Transcriptional slippage controls production of type III secretion apparatus components in Shigella flexneri

Abstract

SummaryDuring transcription, series of approximately 9 As or Ts can direct RNA polymerase to incorporate into the mRNA nucleotides not encoded by the DNA, changing the reading frame downstream from the slippage site. We detected series of 9 or 10 As in spa13, spa33 and mxiA encoding type III secretion apparatus components. Analysis of cDNAs indicated that transcriptional slippage occurs in spa13, mxiA and spa33. Changes in the reading frame were confirmed by using plasmids carrying slippage sites in the 5Ј part of lacZ. Slippage is required for production of Spa13 from two overlapping reading frames and should lead to production of truncated MxiA and Spa33 proteins. Complementation of spa13 and mxiA mutants with plasmids carrying altered sites indicated that slippage in spa13 is required for assembly of the secretion apparatus and that slippage sites in spa13 and mxiA have not been selected to encode Lys residues or to produce two proteins endowed with different activities. The presence of slippage sites decreases production of Spa13 by 70%, of MxiA and Spa33 by 15% and of Spa32 (encoded downstream from spa13) by 50%. These results suggest that transcriptional slippage controls protein production by reducing the proportion of mRNA translated into functional proteins.

Deep Sequencing Whole Transcriptome Exploration of the σE Regulon in Neisseria meningitidis

“…Different mechanisms for differential expression, e.g. transcriptional slippage might be involved, but the lack of genomic differences indicate in favor of the current finding of non-significant differential expression [56].…”

The Fidelity of Transcription

“…Additional observations of transcriptional slippage on homopolymeric tracts have been reported in bacteriophages (10), prokaryotes (11)(12)(13)(14), viruses (15), and eukaryotes (16,17). A clear demonstration of in vivo transcriptional slippage during elongation was provided by Wagner et al (18), who showed that Escherichia coli RNA polymerase made slippage RNA products from templates with a homopolymeric tract greater than 9 nucleotides.…”

The 33 carboxyl-terminal residues of Spa40 orchestrate the multi-step assembly process of the type III secretion needle complex in Shigella flexneri

“…This apparatus is evolutionarily and structurally related to the export machinery of flagellar systems (Macnab, 1999;Blocker et al, 2001). Nine T3SA constituents are related to components of the basal bodies of bacterial flagella (Allaoui et al, 1994;Aizawa, 2001;Blocker et al, 2003;Macnab, 2004), including the innermembrane proteins MxiA (FlhA), Spa9 (FliP), Spa24 (FliR), Spa29 (FliQ) and Spa40 (FlhB), and the cytoplasmic proteins Spa47 (FliI), MxiN (FliH) and Spa33 (FliN) (Kane et al, 2002;Allaoui et al, 1992bAllaoui et al, , 1993bAllaoui et al, , 1995Penno et al, 2006;Jouihri et al, 2003;Andrews & Maurelli, 1992; Sasakawa et al, 1993;Schuch & Maurelli, 2001; MoritaIshihara et al, 2005). Additionally, two cytoplasmic proteins, Spa13 and MxiK, which have no obvious counterpart in the flagellar system, are also implicated in Abbreviations: EPEC, enteropathogenic Escherichia coli; GST, glutathione S-transferase; NC, needle complex; T3S, type III secretion; T3SA, type III secretion apparatus; T3SS, type III secretion system; TEM, transmission electron microscopy.…”