sIHF IS A NOVEL NUCLEOID-ASSOCIATED PROTEIN SPECIFIC TO THE ACTINOBACTERIA

Abstract

<p>The relatively recent discoveries of bacterial small RNAs (sRNAs) and their important regulatory functions prompted us to conduct a genome wide survey for sRNAs in <em>Streptomyces coelicolor</em>. We used a combined bioinformatics and experimental approach to identify and characterize six sRNAs. sRNA expression profiles were determined throughout <em>S. coelicolor</em> development, including vegetative and reproductive growth, during growth on minimal and rich media. Additionally, we also tested sRNA expression in various <em>S. coelicolor</em> developmental mutants. Two sRNAs were expressed exclusively during growth on one medium type and all but one were expressed constitutively throughout growth apart from the late sporulation timepoint. One of the identified sRNAs, scr1906, appeared to be closely associated with development. scr1906 was only expressed in nutrient limiting conditions just prior to aerial development and sporulation. Expression of scr1906 was abolished in a mutant that was defective in sporulation (due to a mutation in the sporulation sigma factor gene, <em>whiG</em>); however, expression was detected in mutants of both known σ^WhiG target genes, <em>whiH</em> and <em>whiI</em>, which encode sporulation transcription factors. Intriguingly, <em>in silico</em> analysis predicted <em>whiH</em> to be a direct target for scr1906-mediated regulation based on potential nucleotide binding sites. The effects of deletion and overexpression of <em>scr1906</em> on WhiH levels were tested, but require further experimentation.</p> <p>In a separate line of investigation, we sought to characterize a novel actinobacterial-specific protein named sIHF. The <em>sIHF</em> mutant strain revealed that sIHF influenced DNA compaction and segregation during <em>S. coelicolor</em> sporulation and also affected antibiotic production. sIHF associated with the nucleoid, and <em>in vitro</em>, it bound to DNA non-specifically in a length dependent manner, although it was determined to have a preference for three distinct DNA motifs. Like most nucleoid-associated proteins, sIHF affected gene expression indicating the potential for an additional role as a transcription factor. Interestingly, sIHF impacted the activity of topoisomerase. Leveraging information that we have gained from the sIHF-DNA co-crystal complex, studies aimed at characterizing the sIHF regions that are important for DNA interaction and topoisomerase modulation are currently underway.</p>