Identification and characterization of small non-coding RNAs in the archaeal diazotroph Methanosarcina mazei Gö1

Small non-coding RNAs (sRNAs) have been identified in all three domains of life. In Eukarya and Bacteria functions have been assigned for many sRNAs. To get a deeper insight into potential regulatory roles of sRNAs in Archaea, we studied the methane-producing archaeon Methanosarcina mazei strain Gö1, which is able to fix molecular nitrogen. Although many aspects of the organism’s adaptation to different stress situations are currently under investigation, e.g. limiting carbon sources (Hovey et al. 2005), nitrogen limitation (Veit et al. 2006, Weidenbach et al. 2008) and osmotic stress (Pflüger et al. 2007, Spanheimer & Müller, 2008), still less is known about the overall transcriptional organization, particularly with regard to the non-coding regions representing 25 % of the genome.

Here we present genome-wide identification of sRNAs in M. mazei, with the main focus on those potentially involved in nitrogen metabolism. Total-RNA from cells grown with ammonium and molecular nitrogen as nitrogen-source was investigated by massive parallel sequencing technology, resulting in two libraries consisting of 228.000 cDNAs in total. The performed RNA-Seq approach revealed the presence of approximately 250 small non-coding RNAs expressed from intergenic regions (IGRs). 76 of those were characterized via northern blot analysis, confirming the expression of 56 sRNAs, 18 of which are differentially transcribed in response to nitrogen availability. Selected (nitrogen regulated) sRNAs have been further analyzed using biochemical and genetic approaches.