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 of spoIIM, a gene of Bacillus subtilis whose product is required for complete septum migration and engulfment of the forespore compartment during sporulation. The spoIIM 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 sigma-E-associated RNA polymerase (E sigma E). A unique 5' end of the in vivo spoIIM 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 sigma E. A partially purified preparation of E sigma E produced a transcript in vitro that initiated at the same nucleotide as the primer extension product generated from in vivo RNA. Ectopic induction of sigma 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 sigma F or sigma G under similar conditions. The results of plasmid integration-excision experiments in which the spoIIM gene was reversibly disrupted by a temperature-sensitive integrational vector suggested that spoIIM expression is required in the forespore compartment, but direct examination of subcellular fractions enriched for mother cell or forespore material indicated that spoIIM expression cannot be confined to the forespore. We conclude that spoIIM is a member of the sigma E regulon and that it may be transcribed exclusively by E sigma E. We discuss the implications of this conclusion for models in which activation of sigma E 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.