Date of Award

1986

Degree Type

Dissertation

Degree Name

Doctor of Philosophy

Abstract

The transcriptional control of two genes rpoB and rpoC encoding b and b', the major protein subunits of E. coli RNA polymerase, was examined in vivo. Restriction fragments from the rplKAJLrpoBC cluster of genes were fused to a lacZ gene carried on a lambda phase vector. The transcriptional activity of each cloned fragment was examined by measuring the b-galactosidase level in a lysogen carrying a transcriptional fusion phage. Two strong promoters, rplKp and rplJp, were identified along with a weak promoter, rplLp. A transcriptional attenuator, atn, located between rplL and rpoB, terminated 60 percent of transcription reading through it regardless of which promoter initiated transcription.;Hybridization studies to estimate the level of rplJL transcripts with respect to rpoBC transcripts in strains carrying rho, nus or sfrB mutations implicated the Rho, NusA and SfrB proteins in termination at atn. S1 mapping experiments confirmed the location of atn, 72 bp 3' to rplL, but suggested that the RNaseIII processing sites were 80 bp 5' to the positions previously reported. S1 mapping studies on rho, nus, and sfrB mutants did not reveal an altered pattern of termination in mutants carrying lesions in any of these genes.;From these studies it is concluded that regardless of which of the two strong promoters, rplKp or rplJp, is used to initiate transcription which reads through atn to transcribe rpoBC, transcription is reduced at atn by 60 percent. Minor promoters do not contribute substantially to rpoBC transcription. Because the frequency of transcription of rplKAJL and rpoBC remains constant at all growth rates, the changing patterns of expression seen for the proteins encoded by these genes is probably controlled post-transcriptionally. On the basis of the effects of rho and nus lesions on the rplJL/rpoBC transcript ratio, the terminator atn is designated Rho-dependent.

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