getting organized – how bacterial cells move proteins and dna anna buch 25.01.2010 martin...

Getting organized –how bacterial cells move

proteins and DNA

Anna Buch25.01.2010

Martin Thanbichler and Lucy Shapiro

Nature Reviews, 2008

Model systems for bacterial cell biology

Box 1

• E. coli: history, genetic tools, physiology

• B. subtilis: cell differentiation, large size

• C. crescentus: cell division, synchonizable

mobile

sessile

• Diffusion and capture

Assembly of stationary protein complexes

SpoIVB

SpoIIQ

Figure 1

Mother cell

Phagocytosis-likeuptake

Septal membrane

• Targeted membrane insertion

Assembly of stationary protein complexes

SpoIVB

SpoIIIAH

SpoIIQ

Figure 1

• Targeted membrane insertion

Assembly of stationary protein complexes

Steinhauer et al., Mol Microbiol. 1999 32:367-77.; Pollard & Cooper, Science 2009 326:1208-12

IcsA: outer membrane protein, N-term is exposed to host cytoplasm

IcsP: Protease that cleaves off IcsA

Shigella flexneri:facultative intracellular pathogen

Dynamic protein scaffolds and cell shape:Bacterial actin-like cytoskeleton

Figure 2

Bundles of two or more protofilaments.

MreB dynamics in C. crescentus

Figure 2

MreB cables

Spiral like during growth

Ring-like during cell division

Architecture of MreB cables

Figure 2; Carballido-Lopez & Errington, Dev Cell. 2003 4:19-28.

B. subtilis, FRAP of GFP-Mbl

Regulation of cell-wall biosynthesis

Carballido-Lopez et al., Dev Cell. 2006 11:399-409

MreB homologues: MreBH and MblLytE: peptidoglycan hydrolase

Peptidoglycan (PG)synthetic machinery

PG-hydrolase subunit

CW binding subdomain

B. subtilis

Role of MreC in bacterial morphogenesis

Divakaruni et al., PNAS 2005 102:18602-7

C. Crescentus

PBC (penicillin-binding protein):

involved in peptidoglykan synthesis

MreC

DAPI

Crescentin

Ausmees et al., Cell. 2003 115:705-13.

C. crescentus:

creS::Tn5 -> no crescentin

creS::Tn5 + creS ->crescentin on plasmid

In-vitro assay

His-CreS filaments,

EM negative stain

Plasmid segregation

• Actin superfamiliy member (type II partitioning system)

• Walker ATPase (type I partitioning system)

• Tubulin homologue

Plasmid segregation by actin-like proteins

Figure 3

Plasmid R1 of E. coli

Plasmid segregation by Walker-type ATPases

• Walker A cytoskeletal ATPase (WACA)

Adapted from Lim et al., PNAS 2005 102:17658-63

Plasmids F and pB171 of E. coli

TubZ: B. thuringiensis serovar israelensis (pBtoxis)

Plasmid segregation by a tubulin homologue

Larsen et al., Genes Dev. 2007 21:1340-52

E.coli, expressing TubZ-GFP, FRAP, time in sec

Model proposes treadmilling

Arrangement of chromosomal DNA

Figure 4, Viollier et al., PNAS 2004 101:9257-62

Divisome: Bacterial cell-division apparatus

Allard & Cytrynbaum PNAS 2009 106:145-50; Erickson, PNAS 2009 106:9238-43

Z-ring: FitsZ filaments

Rod-shaped bacterium (e.g. E. coli)

Division-site placement: The Min system

Figure 5

minCDE operon:

MinD: WACA family

MinCD-complex: inhibit FtsZ-ring formation

MinE: represses MinCD activity

“Fail-safe mechanism”: nucleoid occlusion

B. subtilis: Noc

E. coli: SlmA

Division-site placement: The MipZ system

MipZ: ATPase, inhibits FtsZ-polymerization

ParB: chromosome partitioning protein

parS: cluster of sites, 15 kb away from ori

Conclusions

• Tubulin filaments:– cell-division apparatus, plasmid segregation

• Actin cables:– DNA partitioning, cell-shape determination,

protein localization

• WACA ATPases:– DNA segregation, cell-division plane

Outlook

• Positioning of proteins at cell poles– TipN– Peptidoglycans– Cardiolipin -> ProP

• Biochemical assembly mechanisms– Actin homologues– Tubulin homologues– WACA ATPases

Thank you for your attention!