Abstract
It is well established that in Escherichia coli, the histone‐like nucleoid structuring (H‐NS) protein also functions as negative regulator of rcsA transcription. However, the exact mode of regulation of rcsA transcription by H‐NS has not been studied extensively. Here, we report the multicopy effect of dominant‐negative hns alleles on the transcription of rcsA based on expression of cps‐lac transcriptional fusion in ∆lon, ∆lon rpoB12, ∆lon rpoB77 and lon + strains. Our results indicate that H‐NS defective in recognizing curved DNA fails to repress rcsA transcription significantly, while nonoligomeric H‐NS molecules still retain the repressor activity to an appreciable extent. Together with bioinformatics analysis, our study envisages a critical role for the putative curved DNA region present upstream of rcsA promoter in the transcriptional regulation of rcsA by H‐NS.
Highlights
Introduction of pLGhnsP116S significantly increased the cps-lac expression in all SG20781/pLGhns-P116S the four strains
histone-like nucleoid structuring (H-NS) which is defective in recognizing curved DNA fails to repress rcsA transcription
In our earlier study pertaining to the isolation and characterization of novel RNA polymerase beta subunit (rpoB) mutations capable of suppressing the overproduction of colanic acid capsular polysaccharide (Cps) in lon mutant of E. coli, we have substantiated the role of functional H-NS in the elicitation of capsule expression suppression (Ces) phenotype by the two rpoB mutations, namely rpoB12 (C1576 to T1576; His526 to Tyr526) and rpoB77 (C1535 to T1535; Ser512 to Tyr512) [32]
Summary
Introduction of pLGhnsP116S significantly increased the cps-lac expression in all SG20781/pLGhns-P116S the four strains. Comparative analyses of the multicopy effect of hns+ and hnsP116S alleles on the expression of cps-lac fusion in the relevant strains clearly show the inability of the mutant form of H-NSP116S molecules to exert complete repressor activity like the wild-type it is reported to retain nonspecific DNA-binding activity (for the comparative values, see Table 2) These results strongly support the view that the upstream region of rcsA promoter is likely to contain a bendable/curved region and the inability of mutant form of H-NS (HNSP116S) to bind to such putative bendable/curved region of rcsA promoter could be the cause for higher level of cps-lac expression in the relevant strains. Introduction of pLG-hnsΔ64 increased the cps-lac expression to some extent that clearly implies that the H-NS molecules bearing only the N-terminal region are not completely defective in repression at rcsA promoter
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