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Regulation of gene expression in hepatic cells by the mammalian Target of Rapamycin (mTOR).

Author(s) : Jimenez R, Lee J, Francesconi M, Castellani G, Neretti N, Sanders J, Sedivy J, Gruppuso P,
Journal : PLoS One
BACKGROUND: We investigated mTOR regulation of gene expression by studying rapamycin effect in two hepatic cell lines, the non-tumorigenic WB-F344 cells and the tumorigenic WB311 cells. The latter are resistant to the growth inhibitory effects of rapamycin, thus providing us with an opportunity to study the gene expression effects of rapamycin without confounding effects on cell proliferation. METHODOLOGY/PRINCIPAL FINDINGS: The hepatic cells were exposed torapamycin for 24 hr. Microarray analysis on total RNA preparations identified genes that were affected by rapamycin in both cell lines and, therefore, modulated independent of growth arrest. Further studies showed that the promoterregions of these genes included E-box-containing transcription factor binding sites at higher than expected rates. Based on this, we tested the hypothesis that c-Myc is involved in regulation of gene expression by mTOR by comparing genes altered by rapamycin in the hepatic cells and by c-Myc induction in fibroblasts engineered to express c-myc in an inducible manner. Results showed enrichment for c-Myc targets among rapamycin sensitive genes in both hepatic cell lines. However, microarray analyses on wild type and c-myc null fibroblasts showed similar rapamycin effect, with the set of rapamycin-sensitive genes being enriched for c-Myc targets in both cases. CONCLUSIONS/SIGNIFICANCE: There is considerable overlap in the regulation of gene expression by mTOR and c-Myc. However, regulation of gene expression through mTOR is c-Myc-independent and cannot be attributed to the involvement of specific transcription factors regulated by the rapamycin-sensitive mTOR Complex 1.

Retinoic acid signaling targets Hox genes during the amphioxus gastrula stage: insights into early anterior-posterior patterning of the chordate body plan.

Author(s) : Koop D, Holland N, Semon M, Alvarez S, de Lera A, Laudet V, Holland L, Schubert M,
Journal : Dev Biol
Previous studies of vertebrate development have shown that retinoic acid (RA) signaling at the gastrula stage strongly influences anterior-posterior (A-P) patterning of the neurula and later stages. However, much less is known about the more immediate effects of RA signaling on gene transcription and developmental patterning at the gastrula stage. To investigate the targets of RA signaling during the gastrula stage, we used the basal chordate amphioxus, in which gastrulation involves very minimal tissue movements. First, we determined the effect of altered RA signaling on expression of 42 genes (encoding transcriptionfactors and components of major signaling cascades) known to be expressed in restricted domains along the A-P axis during the gastrula and early neurula stage. Of these 42 genes, the expression domains during gastrulation of only four (Hox1, Hox3, HNF3-1 and Wnt3) were spatially altered by exposure of the embryos to excess RA or to the RA antagonist BMS009. Moreover, blocking protein synthesis with puromycin before adding RA or BMS009 showed that only three of these genes (Hox1, Hox3 and HNF3-1) are direct RA targets at the gastrula stage. From these results we conclude that in the amphioxus gastrula RA signaling primarily acts via regulation of Hox transcription to establish positional identities along theA-P axis and that Hox1, Hox3, HNF3-1 and Wnt3 constitute a basal module of RA action during chordate gastrulation.

Specialization of a Drosophila capping protein essential for the protection of sperm telomeres.

Author(s) : Dubruille R, Orsi G, Delabaere L, Cortier E, Couble P, Marais G, Loppin B,
Journal : Curr Biol
BACKGROUND: A critical function of telomeres is to prevent fusion of chromosome ends by the DNA repair machinery. In Drosophila somatic cells, assembly of the protecting capping complex at telomeres notably involves the recruitment of HOAP, HP1, and their recently identified partner, HipHop. We previously showed that the hiphop gene was duplicated before the radiation of the melanogaster subgroup of species, giving birth to K81, a unique paternal effect gene specifically expressed in the male germline. RESULTS: Here we show that K81 specifically associates with telomeres during spermiogenesis, along with HOAP and HP1, and isretained on paternal chromosomes until zygote formation. In K81 mutant testes, capping proteins are not maintained at telomeres in differentiating spermatids, resulting in the transmission of uncapped paternal chromosomes that fail to properly divide during the first zygotic mitosis. Despite the apparent similar capping roles of K81 and HipHop in their respective domain of expression, we demonstrate by in vivo reciprocal complementation analyses that they are not interchangeable. Strikingly, HipHop appeared to be unable to maintain capping proteins at telomeres during the global chromatin remodeling of spermatid nuclei. CONCLUSIONS: Our data demonstrate that K81 is essential for the maintenance of capping proteins at telomeres in postmeiotic male germ cells. In species of the melanogaster subgroup, HipHop and K81 have not only acquired complementary expression domains, they have also functionally diverged following the gene duplication event. We propose that K81 specialized in the maintenance of telomere protection in the highly peculiar chromatin environment of differentiating male gametes.

Splicing factor and exon profiling across human tissues

Author(s) : de la Grange P, Gratadou L, Delord M, Dutertre M, Auboeuf D,
Journal : Nucleic Acids Res

Splicing factor Spf30 assists exosome-mediated gene silencing in fission yeast.

Author(s) : Bernard P, Drogat J, Dheur S, Genier S, Javerzat J,
Journal : Mol Cell Biol
Heterochromatin assembly in fission yeast relies on the processing of cognate noncoding RNAs by both the RNA interference and the exosome degradation pathways. Recent evidence indicates that splicing factors facilitate the cotranscriptionalprocessing of centromeric transcripts into small interfering RNAs (siRNAs). In contrast, how the exosome contributes to heterochromatin assembly and whether italso relies upon splicing factors were unknown. We provide here evidence that fission yeast Spf30 is a splicing factor involved in the exosome pathway of heterochromatin silencing. Spf30 and Dis3, the main exosome RNase, colocalize atcentromeric heterochromatin and euchromatic genes. At the centromeres, Dis3 helps recruiting Spf30, whose deficiency phenocopies the dis3-54 mutant: heterochromatin is impaired, as evidenced by reduced silencing and the accumulation of polyadenylated centromeric transcripts, but the production of siRNAs appears to be unaffected. Consistent with a direct role, Spf30 binds centromeric transcripts and locates at the centromeres in an RNA-dependent manner. We propose that Spf30, bound to nascent centromeric transcripts, perhapswith other splicing factors, assists their processing by the exosome. Splicing factor intercession may thus be a common feature of gene silencing pathways.

Synthesis, spectroscopic and DNA alkylating properties of malondialdehyde (MDA) bis-imine fluorescent adducts.

Author(s) : Meguellati K, Spichty M, Ladame S,
Journal : Mol Biosyst
The synthesis of a series of malondialdehyde (MDA) fluorescent adducts that mimic the well-known pentamethine cyanine dyes is reported. This new subclass of bis-imino dyes shares some common spectroscopic properties with their polymethine analogues although absorbing at significantly shorter wavelengths. A small library of trimethine and pentamethine cyanine dye bis-imino analogues have beensynthesised and characterised that cover a spectral range from blue to orange. Of particular interest is their capacity to act as mono- and bis-alkylating agents of nucleosides in general and of cytidine (and 2'-deoxycytidine) in particular.

Tax gene expression and cell cycling but not cell death are selected during HTLV-1 infection in vivo.

Author(s) : Zane L, Sibon D, Jeannin L, Zandecki M, Delfau-Larue M, Gessain A, Gout O, Pinatel C, Lancon A, Mortreux F, Wattel E,
Journal : Retrovirology
BACKGROUND: Adult T cell leukemia results from the malignant transformation of aCD4+ lymphoid clone carrying an integrated HTLV-1 provirus that has undergone several oncogenic events over a 30-60 year period of persistent clonal expansion. Both CD4+ and CD8+ lymphocytes are infected in vivo; their expansion relies on CD4+ cell cycling and on the prevention of CD8+ cell death. Cloned infected CD4+but not CD8+ T cells from patients without malignancy also add up nuclear and mitotic defects typical of genetic instability related to the expression of the virus-encoded oncogene tax. HTLV-1 expression is cancer-prone in vitro, but in vivo numerous selection forces act to maintain T cell homeostasis and are possibly involved in clonal selection. RESULTS: Here we demonstrate that the HTLV-1 associated CD4+ preleukemic phenotype and the specific patterns of CD4+ and CD8+ clonal expansion are in vivo selected processes. By comparing the effects of recent (1 month) experimental infections performed in vitro and thoseobserved in cloned T cells from patients infected for >6-26 years, we found thatin chronically HTLV-1 infected individuals, HTLV-1 positive clones are selected for tax expression. In vivo, infected CD4+ cells are positively selected for cell cycling whereas infected CD8+ cells and uninfected CD4+ cells are negatively selected for the same processes. In contrast, the known HTLV-1-dependent prevention of CD8+ T cell death pertains to both in vivo and in vitro infected cells. CONCLUSIONS: Therefore, virus-cell interactions alone are not sufficient to initiate early leukemogenesis in vivo.

Tax gene expression and cell cycling but not cell death are selected during HTLV-1 infection in vivo

Author(s) : Zane L, Sibon D, Jeannin L, Zandecki M, Delfau-Larue M, Gessain A, Gout O, Pinatel C, Lan?on A, Mortreux F, Wattel E,
Journal : Retrovirology

The 3' untranslated region of the Andes hantavirus small mRNA functionally replaces the poly(A) tail and stimulates cap-dependent translation initiation from the viral mRNA.

Author(s) : Vera-Otarola J, Soto-Rifo R, Ricci E, Ohlmann T, Darlix J, Lopez-Lastra M,
Journal : J Virol
In the process of translation of eukaryotic mRNAs, the 5' cap and the 3' poly(A)tail interact synergistically to stimulate protein synthesis. Unlike its cellular counterparts, the small mRNA (SmRNA) of Andes hantavirus (ANDV), a member of theBunyaviridae, lacks a 3' poly(A) tail. Here we report that the 3' untranslated region (3'UTR) of the ANDV SmRNA functionally replaces a poly(A) tail and synergistically stimulates cap-dependent translation initiation from the viral mRNA. Stimulation of translation by the 3'UTR of the ANDV SmRNA was found to be independent of viral proteins and of host poly(A)-binding protein.

Translationally controlled tumor protein is a conserved mitotic growth integrator in animals and plants.

Author(s) : Brioudes F, Thierry A, Chambrier P, Mollereau B, Bendahmane M,
Journal : Proc Natl Acad Sci U S A
The growth of an organism and its size determination require the tight regulation of cell proliferation and cell growth. However, the mechanisms and regulatory networks that control and integrate these processes remain poorly understood. Here, we address the biological role of Arabidopsis translationally controlled tumor protein (AtTCTP) and test its shared functions in animals and plants. The data support a role of plant AtTCTP as a positive regulator of mitotic growth byspecifically controlling the duration of the cell cycle. We show that, in contrast to animal TCTP, plant AtTCTP is not implicated in regulating postmitotic growth. Consistent with this finding, plant AtTCTP can fully rescue cell proliferation defects in Drosophila loss of function for dTCTP. Furthermore, Drosophila dTCTP is able to fully rescue cell proliferation defects in Arabidopsis tctp knockouts. Our data provide evidence that TCTP function in regulating cell division is part of a conserved growth regulatory pathway sharedbetween plants and animals. The study also suggests that, although the cell division machinery is shared in all multicellular organisms to control growth, cell expansion can be uncoupled from cell division in plants but not in animals.