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A dynamically assembled cell wall synthesis machinery buffers cell growth.

ABSTRACT: Assembly of protein complexes is a key mechanism for achieving spatial and temporal coordination in processes involving many enzymes. Growth of rod-shaped bacteria is a well-studied example requiring such coordination; expansion of the cell wall is thought to involve coordination of the activity of synthetic enzymes with the cytoskeleton via a stable complex. Here, we use single-molecule tracking to demonstrate that the bacterial actin homolog MreB and the essential cell wall enzyme PBP2 move on timescales orders of magnitude apart, with drastically different characteristic motions. Our observations suggest that PBP2 interacts with the rest of the synthesis machinery through a dynamic cycle of transient association. Consistent with this model, growth is robust to large fluctuations in PBP2 abundance. In contrast to stable complex formation, dynamic association of PBP2 is less dependent on the function of other components of the synthesis machinery, and buffers spatially distributed growth against fluctuations in pathway component concentrations and the presence of defective components. Dynamic association could generally represent an efficient strategy for spatiotemporal coordination of protein activities, especially when excess concentrations of system components are inhibitory to the overall process or deleterious to the cell.

SUBMITTER: Lee TK

PROVIDER: S-EPMC3970539 | biostudies-other | 2014 Mar

REPOSITORIES: biostudies-other

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A dynamically assembled cell wall synthesis machinery buffers cell growth.

Lee Timothy K TK Tropini Carolina C Hsin Jen J Desmarais Samantha M SM Ursell Tristan S TS Gong Enhao E Gitai Zemer Z Monds Russell D RD Huang Kerwyn Casey KC

Proceedings of the National Academy of Sciences of the United States of America 20140218 12

Assembly of protein complexes is a key mechanism for achieving spatial and temporal coordination in processes involving many enzymes. Growth of rod-shaped bacteria is a well-studied example requiring such coordination; expansion of the cell wall is thought to involve coordination of the activity of synthetic enzymes with the cytoskeleton via a stable complex. Here, we use single-molecule tracking to demonstrate that the bacterial actin homolog MreB and the essential cell wall enzyme PBP2 move on ...[more]

PMID: 24550500

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Project description:UnlabelledThe synthesis of peptidoglycan (PG) in bacteria is a crucial process controlling cell shape and vitality. In contrast to bacteria such as Escherichia coli that grow by dispersed lateral insertion of PG, little is known of the processes that direct polar PG synthesis in other bacteria such as the Rhizobiales. To better understand polar growth in the Rhizobiales Agrobacterium tumefaciens, we first surveyed its genome to identify homologs of (~70) well-known PG synthesis components. Since most of the canonical cell elongation components are absent from A. tumefaciens, we made fluorescent protein fusions to other putative PG synthesis components to assay their subcellular localization patterns. The cell division scaffolds FtsZ and FtsA, PBP1a, and a Rhizobiales- and Rhodobacterales-specific l,d-transpeptidase (LDT) all associate with the elongating cell pole. All four proteins also localize to the septum during cell division. Examination of the dimensions of growing cells revealed that new cell compartments gradually increase in width as they grow in length. This increase in cell width is coincident with an expanded region of LDT-mediated PG synthesis activity, as measured directly through incorporation of exogenous d-amino acids. Thus, unipolar growth in the Rhizobiales is surprisingly dynamic and represents a significant departure from the canonical growth mechanism of E. coli and other well-studied bacilli.ImportanceMany rod-shaped bacteria, including pathogens such as Brucella and Mycobacteriu, grow by adding new material to their cell poles, and yet the proteins and mechanisms contributing to this process are not yet well defined. The polarly growing plant pathogen Agrobacterium tumefaciens was used as a model bacterium to explore these polar growth mechanisms. The results obtained indicate that polar growth in this organism is facilitated by repurposed cell division components and an otherwise obscure class of alternative peptidoglycan transpeptidases (l,d-transpeptidases). This growth results in dynamically changing cell widths as the poles expand to maturity and contrasts with the tightly regulated cell widths characteristic of canonical rod-shaped growth. Furthermore, the abundance and/or activity of l,d-transpeptidases appears to associate with polar growth strategies, suggesting that these enzymes may serve as attractive targets for specifically inhibiting growth of Rhizobiales, Actinomycetales, and other polarly growing bacterial pathogens.

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A dynamically assembled cell wall synthesis machinery buffers cell growth.

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A dynamically assembled cell wall synthesis machinery buffers cell growth.

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