Downloaded from www.microbiologyresearch.org by IP: 54.237.25.84 On: Sat, 22 Oct 2016 06:45:52 Microbiology (2002), 148, 2591–2598 Printed in Great Britain Identification and characterization of novel small RNAs in the aspS–yrvM intergenic region of the Bacillus subtilis genome Satoru Suzuma, 1 Sayaka Asari, 1 Keigo Bunai, 1 Keiko Yoshino, 1 Yoshinari Ando, 1 Hiroshi Kakeshita, 1 Masaya Fujita, 2 Kouji Nakamura 1 and Kunio Yamane 1 Author for correspondence : Kouji Nakamura. Tel : 81 298 53 6419. Fax: 81 298 53 7723. e-mail : nakamura.koujinifty.ne.jp 1 Institute of Biological Sciences, University of Tsukuba, Tsukuba-shi, Ibaraki 305-8572, Japan 2 Department of Molecular and Cellular Biology, The Biological Laboratories, Harvard University, 16 Divinity Avenue, Cambridge, MA 01238, USA A novel RNA species was isolated from Bacillus subtilis, and its sequence was determined and mapped to its genetic position. This RNA was termed BS190 RNA from the length of its mature form (190 nt), and the gene encoding it is located within the aspS–yrvM intergenic region of the B. subtilis genome. Northern blotting revealed that the novel RNA species is transcribed in vegetative cells as a larger precursor (BS201 RNA, 201 nt). After transcription, the 5 end of the precursor is processed to generate the mature form, BS190 RNA. A computer-aided prediction of the secondary structure of BS190 RNA showed that it can be folded into a single hairpin structure with some bulge structures. The authors found that the growth rate of a ΔBS190 mutant strain of B. subtilis was reduced when compared to the wild-type. A phylogenetic comparison of the sequence of the BS190 RNA gene with sequences from the databases suggests that RNA related to BS190 RNA appears to be encoded in the genomes of Bacillus halodurans and Listeria monocytogenes. Keywords : non-coding RNA, RNA processing, RNA secondary structure INTRODUCTION Bacterial and eukaryotic cells contain small RNAs (sRNAs) that are not translated (Wassarman et al., 1999 ; Wightman et al., 1993 ; Lankenau et al., 1994). These sRNAs function as RNA molecules instead of encoding proteins. Non-coding sRNA genes are quite numerous and their functions vary (Eddy, 1999 ; Erdman et al., 2001 ; Storz, 1999). Most of these non-coding sRNAs can be defined as regulatory molecules that seem to fine-tune the cellular responses to stress by integrating environmental signals into global regulation (Altuvia et al., 1997; Zhang et al., 1998). sRNAs act via several mechanisms, such as RNA–RNA base-pairing, RNA– protein interactions and intrinsic RNA activity (Altuvia & Wagner, 2000 ; Wassarman & Storz, 2000 ; Kirsebom, 2001) ; they also regulate diverse cellular functions, including RNA processing, mRNA stabilization, trans- lation, protein stabilization and protein secretion. At least 10 sRNAs are encoded within the Escherichia coli ................................................................................................................................................. Abbreviations : scRNA, small cytoplasmic RNA ; sRNA, small RNA ; tmRNA, transfer-message RNA. genome (Wassarman et al., 1999). Recently, Wassarman et al. (2001) used sequence conservation coupled with micro-array expression analysis to identify 17 new non- coding RNAs in E. coli. Argaman et al. (2001) used sequence conservation coupled with promoter and ρ- independent terminator prediction and identified 14 new non-coding RNAs in the same species. Moreover, Rivas et al. (2001) predicted 275 candidates for structural RNA loci in E. coli using computational analysis and empirically found that at least 11 loci express small, apparently non-coding RNAs. Extensive experimental characterization of the new loci is now under way. Signal recognition particle (SRP) RNA is involved in the translocation of proteins across the endoplasmic reti- culum (Stroud & Walter, 1999). SRP RNAs are ubiqui- tous in nature and contain evolutionarily conserved structural features (Poritz et al., 1988 ; Gorodkin et al., 2001). The 45S RNA of E. coli and the small cytoplasmic RNA (scRNA) of Bacillus subtilis belong to a family of SRP RNAs that function in protein secretion as an integrated component of the SRP-like particle along with an Ffh protein that is a homologue of one of the mammalian SRP components, SRP54 (Bernstein et al., 0002-5508  2002 SGM 2591