Phenotypes of Bacillus subtilis mutants altered in the precursor-specific region of sigma E.
sigma E is a sporulation-specific sigma factor of Bacillus subtilis that is synthesized from an inactive precursor protein (P31). The structural gene (sigE) for P31 was reengineered by oligonucleotide-directed mutagenesis to encode sigma E directly. The sequence specifying the first amino acid of sigma E (GGC) was placed immediately downstream of the initiating codon (ATG) of P31. The resulting sigE allele (sigE delta 84) encodes a sigma E-like protein which differs from the "processed product" by a single Met residue at its amino terminus. B. subtilis strains which carried this allele were Spo- and contained no detectable sigma E. The sigE delta 84 allele generated a product in Escherichia coli which, by quantitative Western immunoblot analysis, was present at 10 to 20% of the level of product (P31) obtained from a wild-type allele. A sigma E-like product was also not detected in two B. subtilis strains with missense mutations in the sequence encoding the processed region of P31. These results suggest that sigma E is a highly labile protein that is stabilized during its synthesis by an element of the precursor sequence. A mutant allele (sigE delta 48) which made an active sigma E-like protein in B. subtilis was isolated. This gene specified a product in which five amino acids, not derived from the P31 processed region, were joined to P31 at a position eight amino acids upstream of the processing site. The sigE delta 48 product was not processed, but it activated the sigma E -dependent spoIID promoter in vivo. The sigE delta 48 product therefore lost both an essential target for processing and a region which inhibited sigma sigma E activity. Cells which carried sig E delta 48 were Spo-. The basis of the sigE delta 48-dependent defect in sporulation is unknown, but the sigma E delta 48 activity appeared to persist beyond the time in development (4 h after onset sporulation) when wild-type sigma E activity declines. Thus, it may interfere with the proper regulation of late sporulation genes.
Jonas
R M
RM
Department of Microbiology, University of Texas Health Science Center, San Antonio 78284.
Peters
H K
HK
Haldenwang
W G
WG
D000483:Alleles; D000595:Amino Acid Sequence; D001412:Bacillus subtilis / Q000235:genetics* / Q000502:physiology; D001483:Base Sequence; D002872:Chromosome Deletion; D002876:Chromosomes, Bacterial; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D008969:Molecular Sequence Data; D009154:Mutation; D010641:Phenotype; D014176:Protein Biosynthesis; D012808:Sigma Factor / Q000235:genetics*; D013171:Spores, Bacterial / Q000502:physiology; D014157:Transcription Factors / Q000235:genetics*; D014169:Transformation, Bacterial
D012808:Sigma Factor; D014157:Transcription Factors