Bacillus subtilis spore coat

Abstract

In response to starvation, bacilli and clostridia undergo a specialized program of development that results in the production of a highly resistant dormant cell type known as the spore. A proteinacious shell, called the coat, encases the spore and plays a major role in spore survival. The coat is composed of over 25 polypeptide species, organized into several morphologically distinct layers. The mechanisms that guide coat assembly have been largely unknown until recently. We now know that proper formation of the coat relies on the genetic program that guides the synthesis of spore components during development as well as on morphogenetic proteins dedicated to coat assembly. Over 20 structural and morphogenetic genes have been cloned. In this review, we consider the contributions of the known coat and morphogenetic proteins to coat function and assembly. We present a model that describes how morphogenetic proteins direct coat assembly to the specific subcellular site of the nascent spore surface and how they establish the coat layers. We also discuss the importance of posttranslational processing of coat proteins in coat morphogenesis. Finally, we review some of the major outstanding questions in the field.

FIG. 1

B. subtilis spore. (A to C) Wild-type spore (B) or arcs of spore coats (A and C). (D) cotE spore. IC, inner coat; OC, outer coat; Cx, cortex; gap, the space between the inner coat and the cortex in a cotE spore; Cl, clump of darkly staining material of unknown composition. Bars, 500 nm (B and D) and 300 nm (A and C).

FIG. 2

Stages of sporulation. (A) Once the cell commits to sporulation, ς^H activity increases. (B) In the next stage, an asymmetrically positioned septum divides the cell into the forespore and mother cell compartments. ς^F becomes active in the forespore, and ς^E becomes active in the mother cell. (C) The forespore engulfs into a membrane-bound protoplast. ς^G becomes active in the forespore, and ς^K directs gene expression in the mother cell. (D) The cortex (the hatched area) forms between the forespore membranes. GerE works in conjunction with ς^K to direct a final phase of gene expression. (E) The coat (the dark ring surrounding the hatched cortex) becomes visible by electron microscopy. (F) In the final stage of sporulation, the mother cell lyses and releases the mature spore into the environment. (G) When nutrient returns to the medium, the spore can germinate and the cell can resume vegetative growth. This involves rehydration of the interior of the spore and cracking open of the coat.

FIG. 3

Program of mother cell gene expression. The stages of sporulation are shown at the top of the figure, and the transcription factors that direct mother cell gene expression at each stage are shown within the mother cell compartment. Below each cell are the coat protein genes that are active at that time. The repressive functions of SpoIIID and GerE are not indicated.

FIG. 4

Model for the assembly of the B. subtilis spore coat. The diagrams represent arcs of the forespore surface. The letters within the diagrams indicate the possible times of assembly and locations of some of the coat proteins. For example, CotJA assembles early, in panel B, within the matrix. Once a coat protein is indicated, it is presumed to remain in the coat for the duration of sporulation; i.e., CotJA is still present although not explicitly shown in panels C and D. (A) After septation and as a result of ς^E activity, SpoIVA localizes to the mother cell side of the forespore membranes. (B) The precoat, consisting of the matrix and the layer of CotE, is assembled over SpoIVA. CotJA and CotJC may be matrix components. (C) After ς^K becomes active, the cortex appears and a large set of coat protein genes are expressed, of which only a subset are represented. CotD, CotH, CotS, and CotT assemble into the inner coat, and CotA and CotM are built into the outer coat. CotH could sit near CotE at the interface of the inner and outer coat layers. CotM may form a substructure within the outer coat. (D) In a final stage, under the control of GerE, an unknown morphogen (represented by ?) directs the completion of the inner coat. CotB and CotG are synthesized and incorporated into the outer coat. Further modifications to the coat, including glycosylation (due to the Sps and Cge proteins), proteolysis, and cross-linking, bring the coat to its final form. Several especially speculative aspects of this model should be noted. The products of coat protein genes that are expressed under the control of ς^K are illustrated as being incorporated into the coat soon after synthesis but may in fact be assembled later, after GerE is active. For example, we do not know that CotD is assembled into the coat before CotB. FS, forespore; MC, mother cell.

FIG. 5

Roles of SpoIVA and SpoVID in attachment of the coat to the forespore membrane. The last two stages represented in Fig. 4 are compressed into a single diagram in panel C. The open boxes represent the early form of SpoVID, when it is not required for matrix attachment. The filled boxes represent the late state of SpoVID, when it becomes required for matrix attachment. FS, forespore; MC, mother cell.