Many aspects of chlamydial division remain undiscovered. The sequenced genome reveals that chlamydiae lack many of the proteins required for septation in other bacterial species, suggesting a unique mechanism for cytokinesis. This work describes a unique chlamydial antigenic structure, termed SEP, which localizes to an apparent septum in dividing chlamydiae. SEP is developmental-stage dependent, localizing to a ring-like structure at the plane of RB division and becoming punctate and irregular after differentiation to EB. SEP is redistributed to distinct sites along the bacterial periphery following treatment of Chlamydia-infected cells with ampicillin or following incubation of infected cells in medium lacking Trp. Removal of these stressors leads to reversion of aberrant forms to typical RB and results in redistribution of SEP back to the apparent plane of bacterial division. This antigen is present in C. trachomatis, C. psittaci, and C. pneumoniae, suggesting that the structure is conserved within the genus. Collectively these results suggest a possible role for SEP in the chlamydial division process.
Recently much has been established surrounding the molecular biology of bacterial cell division. At least nine proteins which localize to the septal ring (a ring of proteins at the site of cytokinesis) are required for bacterial division in E. coli
: FtsZ, ZipA, FtsW, FtsA, FtsL, FtsN, FtsQ, FtsK, and FtsI (PBP3) (6
). Other Fts proteins, FtsH, FtsJ, FtsY, FtsX, and FtsE, may play an indirect role in septation. Essential to activating the septal protein assembly pathway is FtsZ ring formation at the division site. Paradoxically, chlamydiae lack an ftsZ
homolog but do encode predicted proteins that are likely homologous to proteins involved in septation, including FtsK, FtsW, FtsY, FtsH, and FtsI (27
). Of these proteins, FtsI, FtsW, and FtsK show immunofluorescence staining patterns in E. coli
similar to that seen with SEP localization to the ring in chlamydiae (1
). However, immunoblot analyses with anti-SEP antisera did not identify candidate proteins that might be the target antigen, suggesting that the SEP antigen may be nonproteinaceous.
Of the few fts
genes present in the chlamydial genome, homologs to ftsI
and possibly ftsW
are involved in PG biosynthesis at the septal plane in E. coli
). These findings, along with the facts that the chlamydial genome contains all genes necessary for PG synthesis and that Chlamydia
is highly sensitive to inhibitors of PG synthesis, are contradictory to other studies which conclude an absence of PG within the chlamydial cell.
Cell wall inhibitors block PG assembly through several mechanisms (25
). In some cases this leads to accumulation of PG precursors within the bacterial cell. Treatment of E. coli
with moenomycin, which inhibits the transglycosylation reaction, promotes the accumulation of several PG precursors prior to cell lysis. In contrast, treatment with penicillin G, which inhibits the transpeptidation reaction, results in unchanged or decreased concentrations of such precursors (17
). In the present study, while treatment with ampicillin and culture in Trp-deficient medium both led to SEP redistribution, there were differences in the observed phenotype among the different chlamydial species. Within C. psittaci
GPIC, SEP distribution patterns following ampicillin treatment and following Trp starvation were very similar. However, within C. trachomatis
L2, SEP was very abundant following aberrancy produced by ampicillin treatment but virtually undetectable following Trp starvation. The observed differences in SEP accumulation between these two species are perplexing. Because ampicillin treatment has no effect on chlamydial protein synthesis, production and apparent accumulation of enzymes and possibly PG precursors may occur. This effect is markedly more evident in C. trachomatis
than in C. psittaci
. In contrast, depletion of available intracellular Trp affects the production of many proteins; 15 of the 18 PG biosynthesis proteins contain Trp. This may result in only small amounts of critical enzymes accumulating within treated chlamydiae. The difference in SEP accumulation observed between Trp-starved C. psittaci
and Trp-starved C. trachomatis
may reflect differences in their needs for Trp in the synthesis of various PG precursors.
In most walled bacteria PG serves two purposes. It forms a structural sacculus providing osmotic stability to the organism, and it forms a scaffold during initiation of the septation process (20
). There is considerable evidence that the chlamydiae probably do not require PG for structural stability within the cell envelope, as this function is provided by disulfide-linked outer membrane proteins. However, the second function remains a possible role for chlamydial PG. We hypothesize that small amounts of PG may function during septum formation within dividing RB. Our studies identify an antigen that either may be this PG or may colocalize with this theoretical structure during growth. The ability of MCWS to adsorb anti-SEP activity and the affinity of anti-SEP for the E. coli
cell wall suggests that anti-SEP may be binding directly to an antigen in common between MCWS and the E. coli
cell wall. Since E. coli
is contained within a murein sacculus, one such candidate antigen is PG. Further experiments are in progress to more clearly identify the target of anti-SEP and to examine the function of SEP in chlamydial growth.