Rate and topography of cell wall synthesis during the division cycle of Salmonella typhimurium

Abstract

The rates of synthesis of peptidoglycan and protein during the division cycle of Salmonella typhimurium have been measured by using the membrane elution technique and differentially labeled diaminopimelic acid and leucine. The cells were labeled during unperturbed exponential growth and then bound to a nitrocellulose membrane by filtration. Newborn cells were eluted from the membrane with fresh medium. The radioactivity in the newborn cells in successive fractions was determined. As the cells are eluted from the membrane as a function of their cell cycle age at the time of labeling, the rate of incorporation of the different radioactive compounds as a function of cell cycle age can be determined. During the first part of the division cycle, the ratio of the rates of protein and peptidoglycan synthesis was constant. During the latter part of the division cycle, there was an increase in the rate of peptidoglycan synthesis relative to the rate of protein synthesis. These results support a simple, bipartite model of cell surface increase in rod-shaped cells. Before the start of constriction, the cell surface increased only by cylindrical extension. After cell constriction started, the cell surface increased by both cylinder and pole growth. The increase in surface area was partitioned between the cylinder and the pole so that the volume of the cell increased exponentially. No variation in cell density occurred because the increase in surface allowed a continuous exponential increase in cell volume that accommodated the exponential increase in cell mass. Protein was synthesized exponentially during the division cycle. The rate of cell surface increase was described by a complex equation which is neither linear nor exponential.There have been many measurements of the rate of cell wall synthesis during the division cycle (25,38,54,60,64,69), and although they differ in detail, they have generally arrived at similar results. There is a continuous increase in the rate of surface synthesis during the division cycle, with some reports of peaking in the rate of synthesis near the end of the division cycle. Much of the theoretical analysis of these results has been directed toward distinguishing among simple mathematical descriptions of synthesis such as exponential or linear growth patterns or combinations of these (3,

Peptidoglycan synthesis in Salmonella typhimurium 2616

“…We observed an increase in the ratio of DAP to leucine in the older cells, peaking at 0-7 during the cell cycle. This is in agreement with results obtained using the backwards membrane-elution method (Cooper, 1988 ; ; unpublished results). The bars in Fig.…”

“…Results obtained using the membrane-elution method have led to our understanding of the pattern of DNA replication during the bacterial division cycle (Cooper,199 1 a). Membrane-elution studies with S. typhimurium 2616 and E. coli NC3 have also shown that there is a reproducible non-exponential pattern of peptidoglycan synthesis during the division cycle (Cooper, 1988 ;. Theoretical considerations prompted by these findings suggest that the point at which septal peptidoglycan synthesis is greatest may not be at the end of the division cycle.…”

“…Salmonella typhimurium 26 16 (lysA) (from Dr K. E. Sanderson, University of Calgary) was grown in C medium supplemented with 50 pg lysine ml-' (Cooper, 1988). Carbon sources (glucose, glycerol, succinate or acetate) were added to a final concentration of 0.4%.…”

“…4(a, b) indicate the time when the DAP to leucine ratio is maximal. This may indicate the time when septum synthesis, as a fraction of total cell wall, is maximal (Cooper, 1988). D. Gully and S. Cooper The percentage of radiolabelled DAP in tetra, tetratetra and tetra-tri muropeptides during the division cycle are shown in Fig.…”

“…the radiolabelling of an exponential culture followed by fractionation of cell ages by elution from a nitrocellulose membrane (Cooper, 1988;, Gally et al, 1993. Maturational changes exhibited by pulselabelled peptidoglycan restrict the utility of the backwards method for the analysis of the composition of peptidoglycan synthesized at different times during the division cycle.…”