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  • Title: Expression of the E.coli hemolysin secretion gene hlyB involves transcript anti-termination within the hly operon.
    Author: Koronakis V, Cross M, Hughes C.
    Journal: Nucleic Acids Res; 1988 Jun 10; 16(11):4789-800. PubMed ID: 2455277.
    Abstract:
    The genes hly A and hly B dictating synthesis and secretion of hemolytic toxin by Escherichia coli are transcribed as part of an operon hly C, hly A, hly B but are separated by an inverted repeat and poly-T sequence characteristic of a rho-independent terminator. The hly A-hly B intergenic sequence caused a reduction of in vivo transcriptional read-through from the gal promotor into gal K when inserted in the pKG1900 terminator-probe vector and also in vitro 3' to the bacteriophage T7 luminal diameter 10 promotor. Hybridisation of mRNA generated by the hly determinant of recombinant DNA pBR202-312 to an antisense RNA probe spanning the hly intergenic sequence revealed specific termination of 80% of in vivo hly transcripts within the poly-T sequence, immediately preceding the hly B translation start. Extension of the hly determinant with 3.5kbp of hly promotor-proximal DNA sequence raised intracellular and cell-free hemolytic activity 3-fold and 20-fold respectively and increased markedly the secretion of the 107Kd hemolysin protein (HlyA). Parallel hybridisation of in vivo mRNA to hly A and hly B antisense probes revealed that levels of hly A and hly B mRNA were increased by approximately 3-fold and 90-fold. The dramatically enhanced level of hly B mRNA resulted primarily from specific suppression of transcription termination at the intergenic rho-independent terminator from 80% to 1%, thus allowing virtually all hly A transcripts to elongate into hly B.
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