: how can a hub protein bind so many different partners? Trends Bio-chem. Sci. 34, 594600.

Tuller, T., Carmi, A., Vestsigian,K., Navon, S., Dorfan, Y., Zaborske, J.,Pan, T.,

Dahan, O., Furman, I., and Pilpel, Y. (2010). An evolutionarily conserved mech-anism for controlling the efficiency of protein translation. Cell 141, 344354.

Turner, G.C., and Varshavsky, A. (2000). Detecting and measuring cotransla-tional protein degradation in vivo. Science 289, 21172120.

Tyedmers,J., Mogk, A.,and Bukau, B. (2010). Cellularstrategiesfor controllingprotein aggregation. Nat. Rev. Mol. Cell Biol. 11, 777788.

Vabulas, R.M., and Hartl, F.U. (2005). Protein synthesis upon acute nutrientrestriction relies on proteasome function. Science 310, 19601963.

Vainberg, I.E., Lewis, S.A., Rommelaere, H., Ampe, C., Vandekerckhove, J.,Klein, H.L., and Cowan, N.J. (1998). Prefoldin, a chaperone that deliversunfolded proteins to cytosolic chaperonin. Cell 93, 863873.

vanHoof, A., and Parker, R. (2002). Messenger RNA degradation: beginningatthe end. Curr. Biol. 12, R285R287.

Vavouri, T., Semple, J.I., Garcia-Verdugo, R., and Lehner, B. (2009). Intrinsicprotein disorder and interaction promiscuity are widely associated withdosage sensitivity. Cell 138, 198208.

Wagner, A. (2008). Robustness and evolvability: a paradox resolved. Proc.Biol. Sci. 275, 91100.

Whitesell, L., andLindquist, S.L.(2005).HSP90 andthe chaperoning of cancer.Nat. Rev. Cancer 5, 761772.

Willmund, F., Del Alamo, M., Pechmann, S., Chen, T., Albane` se, V., Dammer,E.B., Peng, J., and Frydman, J. (2013). The cotranslational function of ribo-some-associatedHsp70 in eukaryotic proteinhomeostasis.Cell152, 196209.

Wilson, D.N., and Beckmann, R. (2011). The ribosomal tunnel as a functionalenvironment for nascent polypeptide folding and translational stalling. Curr.Opin. Struct. Biol. 21, 274282.


Molecular Cell

Review


11/11

Woolhead, C.A., McCormick, P.J., and Johnson, A.E. (2004). Nascentmembrane and secretory proteins differ in FRET-detected folding far insidethe ribosome and in their exposure to ribosomal proteins. Cell 116, 725736.

Wright, C.F., Teichmann, S.A.,Clarke, J., and Dobson, C.M.(2005).The impor-

tance of sequence diversity in the aggregation and evolution of proteins.Nature 438, 878881.

Yam, A.Y.-W., Albane` se, V., Lin, H.-T.J., and Frydman, J. (2005). Hsp110cooperates with different cytosolic HSP70 systems in a pathway for de novofolding. J. Biol. Chem. 280, 4125241261.

Yam, A.Y., Xia, Y., Lin, H.-T.J., Burlingame, A., Gerstein, M., and Frydman, J.(2008). Defining the TRiC/CCT interactome links chaperonin function tostabilization of newly made proteins with complex topologies. Nat. Struct.Mol. Biol. 15, 12551262.

Yewdell, J.W., and Nicchitta, C.V. (2006). The DRiP hypothesis decennial:support, controversy, refinement and extension. Trends Immunol. 27,368373.

Young, J.C., Hoogenraad, N.J., and Hartl, F.U. (2003). Molecular chaperonesHsp90 and Hsp70 deliver preproteins to the mitochondrial import receptorTom70. Cell 112, 4150.

Zaher, H.S.,and Green, R. (2009a).Fidelityat themolecular level: lessons fromprotein synthesis. Cell 136, 746762.

Zaher, H.S., and Green, R. (2009b). Quality control by the ribosome followingpeptide bond formation. Nature 457, 161166.

Zhang, G., and Ignatova, Z. (2011). Folding at the birth of the nascent chain:coordinating translation with co-translational folding. Curr. Opin. Struct. Biol.21, 2531.

Zhang, F., Saha, S., Shabalina, S.A., and Kashina, A. (2010). Differentialarginylation of actin isoforms is regulated by coding sequence-dependentdegradation. Science 329, 15341537.

Zhang, Y., Berndt, U., Golz, H., Tais, A., Oellerer, S., Wolfle, T., Fitzke, E., andRospert, S. (2012). NAC functions as a modulator of SRP during the earlysteps of protein targeting to the endoplasmic reticulum. Mol. Biol. Cell 23,30273040.

Molecular Cell 49 February 7 2013 2013 Elsevier Inc 421

Molecular Cell

Review

the ribosome as a hub for protein quality control - mol cell 2013

Documents