RNase E Affects the Expression of the Acyl-Homoserine Lactone Synthase Gene sinI in Sinorhizobium meliloti

Abstract

c Quorum sensing of Sinorhizobium meliloti relies on N-acyl-homoserine lactones (AHLs) as autoinducers. AHL production increases at high population density, and this depends on the AHL synthase SinI and two transcriptional regulators, SinR and ExpR. Our study demonstrates that ectopic expression of the gene rne, coding for RNase E, an endoribonuclease that is probably essential for growth, prevents the accumulation of AHLs at detectable levels. The ectopic rne expression led to a higher level of rne mRNA and a lower level of sinI mRNA independently of the presence of ExpR, the AHL receptor, and AHLs. In line with this, IPTG (isopropyl-␤-D-thiogalactopyranoside)-induced overexpression of rne resulted in a shorter half-life of sinI mRNA and a strong reduction of AHL accumulation. Moreover, using translational sinI-egfp fusions, we found that sinI expression is specifically decreased upon induced overexpression of rne, independently of the presence of the global posttranscriptional regulator Hfq. The 28-nucleotide 5= untranslated region (UTR) of sinI mRNA was sufficient for this effect. Random amplification of 5= cDNA ends (5=-RACE) analyses revealed a potential RNase E cleavage site at position ؉24 between the Shine-Dalgarno site and the translation start site. We postulate therefore that RNase E-dependent degradation of sinI mRNA from the 5= end is one of the steps mediating a high turnover of sinI mRNA, which allows the Sin quorum-sensing system to respond rapidly to changes in transcriptional control of AHL production.

RNase E and RNase J are needed for S-adenosylmethionine homeostasis in Sinorhizobium meliloti

“…We used the S. meliloti strain Rm2011 (referred to here as strain 2011; [48,49] and its RNase E and RNase J mutants 2011rne :: Tn5 and 2011rnj :: Tn5, which harbour mini-Tn5 insertions in the respective genes rne (SMc01336) and rnj (SMc01929) [7,31,37]. The mutant strains are referred to here as 2011rne and 2011rnj, respectively.…”

“…Additionally, the SinR homologue ExpR senses AHLs and transcriptionally regulates sinR, sinI and other genes depending on population density, which is reflected by the AHL concentration [43,44]. As mentioned above, RNase E and the sRNA RcsR1 are involved in sinI mRNA regulation [7,21]. Furthermore, posttranscriptional regulation of sinI and expR by the RNA chaperone Hfq has also been described [45][46][47].…”

“…In contrast, an S. meliloti strain with a knockout mutation of the RNase E encoding gene rne is still not available. A previous analysis of an RNase E mutant with a mini-Tn5 insertion in the 3¢-half of rne, which constitutively expresses rne from a plasmid, revealed rapid degradation of the autoinducer synthase mRNA sinI mRNA and diminished autoinducer (acylhomoserine lactone) amounts due to rne overexpression [7]. RNase E seems to be essential in S. meliloti and the RNase E mutant most probably expresses a truncated RNase E that has defects in degradosome formation [7].…”

“…A previous analysis of an RNase E mutant with a mini-Tn5 insertion in the 3¢-half of rne, which constitutively expresses rne from a plasmid, revealed rapid degradation of the autoinducer synthase mRNA sinI mRNA and diminished autoinducer (acylhomoserine lactone) amounts due to rne overexpression [7]. RNase E seems to be essential in S. meliloti and the RNase E mutant most probably expresses a truncated RNase E that has defects in degradosome formation [7]. The S. meliloti degradosome probably plays a role in the posttranscriptional regulation of sinI mRNA, since full-length RNase E is needed for degradation of this mRNA upon overexpression of the small RNA (sRNA) RcsR1 [21].…”

“…One of the most important bacterial RNases is the endoribonuclease, RNase E, which is essential for growth in most bacteria analysed in this respect, including S. meliloti [6][7][8]. E. coli RNase E participates in rRNA [9,10] and tRNA processing [11], is crucial for mRNA turnover [12][13][14], prefers 5¢-monophosphorylated over 5¢-triphosphorylated RNA [15], cleaves at single-stranded, AU-rich regions [12] and is localized at the membrane [16].…”