A real-time view of the TAR:Tat:P-TEFb complex at HIV-1 transcription sites

HIV-1 transcription is tightly regulated: silent in long-term latency and highly active in acutely-infected cells. Transcription is activated by the viral protein Tat, which recruits the elongation factor P-TEFb by binding the TAR sequence present in nascent HIV-1 RNAs. In this study, we analyzed the dynamic of the TAR:Tat:P-TEFb complex in living cells, by performing FRAP experiments at HIV-1 transcription sites. Our results indicate that a large fraction of Tat present at these sites is recruited by Cyclin T1. We found that in the presence of Tat, Cdk9 remained bound to nascent HIV-1 RNAs for 71s. In contrast, when transcription was activated by PMA/ionomycin, in the absence of Tat, Cdk9 turned-over rapidly and resided on the HIV-1 promoter for only 11s. Thus, the mechanism of trans-activation determines the residency time of P-TEFb at the HIV-1 gene, possibly explaining why Tat is such a potent transcriptional activator. In addition, we observed that Tat occupied HIV-1 transcription sites for 55s, suggesting that the TAR:Tat:P-TEFb complex dissociates from the polymerase following transcription initiation, and undergoes subsequent cycles of association/dissociation

ADAR2-mediated editing of RNA substrates in the nucleolus is inhibited by C/D small nucleolar RNAs

Posttranscriptional, site-specific adenosine to inosine (A-to-I) base conversions, designated as RNA editing, play significant roles in generating diversity of gene expression. However, little is known about how and in which cellular compartments RNA editing is controlled. Interestingly, the two enzymes that catalyze RNA editing, adenosine deaminases that act on RNA (ADAR) 1 and 2, have recently been demonstrated to dynamically associate with the nucleolus. Moreover, we have identified a brain-specific small RNA, termed MBII-52, which was predicted to function as a nucleolar C/D RNA, thereby targeting an A-to-I editing site (C-site) within the 5-HT2C serotonin receptor pre-mRNA for 2′-O-methylation. Through the subcellular targeting of minigenes that contain natural editing sites, we show that ADAR2- but not ADAR1-mediated RNA editing occurs in the nucleolus. We also demonstrate that MBII-52 forms a bona fide small nucleolar ribonucleoprotein particle that specifically decreases the efficiency of RNA editing by ADAR2 at the targeted C-site. Our data are consistent with a model in which C/D small nucleolar RNA might play a role in the regulation of RNA editing

Live cell imaging reveals 3 '-UTR dependent mRNA sorting to synapses

mRNA transport restricts translation to specific subcellular locations, which is the basis for many cellular functions. However, the precise process of mRNA sorting to synapses in neurons remains elusive. Here we use Rgs4 mRNA to investigate 3'-UTR-dependent transport by MS2 live-cell imaging. The majority of observed RNA granules display 3'-UTR independent bidirectional transport in dendrites. Importantly, the Rgs4 3'-UTR causes an anterograde transport bias, which requires the Staufen2 protein. Moreover, the 3'-UTR mediates dynamic, sustained mRNA recruitment to synapses. Visualization at high temporal resolution enables us to show mRNA patrolling dendrites, allowing transient interaction with multiple synapses, in agreement with the sushi-belt model. Modulation of neuronal activity by either chemical silencing or local glutamate uncaging regulates both the 3'-UTR-dependent transport bias and synaptic recruitment. This dynamic and reversible mRNA recruitment to active synapses would allow translation and synaptic remodeling in a spatially and temporally adaptive manner

Real-time imaging of cotranscriptional splicing reveals a kinetic model that reduces noise: implications for alternative splicing regulation

A combination of several rate-limiting steps allows for efficient control of alternative splicing

THE ANALYSIS OF THE GENETIC DETERMINANTS, DEFINING THE MICROCINES SYNTHESIS

The object of investigation: the peptide antibiotics of the enterobacteria (microcines). The work is aimed at studying the genetic determinants, connected with the synthesis of microcines in cells E. coli. The high conservativeness of the plasmide genes, determining the synthesis of the three known microcines of type B, has been shown. It has been established, that in the synthesis of microcine C51, three plasmide genes take part, for the immunity expression at least two genes are required for the microcine. The nucleotide consequence of the DNA fragment, determining the partial immunity to microcine C51, has been determined. The field of application: studying the paptide antibiotics, organisation and functioning of the specific plasmide genesAvailable from VNTIC / VNTIC - Scientific & Technical Information Centre of RussiaSIGLERURussian Federatio

Localisation des ARN dans le cytoplasme

La localisation de certains ARN dans des endroits précis du cytoplasme cellulaire a été découverte il y a une vingtaine d’années, à la fois dans l’embryon précoce d’ascidie, dans l’oeuf de drosophile et dans les fibroblastes embryonnaires de poulet. On connaît maintenant plus d’une centaine d’exemples d’ARN localisés, et cela dans de nombreuses espèces, de la levure à l’homme. Dans la plupart des cas, la localisation de ces ARN est liée à des événements de polarité cellulaire. À l’aide de deux exemples, l’ARNm Ash1 de la levure et les ARN rétroviraux murins, cet article aborde la fonction et les mécanismes de la localisation des ARN. On verra notamment que ce phénomène est impliqué dans de nombreux processus cellulaires et que le transport des ARN est intimement lié au cytosquelette et au trafic membranaire.RNA localization in subcytoplasmic areas is a process known for more than twenty years, and more than a hundred RNAs have now been shown to be spatially regulated. In most cases, RNA localization is involved in cell polarity, either by reading spatial clues and translating them into a spatial regulation of gene expression, or more directly by controlling cytoskeletal polarity. In this review, the various functions of RNA localization will be presented, and by analyzing two examples, Ash1 mRNA in yeast and retroviral genomic RNAs in mammals, the reader will be taken step by step into the detailed mechanisms of this fascinating process