Expression of a germination-specific amidase, SleB, of bacilli in the forespore compartment of sporulating cells and its localization on the exteriorside of the cortex in dormant spores

A germination-specific amidase of bacilli is a major spore-lytic enzyme tha t is synthesized with a putative signal sequence and hydrolyses spore corte x in situ. The sleB gene encoding this amidase in Bacillus subtilis and Bac illus cercus was expressed in the forespore compartment of sporulating cell s under the control of sigma(G), as shown by Northern blot and primer exten sion analyses, The forespore-specific expression of B. subtilis sleB was fu rther indicated by the forespore-specific accumulation of a SleB-green fluo rescent protein fusion protein from which a putative secretion signal of Sl eB was deleted. Immunoelectron microscopy with anti-SleB antiserum and a co lloidal gold-immunoglobulin G complex showed that the enzymes from both Bac illus species are located just inside the spore coat layer in the dormant s pore, and in the dormant spore, the amidases appear exist in a mature form lacking a signal sequence, These results indicate that SleB is translocated across the forespore's inner membrane by a secretion signal peptide and is deposited in cortex layer synthesized between the forespore inner and oute r membranes. The peripheral location of the spore-lytic enzymes in the dorm ant spore suggests that spore germination is initiated at the exterior of t he cortex.