 |
News
from Harvard Medical, Dental, & Public Health Schools |
||
|
February 19, 1999 MICROBIOLOGYUncovering the Secrets of Bacterial DivisionTo the untrained observer, the life of a bacterium could seem utterly boring. Grow and divide, sense the environment, and if things look good, grow and divide again. In a bug's life, it appears, to divide or not to divide is the main question.
But cell division is no trivial matter. An essential decision in the life of a bacterium like E. coli, division is allowed only when all other systems are go--if food is plentiful and no damage has been done to the bacterial DNA, among other things. Once DNA replication has occurred, a septum forms at a specific place and time, creating the constriction that will split the cell in two. How this septum is formed and how its formation is regulated have remained major questions for microbiologists. A trio of recent articles from the laboratory of Jonathan Beckwith, American Cancer Society professor of microbiology and molecular genetics, provides new insights into the assembly of the septum and the key players in this process. Like puzzle pieces coming together to reveal a bigger picture, the articles published in the January issues of the Journal of Bacteriology and Molecular Microbiology present a complementary analysis of three membrane proteins that localize at the division septum--FtsL, FtsI, and FtsQ--and show that their recruitment to the septum depends on a linear, ordered chain of events. String of Cells Back in the 1960s, a temperature-sensitive screen in the Pasteur Institute led to the identification of E. coli mutants with an unusual shape. Elongated and thin, these mutants were said to "filament," and were termed Fts mutants (for temperature-sensitive filamentation). The striking phenotype suggested genes involved in cell division, because these filaments were, in fact, strings of cells that failed to completely divide and remained attached at the ends.
It later became clear that not all mutants that formed filaments were defective in the process of septation. Mutations in genes involved in other cellular processes that indirectly affect division can also result in filaments. Localization of proteins at the division septum therefore became an important criterion for identifying proteins that play a role in septation. For a long time, though, only one of the septal proteins, FtsZ, was abundant enough to be clearly visualized forming a ring at the future division site. Although formation of this "Z ring" was thought to be essential for the recruitment of eight other proteins to the division site (including FtsL, Fts I, and FtsQ), technical difficulties made further studies more challenging. "It seemed clear to everyone in the field that a lot of the proteins involved in cell division must localize at the septum," says Beckwith. But immunolocalization of these proteins was not always successful. The proteins are made in very small quantities, which might explain why they were hard to detect. "This is why Green Fluorescence Protein [GFP] fusions have become incredibly useful for us," says Beckwith. The scientists fused several of the elusive Fts proteins to GFP, which served as a luminous tag revealing their location. They used a novel approach to introduce a single copy of the GFP fusion construct into the bacterial genome, thereby expressing the recombinant protein at physiological levels. When the researchers looked at the localization of GFP fusion versions of FtsI, FtsL, and FtsQ, they observed specific and clear restriction of the proteins to the Z ring. How They Get There--and When With an effective way to visualize these proteins at the division septum, the next goal was to understand how they got there. Experiments in which the distinct domains in FtsI, FtsL, and FtsQ were replaced with domains from an unrelated protein showed that each septal protein has different requirements for proper localization to the Z ring. For FtsQ, only one domain is needed, whereas FtsI requires all its domains for localization to the ring. Finally, Beckwith's group helped to elucidate a sequential relationship between the septal proteins. Using the GFP fusions, they analyzed how FtsI, FtsL, and FtsQ localized in different E. coli strains that were deficient in other septal proteins. These experiments allowed them to establish a linear order of appearance at the septum, where the presence of one Fts protein is necessary for the localization of the more downstream components. The reconstruction of this linear pathway of septal ring assembly reflects work from his and other labs, Beckwith says. "We expect that more proteins involved in this pathway will be identified," adds graduate student Joseph Chen. Not Just Bacteria Bacterial cells are not the only ones dealing with the complex issue of septation. Septal ring equivalents are present in eukaryotic cells, such as the actomyosin ring in yeast and the contractile ring in mammalian cells. How homologous these structures are, however, remains unclear. For example, bacterial cells lack actin and myosin, which are components of the rings of eukaryotic cells. The differences between the septal ring in bacteria and the ring in eukaryotic cells are currently being explored by several biotechnology companies, Beckwith says. Because they are specific to bacteria, septal proteins represent a potential target for antibiotic therapy, he adds. --Sylvia Pagán Westphal
|
