Detection of 50- and 30-UTR-derived small RNAs and cis-encoded antisense RNAs in Escherichia coli (original) (raw)

Detection of 5'- and 3'-UTR-derived small RNAs and cis-encoded antisense RNAs in Escherichia coli

Nucleic Acids Research, 2005

Evidence is accumulating that small, noncoding RNAs are important regulatory molecules. Computational and experimental searches have led to the identification of 60 small RNA genes in Escherichia coli. However, most of these studies focused on the intergenic regions and assumed that small RNAs were .50 nt. Thus, the previous screens missed small RNAs encoded on the antisense strand of protein-coding genes and small RNAs of ,50 nt. To identify additional small RNAs, we carried out a cloning-based screen focused on RNAs of 30-65 nt. In this screen, we identified RNA species corresponding to fragments of rRNAs, tRNAs and known small RNAs. Several of the small RNAs also corresponded to 5 0-and 3 0-untranslated regions (UTRs) and internal fragments of mRNAs. Four of the 3 0-UTR-derived RNAs were highly abundant and two showed expression patterns that differed from the corresponding mRNAs, suggesting independent functions for the 3 0-UTR-derived small RNAs. We also detected three previously unidentified RNAs encoded in intergenic regions and RNAs from the long direct repeat and hok/sok elements. In addition, we identified a few small RNAs that are expressed opposite protein-coding genes and could base pair with 5 0 or 3 0 ends of the mRNAs with perfect complementarity.

RNAs and cis-encoded antisense RNAs in Escherichia coli

2004

Evidence is accumulating that small, noncoding RNAs are important regulatory molecules. Computational and experimental searches have led to the identification of 60 small RNA genes in Escherichia coli. However, most of these studies focused on the intergenic regions and assumed that small RNAs were.50 nt. Thus, the previous screens missed small RNAs encoded on the antisense strand of protein-coding genes and small RNAs of,50 nt. To identify additional small RNAs, we carried out a cloning-based screen focused on RNAs of 30–65 nt. In this screen, we identified RNA species corresponding to fragments of rRNAs, tRNAs and known small RNAs. Several of the small RNAs also corresponded to 50- and 30-untranslated regions (UTRs) and internal fragments of mRNAs. Four of the 30-UTR-derived RNAs were highly abundant and two showed expression patterns that differed from the corresponding mRNAs, suggesting independent functions for the 30-UTR-derived small RNAs. We also detected three previously un...

A survey of small RNA-encoding genes in Escherichia coli

Nucleic Acids Research, 2003

Small RNA (sRNA) molecules have gained much interest lately, as recent genome-wide studies have shown that they are widespread in a variety of organisms. The relatively small family of 10 known sRNA-encoding genes in Escherichia coli has been signi®cantly expanded during the past two years with the discovery of 45 novel genes. Most of these genes are still uncharacterized and their cellular roles are unknown. In this survey we examined the sequence and genomic features of the 55 currently known sRNA-encoding genes in E.coli, attempting to identify their common characteristics. Such characterization is important for both expanding our understanding of this unique gene family and for improving the methods to predict and identify sRNA-encoding genes based on genomic information.

Novel small RNA-encoding genes in the intergenic regions of Escherichia coli

Current Biology, 2001

To search the Escherichia coli genome sequence for genes hanah@md2.huji.ac.il encoding small RNAs, we developed a computational strategy employing gerhart.wagner@icm.uu.se transcription signals and genomic features of the known small RNA-encoding genes. The search, for which we used rather restrictive criteria, has led ‡ These authors contributed equally to this work.

Small Regulatory RNAs in Bacteria

Nucleic Acids and Molecular Biology, 2006

In recent years, small regulatory RNAs have been discovered at a staggering rate both in prokaryotes and eukaryotes. By now it is clear that post-transcriptional regulation of gene expression mediated by such RNAs is the rule rather than-as previously believed-the exception. In this chapter, we focus on small RNAs (sRNAs) encoded by bacterial chromosomes. The strategies for their discovery, their biological roles, and their mechanisms of action are discussed. Even though the number of well-characterized sR-NAs in, for example, the best studied model enterobacterium Escherichia coli,i ss t i l l small, the emerging pattern suggests that antisense mechanisms predominate. In terms of their roles in bacterial physiology, most of these RNAs appear to be involved in stress response regulation. Some other examples indicate functions in regulation of virulence. Two aspects of sRNA-mediated control arising from recent observations are addressed as well. Firstly, some sRNAs need proteins (notably Hfq) as helpers in their antisense activities-at this point the reason for this requirement is not understood. Secondly, only limited sequence complementarity is generally observed in antisense-target RNA pairs. This raises the fundamental question of how specific recognition is accomplished, and what the structure/sequence determinants for rapid and productive interaction are. 2 E.G.H. Wagner · F. Darfeuille

Identification of bacterial small non-coding RNAs: experimental approaches

Current Opinion in Microbiology, 2007

Almost 140 bacterial small RNAs (sRNAs; sometimes referred to as non-coding RNAs) have been discovered in the past six years. The majority of these sRNAs were discovered in Escherichia coli, and a smaller subset was characterized in other bacteria, many of which were pathogenic. Many of these genes were identified as a result of systematic screens using computational prediction of sRNAs and experimental-based approaches, including microarray and shotgun cloning. A smaller number of sRNAs were discovered by direct labeling or by functional genetic screens. Many of the discovered genes, ranging in size from 50 to 500 nucleotides, are conserved and located in intergenic regions, in-between open reading frames. The expression of many of these genes is growth phase dependent or stress related. As each search employed specific parameters, this led to the identification of genes with distinct characteristics. Consequently, unique sRNAs such as those that are species-specific, sRNA genes that are transcribed under unique conditions or genes located on the antisense strand of protein-encoding genes, were probably missed.

A bioinformatics based approach to discover small RNA genes in the Escherichia coli genome

Biosystems, 2002

The recent explosion in available bacterial genome sequences has initiated the need to improve an ability to annotate important sequence and structural elements in a fast, efficient and accurate manner. In particular, small non-coding RNAs (sRNAs) have been difficult to predict. The sRNAs play an important number of structural, catalytic and regulatory roles in the cell. Although a few groups have recently published prediction methods for annotating sRNAs in bacterial genome, much remains to be done in this field. Toward the goal of developing an efficient method for predicting unknown sRNA genes in the completed Escherichia coli genome, we adopted a bioinformatics approach to search for DNA regions that contain a s70 promoter within a short distance of a rho-independent terminator. Among a total of 227 candidate sRNA genes initially identified, 32 were previously described sRNAs, orphan tRNAs, and partial tRNA and rRNA operons. Fifty-one are mRNAs genes encoding annotated extremely small open reading frames (ORFs) following an acceptable ribosome binding site. One hundred forty-four are potentially novel non-translatable sRNA genes. Using total RNA isolated from E. coli MG1655 cells grown under four different conditions, we verified transcripts of some of the genes by Northern hybridization. Here we summarize our data and discuss the rules and advantages/disadvantages of using this approach in annotating sRNA genes on bacterial genomes.

Comparative Genomics of Small RNAs in Bacterial Genomes

OMICS: A Journal of Integrative Biology, 2007

In recent years, various families of small non-coding RNAs (sRNAs) have been discovered by experimental and computational approaches, both in bacterial and eukaryotic genomes. Although most of them await elucidation of their function, it has been reported that some play important roles in gene regulation. Here we carried out comparative genomics analysis of possible sRNAs that are computationally identified in 30 bacterial genomes from ␥and ␣-proteobacteria and Deinococcus radiodurans. Identified sRNAs are clustered by a complete-linkage clustering method to see conservation among the organisms. On average, sRNAs are found in approximately 30% of intergenic regions of each genome sequence. Of these, 25.7% are conserved among three or more organisms. Approximately 60% of the conserved sRNAs do not locate in orthologous intergenic regions, implying that sRNAs may be shuffled their positions in genomes. The current study implies that sRNAs may be involved in a more extensive range of functions in bacteria.

A cohabiting bacterium alters the spectrum of short RNAs secreted by Escherichia coli

FEMS microbiology letters, 2018

Recently, it has been found that bacteria secrete short RNAs able to affect gene expression in eukaryotic cells, while certain mammalian microRNAs 'shape the gut microbiome' altering bacterial transcriptome. The involvement of bacterial RNAs in communication with other bacteria is also expected, but has not been documented yet. Here, we compared the fractions of extremely short (12-22 nucleotides) RNAs secreted by Escherichia coli grown in a pure culture and jointly with bacteria of the Paenibacillus genus. Besides fragments of rRNAs and tRNAs, abundant in all samples, secreted oligonucleotides (exoRNAs) predominantly contained GC-rich fragments of messenger and antisense RNAs processed from regions with stable secondary structures. They differed in composition from oligonucleotides of intracellular fraction, where fragments of small regulatory RNAs were prevalent. Both fractions contained RNAs capable to form complementary duplexes, while for exoRNA samples a higher percent...