RNase III family enzymes which are perhaps the most widely conserved

RNase III family enzymes which are perhaps the most widely conserved of all ribonucleases are known primarily for their role in the processing and maturation of small RNAs. mechanism for global ribonuclease-mediated control of gene expression in streptomycetes. PNU 282987 Introduction species are prokaryotic soil-dwelling organisms that have a complex life cycle involving mycelial growth and spore formation. In addition to producing more than two-thirds of the antibiotics currently in clinical and veterinary use are the source of multiple other types of pharmaceutically useful compounds (Challis and Hopwood 2003 For more than 40 years has been widely employed as a model organism in studies of morphological development and antibiotic regulation in streptomycetes. These studies (reviewed in Bibb 1996 Chater 2006 Flardh and Buttner 2009 plus more recent microarray-based investigations (Huang is controlled by a complex multifaceted regulatory network. The gene was discovered in a screen for mutants that are defective in antibiotic biosynthesis but which do not interfere with the morphological development associated with the production of antibiotics and other secondary metabolites in this micro-organism (Adamidis and Champness 1992 Aceti and Champness 1998 DNA sequence analysis suggested that the protein encoded by is a member of the RNase III family of endoribonucleases (Price gene were found to accumulate 30s rRNA precursors (Price (Chang (Gravenbeek and Jones 2008 Xu can dramatically and broadly affect the abundance of individual mRNAs in mutant strain (Huang (Adamidis and Champness 1992 implies that these mutation may be targets of ribonucleolytic digestion by the AbsB protein. Here we report the discovery that AbsB/RNase III regulation of genes implicated in morphological development is mediated by endoribonucleolytic cleavage of mRNA encoding AdpA initially identified in as an pleiotropic AraC/XylS family transcription factor (Ohnishi mutant bacteria in microarray studies was (SCO0762) (Huang (Taguchi point mutant strain C120 which replaces leucine by PNU 282987 proline at position 120 of the open reading frame (Adamidis and Champness 1992 showed increased sporulation (Fig. S1) and RT-PCR studies in the bHLHb24 same strain as well as in an null mutant strain (J-5572) constructed in our lab (Fig. 1A) confirmed both the dependence and the temporally regulated elevation of mRNA abundance relative to parental bacteria (J1501). This increase was most prominent 48 h after plating of spores – the time in the life cycle associated with the formation of aerial mycelium. However notwithstanding the dramatic increase in transcript abundance observed in bacteria defective in (both in C120 and The null mutant strain J-5572) we did not detect cleavage of transcripts by purified AbsB/RNase III protein (Fig. S2A) under conditions where the ribonuclease cleaved its own mRNA (Fig. S2B). These observations suggest that the regulation of at least by AbsB is accomplished indirectly rather than by AbsB/RNase III digestion of transcripts. Potentially such indirect regulation could PNU 282987 be mediated by either an AbsB-targeted regulatory RNA or an AbsB-targeted mRNA encoding a transcription-activating protein. Fig. 1 Quantitative RT-PCR analysis of abundance of (A) and (B) mRNA in J1501 (morphologically wild type) C120 (point mutant) and J-5572 (Δis only one of many sporulation-gene-associated genes whose transcripts are elevated in PNU 282987 mutant bacteria (Huang (SCO0762) is TTA-dependent and is regulated in by AdpA an AraC/XylS family transcription factor (Kim mutant bacteria. Moreover as we had observed for transcripts mRNA also was not targeted by the ribonucleolytic action of purified AbsB/RNase III protein (Fig. S2C). AdpA was discovered initially because of its effects on antibiotic biosynthesis and sporulation in (Ohnishi (Nguyen gene has also been called (Merrick 1976 Lawlor genes of all sequenced species are located at exactly the same placement in the proteins coding series recommending that their incredibly conserved area may have natural relevance. The above mentioned observations and factors raised the chance that upregulation of sporulation-associated genes in mutant bacterias could be mediated by AdpA. Consistent.

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