Evidence that the spoIIM gene of Bacillus subtilis is transcribed by RNA polymerase associated with sigma E

Abstract

We have investigated the temporal and spatial regulation ofspoHIM, a gene ofBacillus subtilis whose product is required for complete septum migration and engulfment of the forespore compartment during sporulation. The spoHIM promoter was found to become active about 2 h after the initiation of sporulation. The effects of mutations on the expression of a spoIIM-lacZ fusion were most consistent with its utilization by &-Fassociated RNA polymerase (Eci). A unique 5' end of the in vivo spoHIM transcript was detected by primer extension analysis and was determined to initiate at the appropriate distance from a sequence conforming very closely to the consensus for genes transcribed by E&F. A partially purified preparation of E&X produced a transcript in vitro that initiated at the same nucleotide as the primer extension product generated from in vivo RNA. Ectopic induction of &E synthesis during growth resulted in the immediate and strong expression of a spoIIM-lacZ fusion, but an identical fusion was completely unresponsive to induced synthesis of either oF or rG under similar conditions. The results of plasmid integration-excision experiments in which the spolIM gene was reversibly disrupted by a temperature-sensitive integrational vector suggested that spolIM expression is required in the forespore compartment, but direct examination of subcellular fractions enriched for mother cell or forespore material indicated that spolIM expression cannot be confined to the forespore. We conclude that spoHIM is a member of the &E regulon and that it may be transcribed exclusively by Ecr-. We discuss the implications of this conclusion for models in which activation of o1r in the mother cell is proposed to be a part of the mechanism responsible for initiating separate programs of gene activity in the two sporangium compartments.Endospore formation in Bacillus subtilis involves a series of temporally and spatially ordered changes in cell morphology and gene expression (42,54). At a critical stage in this process, an asymmetric septation event creates two cell compartments, the mother cell and the forespore, each containing a transcriptionally active chromosome. At some point very soon after asymmetric septation, the mother cell and the forespore compartments begin to execute quite distinct programs of gene expression. The mechanisms responsible for establishing these compartment-specific programs are not fully understood, but they apparently involve, in part, the compartment-specific activation of RNA polymerase sigma factors (16,40,41,76). This is most obvious at the later stages of sporulation, in which a polymerase species associated with g is active exclusively in the mother cell compartment (11,36,43) and a species associated with cyG is active exclusively in the forespore (31, 78). At earlier stages, the situation is less certain and the basis for compartmentspecific gene activity is more speculative. Soon after asymmetric septation, a polymerase species associated with &rF becomes active in the forespore (45, 67). The o-F...

Compartmentalization of Gene Expression duringBacillus subtilisSpore Formation

“…Transcription of the spoIID (246), spoIIM (272), and spoIIP (85) loci is directed by E , suggesting that initiation of engulfment is driven by proteins synthesized in the mother cell. Consistent with a direct role in engulfment, GFP fusions with all three of the encoded proteins localized to the asymmetric septum and engulfing prespore membrane (1,57).…”

“…Given that E -directed genes are required to cause septal wall autolysis during engulfment (1,37,57,85,228,232,246,271,272), it seemed plausible that similar activity might enlarge a breach in the septum resulting from loss of SpoIIIE. Indeed, E -directed transcription of spoIID and spoIIP is required to disrupt compartmentalization in spoIIIE class II mutants (110).…”

“…Members of the E regulon that are important for sporulation include spoIID, spoIIM, and spoIIP, which are required for engulfment and to prevent a second asymmetric division from occurring in the mother cell (1,37,57,85,228,232,246,271,272); spoIVA, cotE, and spoVID, which encode scaffold proteins for spore coat assembly (18,245,273,317,318); the spoIIIA operon, which is required for activation of the lateprespore specific sigma factor G (125, 146); sigK, the composite gene for the late mother cell-specific transcription factor K (155,280); and spoIVCA, the gene for the recombinase that generates sigK via a chromosomal rearrangement (155,235,256). In addition, E activates transcription of spoIIID, which encodes a regulator of some E -dependent genes (102).…”

Contrasting Effects of σEon Compartmentalization of σFActivity during Sporulation ofBacillus subtilis

“…There are three proteins essential for engulfment that are thought to be directly involved in septal thinning: SpoIID, SpoIIM, and SpoIIP (1,7,13,29,40). They are all expressed in the mother cell under the control of E (13,35,41) and localize to the engulfing septum (1,11). Recently, SpoIID has been reported to degrade the bacterial cell wall in vitro (1).…”

cse15, cse60, and csk22 are new members of mother-cell-specific sporulation regulons in Bacillus subtilis

“…Conversion of pro-E to its active form, E , which occurs exclusively in the mother cell, inaugurates the program of gene expression in this compartment (19,35,44,68). The E form of RNA polymerase (E E ) directs the transcription of a heterogeneous group of genes controlling functions as diverse as engulfment (23,56,58,62), germination (49,70), cortex synthesis (2,9,14,43,52,54,60,64), and coat morphogenesis (3,18,28,49,54,60,70), as well as genes that may assist in the control of the metabolic state of the mother cell (7,8,26,37). E E also drives expression of at least two regulatory genes, spoIIID (40,62,68) and sigK (39,68).…”