7240–7251 Nucleic Acids Research, 2008, Vol. 36, No. 22 doi:10.1093/nar/gkn898 Published online 12 November 2008 Small non-coding RNAs in Streptomyces coelicolor Julia P. Swiercz1, Hindra1, Jan Bobek 1 , Henry J. Haiser1, Christina Di Berardo 1 , Brian Tjaden2 and Marie A. Elliot1,* 1Department of Biology and Institute for Infectious Disease Research, McMaster University, 1280 Main Street West, Hamilton, ON, Canada L8S 4K1 and 2Computer Science Department, Wellesley College, Wellesley, MA 02481, USA Received August 25, 2008; Revised October 23, 2008; Accepted October 24, 2008 ABSTRACT In bacteria, small RNAs (sRNAs) make important regulatory contributions to an ever increasing number of cellular processes. To expand the reper- toire of known sRNAs, we sought to identify novel sRNAs in the differentiating, multicellular bacterium Streptomyces coelicolor. We describe a combined bioinformatic and experimental approach that enabled the identification and characterization of nine novel sRNAs in S. coelicolor, including a cis- encoded antisense sRNA. We examined sRNA expression throughout the S. coelicolor develop- mental cycle, which progresses from vegetative mycelium formation, to aerial mycelium formation and finally sporulation. We further determined the effects of growth medium composition (rich versus minimal medium) on sRNA gene expression, and compared wild-type sRNA expression profiles with those of four developmental mutants. All but two of the sRNAs exhibited some degree of medium dependence, with three sRNAs being expressed exclusively during growth on one medium type. Unlike most sRNAs characterized thus far, several sRNA genes in S. coelicolor were expressed consti- tutively (apart from during late sporulation), sug- gesting a possible housekeeping role for these transcripts. Others were expressed at specific developmental stages, and their expression profiles were altered in response to developmental muta- tions. Expression of one sRNA in particular was dependent upon the sporulation-specific sigma factor pWhiG. INTRODUCTION Biological complexity can often be correlated with regula- tory complexity. It is well established that bacteria living in a highly variable environment, or having a complex life cycle, devote a large proportion of their genome to regu- latory and signaling functions (1). In the case of the soil- dwelling bacterium Streptomyces coelicolor, >10% of all protein-coding genes are predicted to have a regulatory function (2). This reﬂects not only the variable habitat of S. coelicolor, but also its intricate life cycle that involves a series of remarkable morphological and metabolic trans- formations (3). S. coelicolor is a ﬁlamentous bacterium that grows by hyphal tip extension to form a vegetative mycelium. The mycelium is then presumed to be partially canni- balized to provide nutrients for the production of second- ary metabolites (including many antibiotics) and the raising of aerial hyphae, which mature to form chains of exospores. Morphogenesis and antibiotic production are stringently controlled, and are genetically coupled (sharing some regulatory elements) (4). Whilst progress has been made in elucidating the regulatory networks gov- erning development and antibiotic production, many of the components involved in these networks have yet to be identiﬁed. The cellular regulatory arsenal comprises not only regulatory proteins, but also non-coding RNAs, or small RNAs (sRNAs) as they are commonly referred to in bac- teria because of their small size [50–500 nucleotides (nt)]. sRNAs exert their regulatory inﬂuence through inter- actions with mRNAs (modulating transcript stability or translation eﬃciency) or proteins (aﬀecting protein activity). In Escherichia coli, sRNAs have roles in processes ranging from the control of outer membrane protein expression to the sequestration of sigma factors (5–8). sRNAs have been further implicated in sporulation in Bacillus subtilis (9), quorum sensing in Vibrio harveyi and V. cholerae (10), life cycle diﬀerentiation in Chlamydia trachomatis (11) and virulence in Shigella dysenteriae (12). Despite the obvious importance of sRNAs as regulators, their relative abundance across the bacteria has not yet been fully elucidated, and it is likely that they have an important regulatory function in Streptomyces. *To whom correspondence should be addressed. Tel: +1 905 525 9140 (ext. 24225); Fax: +1 905 522 6066; Email: melliot@mcmaster.ca The authors wish it to be known that, in their opinion, the first two authors should be regarded as joint First Authors. 2008 The Author(s) This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/ by-nc/2.0/uk/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.