Functional annotation of human long noncoding RNAs via molecular phenotyping.

Long noncoding RNAs (lncRNAs) constitute the majority of transcripts in the mammalian genomes, and yet, their functions remain largely unknown. As part of the FANTOM6 project, we systematically knocked down the expression of 285 lncRNAs in human dermal fibroblasts and quantified cellular growth, morphological changes, and transcriptomic responses using Capped Analysis of Gene Expression (CAGE). Antisense oligonucleotides targeting the same lncRNAs exhibited global concordance, and the molecular phenotype, measured by CAGE, recapitulated the observed cellular phenotypes while providing additional insights on the affected genes and pathways. Here, we disseminate the largest-to-date lncRNA knockdown data set with molecular phenotyping (over 1000 CAGE deep-sequencing libraries) for further exploration and highlight functional roles for ZNF213-AS1 and lnc-KHDC3L-2.

Streptomyces rare codon UUA: from features associated with 2 adpA related locations to candidate phage regulatory translational bypassing.

In Streptomyces species, the cell cycle involves a switch from an early and vegetative state to a later phase where secondary products including antibiotics are synthesized, aerial hyphae form and sporulation occurs. AdpA, which has two domains, activates the expression of numerous genes involved in the switch from the vegetative growth phase. The adpA mRNA of many Streptomyces species has a UUA codon in a linker region between 5' sequence encoding one domain and 3' sequence encoding its other and C-terminal domain. UUA codons are exceptionally rare in Streptomyces, and its functional cognate tRNA is not present in a fully modified and acylated form, in the early and vegetative phase of the cell cycle though it is aminoacylated later. Here, we report candidate recoding signals that may influence decoding of the linker region UUA. Additionally, a short ORF 5' of the main ORF has been identified with a GUG at, or near, its 5' end and an in-frame UUA near its 3' end. The latter is commonly 5 nucleotides 5' of the main ORF start. Ribosome profiling data show translation of that 5' region. Ten years ago, UUA-mediated translational bypassing was proposed as a sensor by a Streptomyces phage of its host's cell cycle stage and an effector of its lytic/lysogeny switch. We provide the first experimental evidence supportive of this proposal.

Two Cobalt Chelatase Subunits Can Be Generated from a Single chlD Gene via Programed Frameshifting.

Magnesium chelatase chlIDH and cobalt chelatase cobNST enzymes are required for biosynthesis of (bacterio)chlorophyll and cobalamin (vitamin B12), respectively. Each enzyme consists of large, medium, and small subunits. Structural and primary sequence similarities indicate common evolutionary origin of the corresponding subunits. It has been reported earlier that some of vitamin B12 synthesizing organisms utilized unusual cobalt chelatase enzyme consisting of a large cobalt chelatase subunit (cobN) along with a medium (chlD) and a small (chlI) subunits of magnesium chelatase. In attempt to understand the nature of this phenomenon, we analyzed >1,200 diverse genomes of cobalamin and/or chlorophyll producing prokaryotes. We found that, surprisingly, genomes of many cobalamin producers contained cobN and chlD genes only; a small subunit gene was absent. Further on, we have discovered a diverse group of chlD genes with functional programed ribosomal frameshifting signals. Given a high similarity between the small subunit and the N-terminal part of the medium subunit, we proposed that programed translational frameshifting may allow chlD mRNA to produce both subunits. Indeed, in genomes where genes for small subunits were absent, we observed statistically significant enrichment of programed frameshifting signals in chlD genes. Interestingly, the details of the frameshifting mechanisms producing small and medium subunits from a single chlD gene could be prokaryotic taxa specific. All over, this programed frameshifting phenomenon was observed to be highly conserved and present in both bacteria and archaea.

Prediction of lncRNAs and their interactions with nucleic acids: benchmarking bioinformatics tools.

The genomes of mammalian species are pervasively transcribed producing as many noncoding as protein-coding RNAs. There is a growing body of evidence supporting their functional role. Long noncoding RNA (lncRNA) can bind both nucleic acids and proteins through several mechanisms. A reliable computational prediction of the most probable mechanism of lncRNA interaction can facilitate experimental validation of its function. In this study, we benchmarked computational tools capable to discriminate lncRNA from mRNA and predict lncRNA interactions with other nucleic acids. We assessed the performance of 9 tools for distinguishing protein-coding from noncoding RNAs, as well as 19 tools for prediction of RNA-RNA and RNA-DNA interactions. Our conclusions about the considered tools were based on their performances on the entire genome/transcriptome level, as it is the most common task nowadays. We found that FEELnc and CPAT distinguish between coding and noncoding mammalian transcripts in the most accurate manner. ASSA, RIBlast and LASTAL, as well as Triplexator, turned out to be the best predictors of RNA-RNA and RNA-DNA interactions, respectively. We showed that the normalization of the predicted interaction strength to the transcript length and GC content may improve the accuracy of inferring RNA interactions. Yet, all the current tools have difficulties to make accurate predictions of short-trans RNA-RNA interactions-stretches of sparse contacts. All over, there is still room for improvement in each category, especially for predictions of RNA interactions.

Genome sequences for five strains of the emerging pathogen Haemophilus haemolyticus.

We report the first whole-genome sequences for five strains, two carried and three pathogenic, of the emerging pathogen Haemophilus haemolyticus. Preliminary analyses indicate that these genome sequences encode markers that distinguish H. haemolyticus from its closest Haemophilus relatives and provide clues to the identity of its virulence factors.