Harvard Medical School


Contact Information:

Harvard Medical School

HIM, Room 1026

4 Blackfan Circle

Boston, MA 02115

Phone: (617) 466-9851
Fax: (617) 432-4787

 

e-mail: thomas_bernhardt@hms.harvard.edu

Research Summary

Bacterial cells come in a wide variety of shapes and sizes. Within a given class, however, cells typically grow and divide in such a way as to maintain the particular shape and size distribution programmed by their genome. Research in my laboratory is focused on understanding how these programs are executed by uncovering the molecular mechanisms organizing cell envelope assembly and promoting cell division. The general problems we wish to address are: i) how the synthetic machineries that make the cell membrane, cell wall (peptidoglycan), and other envelope layers are coordinated in space and time to construct a cell with uniform shape, ii) how the cell division machinery drives the invagination of the cell envelope and builds the new daughter cell poles, and iii) how cells know where and when to divide in the first place. A multidisciplinary approach including genetics, biochemistry, microscopy, and genomics will be taken to address these questions using the Gram-negative rod Escherichia coli as our primary model bacterium. Ideally, our work will not only help further our understanding of fundamental aspects of bacterial cell biology, but will also provide new avenues with which to short-circuit the bacterial growth program for potential therapeutic intervention.

Current Projects

Division-site placement:

To begin the division process, the bacterial tubulin protein FtsZ polymerizes into a dynamic ring-like structure just underneath the cell membrane at the prospective site of fission. This Z-ring then recruits a host of additional division proteins to organize the assembly of a multi-protein molecular machine that promotes cytokinesis. We have recently identified SlmA as a chromosome-associated, FtsZ-interacting protein that controls Z-ring placement and helps prevent cells from dividing over their chromosomes. Genetic, cell biological, biochemical, and genomic experiments are now underway to further explore SlmA function and the mechanism by which it regulates Z-ring assembly. Additional global genetic screens will also be performed to help us discover other regulators governing the timing and placement of Z-ring assembly. Please see publication (7) for more details.

Peptidoglycan synthesis and remodeling:

Most bacteria surround themselves with a crosslinked polysaccharide called peptidoglycan (PG) or murein. This PG layer is essential for the integrity of most bacterial cells with any breach in its continuity leading to rapid cell lysis and death. In addition to its essentiality, PG is unique to the bacterial world, and many of the enzymes required for its assembly are surface exposed. These properties make PG synthesis an ideal target for antibacterial agents. Indeed, many of our most successful antibacterial therapies target PG production (penicillin and vancomycin, for example).

Construction of the PG layer requires enzymes that make the polysaccharide strands (transglycosylases) and enzymes that form the peptide crosslinks (transpeptidases). Adding new material to the existing PG structure is also thought to require the action of murein hydrolases, enzymes that break PG linkages. The E. coli genome codes for over thirty potential murein hydrolases. In all but a few cases, the physiological role(s) of these enzymes are unknown. In addition, it is unclear how the activities of these enzymes are held in check to prevent uncontrolled PG degradation and cell lysis. To better understand the function of these enzymes and their potential role(s) in PG assembly and remodeling, we are currently taking advantage of new cell biological tools to study their subcellular localization in addition to initiating genetic and biochemical studies to define their precise enzymatic activities and how they may be regulated. Please see publications (1, 2, 4, 8, 9 and 11-13 for more details.

Future Projects

We are always open to new ideas for projects aimed at understanding the cell biology of bacteria. As the laboratory grows, we plan on pursuing additional projects geared towards understanding interesting cell biological problems in bacterial species other than E. coli. Please feel free to contact us with your ideas and inquire about positions available in the Bernhardt lab.

Publications

1. Cho, H., McManus, H. R., Dove, S.L., Bernhardt, T.G. (2011) The nucleoid occlusion factor SlmA is a DNA-activated FtsZ polymerization antagonist. Proc. Natl. Acad. Sci.
(in press)

2. Paradis-Bleau C.1, Markovski M.1, Uehara T.1, Lupoli T., Walker S., Kahne D., Bernhardt T.G. (2010) Lipoprotein cofactors located in the outer membrane activate bacterial cell wall polymerases. Cell 143:1110-1120 (1co-first authors)

3. Uehara, T, Parzych, KR, Dinh, T, and Bernhardt, T.G. (2010) Daughter cell separation is controlled by cytokinetic ring-activated cell wall hydrolysis. EMBO J. 29: 1412-1422 [PMC2868575]

4. Morlot, C.1, Uehara, T.1, Marquis, K. A., Bernhardt, T.G.2 and Rudner, D.R.2 (2010) A highly coordinated cell wall degradation machine governs spore morphogenesis in Bacillus subtilis. Genes Dev 24: 411-422 [PMC2816739]
1co-first authors, 2corresponding authors

5. Gerding, M.A., Liu, B, Bendezú, F.O., Hale, C.A., Bernhardt, T.G. and de Boer, PA (2009) Self-enhanced accumulation of FtsN at division sites, and roles for other proteins with a SPOR domain (DamX, DedD, and RlpA) in Escherichia coli cell constriction. J Bacteriol 191:7383-7401 [PMC2786604]

6. Uehara, T., Dinh, T., and Bernhardt, T.G. (2009) LytM-domain factors are required for daughter cell separation and rapid ampicillin-induced lysis in Escherichia coli. J Bacteriol 191:5094-5107 [PMC2725582]

7. Bendezu, F.O., Hale, C.A., Bernhardt, T.G., and de Boer, P. (2009) RodZ (YfgA) is required for proper assembly of the MreB actin cytoskeleton and cell shape in E. coli. EMBO J. 28: 193-204 [PMC2637328]

8. Zheng., Y., Struck, D.K., Bernhardt, T.G., and Young, R. (2008) Genetic analysis of MraY inhibition by the φX174 E protein. Genetics 180: 1459-1466 [PMC2581948]

9. Bernhardt, T.G. and de Boer, P. (2005). SlmA, a nucleoid associated, FtsZ-binding protein required for blocking septal ring assembly over chromosomes in E. coli. Mol. Cell 18: 555-564.

10. Bernhardt, T.G. and de Boer, P. (2004). Screening for synthetic lethal mutants in Escherichia coli and identification of EnvC(YibP) as a periplasmic septal ring factor with murein hydrolase activity. Mol. Microbiol. 52: 1255-1269.

11. Bernhardt, T.G. and de Boer, P. (2003). The Escherichia coli amidase AmiC is a periplasmic septal ring component exported via the twin arginine transport pathway. Mol. Microbiol. 48: 1171-1182.

12. Bernhardt, T.G., Roof, W.D., Young, R.  (2002).  The E. coli FKBP-type PPIase is required for the stabilization of the E lysis protein of bacteriophage φX174. Mol. Microbiol. 45: 99-108.

13. Bernhardt, T.G.*, Wang, I.*, Struck, D.K., and Young, R.  (2001).  A protein antibiotic in the phage Qβ virion: Diversity in lysis targets. Science 292:2326-2329. *(co-first author)

14. Bernhardt, T.G., Struck, D.K., and Young, R.  (2001).  The lysis protein E of φX174 is a specific inhibitor of the MraY-catalyzed step in peptidoglycan synthesis. J. Biol. Chem.  276:6093-6097.

15. Bernhardt, T.G., Roof, W.D., Young, R.  (2000).  Genetic evidence that the bacteriophage φX174 lysis protein inhibits cell wall synthesis. Proc. Natl. Acad. Sci.  97:4297-4302.