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BBA - Gene Regulatory Mechanisms (v.1819, #8)
Expression dynamics of microRNA biogenesis during preimplantation mouse development by Garcia-Lopez Jesús García-López; Jesús del Mazo (pp. 847-854).
The role of microRNAs (miRNAs) in early development, and particularly in the post-transcriptional regulation of maternal mRNAs remains controversial. Hence, we have assessed how miRNA processing is regulated during preimplantation mouse development, from the fully-grown oocyte to the blastocyst, quantifying the expression of genes whose proteins are involved in miRNAs biogenesis and function. The expression of the Drosha, Dgcr8, Exportin 5, Dicer, Ago1, Ago2, Ago3, Ago4 and Ago5 genes was downregulated from the zygotic cleavage stage, except for the increase of Ago1, Ago3 and Ago4 expression in the 2-cell embryo, and of Ago2 in 4- and 8-cell embryos. These findings suggest that the capacity to process miRNAs, by the considered canonical pathway, diminishes after fertilization, primarily reducing miRNA activity in the later stages of preimplantation development. However, by analyzing the different precursor and mature forms of specific miRNAs that are abundantly expressed in the blastocyst, such as miR-292-3p and miR-292-5p, we identified miRNA-duplexes and/or miRNAs bound to target mRNAs that may serve as potential stockpiles of miRNAs. In response to the demand, such stockpile could directly provide functional and mature miRNAs.► Genes involved in miRNA biogenesis are downregulated from the oocyte to blastocyst stages. ► However, Ago2 gene was highly upregulated in 4- and 8-cell embryos. ► Mature miRNAs could be recycled from miRNA duplex and miRNA bound to target mRNAs.
Keywords: MicroRNA; miRNA; Preimplantation embryo; Biogenesis; Oocyte; Zygote
Binding of FoxM1 to G2/M gene promoters is dependent upon B-Myb by Christin F. Down; Julie Millour; Eric W.-F. Lam; Roger J. Watson (pp. 855-862).
The promoters of genes which regulate entry into and progress through mitosis are typically induced maximally in G2 by transcription factors that include B-Myb and FoxM1. As FoxM1 gene transcription is a target of B-Myb, we investigated in this study how these transcription factors functionally interact to regulate these G2/M genes. Using a 3T3 cell line containing floxed B-myb alleles ( B-myb F/F) that could be conditionally deleted by Cre recombinase, we confirmed that B-myb knockout caused both decreased mRNA expression of several G2/M genes, including FoxM1, and delayed entry into mitosis. Although FoxM1 protein expression was actually unaffected by B-myb knockout when quiescent B-myb F/F 3T3 cells re-entered the cell cycle upon serum-stimulation, chromatin immunoprecipitation revealed that FoxM1 binding to G2/M promoters was substantially reduced. FoxM1 transcriptional activity requires sequential phosphorylation by Cyclin-dependent kinases and Plk1, which are B-Myb target genes, and we found that phosphorylation at Plk1-specific sites was somewhat reduced upon B-myb knockout. Neither this effect nor nuclear accumulation of FoxM1, which was unaffected by B-myb knockout, was sufficient to account for the dependence on B-Myb for FoxM1 promoter binding, however. More significantly, assays using paired Birc5 ( survivin) promoter-luciferase reporters with either wild-type or mutated Myb binding sites showed that FoxM1 was unable to bind and activate the promoter in the absence of B-Myb binding. Our data suggest that B-Myb is required as a pioneer factor to enable FoxM1 binding to G2/M gene promoters and explains how these transcription factors may collaborate to induce mitosis.► B-myb knockout in a 3T3 cell line decreased G2/M gene expression and delayed mitosis. ► Following B-myb knockout, FoxM1 binding to G2/M promoters was substantially reduced. ► FoxM1 was unable to bind to a Birc5 promoter if B-Myb binding sites were mutated. ► Our data suggest B-Myb acts as a pioneer factor to enable FoxM1 to induce mitosis.
Keywords: B-Myb (; MybL2; ); FoxM1; Cell cycle; Mitosis; Pioneer transcription factor
Evolution goes GAGA: GAGA binding proteins across kingdoms by Nathalie Berger; Bertrand Dubreucq (pp. 863-868).
Chromatin-associated proteins (CAP) play a crucial role in the regulation of gene expression and development in higher organisms. They are involved in the control of chromatin structure and dynamics. CAP have been extensively studied over the past years and are classified into two major groups: enzymes that modify histone stability and organization by post-translational modification of histone N-Terminal tails; and proteins that use ATP hydrolysis to modify chromatin structure. All of these proteins show a relatively high degree of sequence conservation across the animal and plant kingdoms. The essential Drosophila melanogaster GAGA factor (dGAF) interacts with these two types of CAP to regulate homeobox genes and thus contributes to a wide range of developmental events. Surprisingly, however, it is not conserved in plants. In this review, following an overview of fly GAF functions, we discuss the role of plant BBR/BPC proteins. These appear to functionally converge with dGAF despite a completely divergent amino acid sequence. Some suggestions are given for further investigation into the function of BPC proteins in plants.► Chromatin-associated proteins (CAP) play a key role in the regulation of gene expression. ► CAP are relatively well conserved across the animal and plant kingdoms. ► The Drosophila GAGA factor (dGAF) interacts with CAP but is not conserved in plants. ► Plant BBR/BPC proteins functionally converge with dGAF despite divergent sequences.
Keywords: GAGA factor; Chromatin; Plant; BPC protein
GATA-4/-6 and HNF-1/-4 families of transcription factors control the transcriptional regulation of the murine Muc5ac mucin during stomach development and in epithelial cancer cells by Nicolas Jonckheere; Audrey Vincent; Hélène Franquet-Ansart; Janneke Witte-Bouma; Anita Korteland-van Male; Emmanuelle Leteurtre; Ingrid B. Renes; Isabelle Van Seuningen (pp. 869-876).
During human embryonic and fetal development of the gastrointestinal tract, the gene encoding the MUC5AC mucin has a spatio–temporal pattern of expression restricted to the stomach. In order to better understand the molecular mechanisms responsible for this restricted pattern of expression, we have studied Muc5ac expression in the developing stomach of the mouse and correlated it to that of transcription factors known to be involved in cell differentiation programs during development. Our results indicate that GATA-6 and HNF-4α expression increased concomitantly with the induction of Muc5ac expression in embryonic stomach. We then studied Muc5ac transcriptional regulation by these transcription factors and showed that they all transactivate Muc5ac promoter. We also identified several active GATA-4/-5/-6 and HNF-1/-4 cis-elements using gel shift assays, chromatin immunoprecipitation and site-directed mutagenesis. Among all Muc5ac regulators, only GATA-6 and HNF-4 a expression was concomitant to that of Muc5ac in the developing stomach. This is thus in favor of an important role for these two transcription factors as regulators of expression of the Muc5ac mucin during stomach development and in epithelial cancer cells.► Muc5ac, GATA-6 and HNF-4α are expressed concomitantly in the developing mouse stomach. ► GATA-4/-5/-6 and HNF-1α/-1β/-4α/-4γ TF promote Muc5ac transcription. ► Muc5ac promoter contains one active GATA cis-element. ► Two active HNF cis-elements were identified within Muc5ac promoter. ► HNF and GATA TF are regulators of Muc5ac mucin expression in the developing stomach.
Keywords: Abbreviations; Bp; basepair; ED; embryonic day; EMSA; electrophoretic mobility shift assay; HNF; hepatocyte nuclear factor; kb; kilobase; PCR; polymerase chain reaction; PND; post natal day; RT; reverse transcriptase; TF; transcription factor Muc5ac; Stomach; Transcription; Development; HNF; GATA
Nicotinamide treatment reduces the levels of histone H3K4 trimethylation in the promoter of the mper1 circadian clock gene and blocks the ability of dexamethasone to induce the acute response by M. Xydous; K.E. Sekeri-Pataryas; A. Prombona; T.G. Sourlingas (pp. 877-884).
Circadian rhythms, which measure time on a scale of 24h, are generated by one of the most ubiquitous endogenous mechanisms, the circadian clock. SIRT1, a class III histone deacetylase, and PARP-1, a poly(ADP-ribose) polymerase, are two NAD+-dependent enzymes that have been shown to be involved in the regulation of the clock. Here we present evidence that the metabolite nicotinamide, an inhibitor of SIRT1, PARP-1 and mono(ADP-ribosyl) transferases, blocks the ability of dexamethasone to induce the acute response of the circadian clock gene, mper1, while it concomitantly reduces the levels of histone H3 trimethylation of lysine 4 (H3K4me3) in the mper1 promoter. Moreover, application of alternative inhibitors of SIRT1 and ADP-ribosylation did not lead to similar results. Therefore, inhibition of these enzymes does not seem to be the mode by which NAM exerts these effects. These results suggest the presence of a novel mechanism, not previously documented, by which NAM can alter gene expression levels via changes in the histone H3K4 trimethylation state.► We study SIRT1's role in the dexamethasone-induced acute response of mper1. ► We used two SIRT1 inhibitors, nicotinamide (NAM) and sirtinol. ► NAM attenuates the acute response, while sirtinol doesn't affect it. ► NAM's ability to inhibit either SIRT1 or PARP-1 is not involved in this effect. ► We report for the first time that NAM reduces H3K4me3 levels in the mper1 promoter.
Keywords: Abbreviations; dex; dexamethasone; NAM; nicotinamide; TSA; trichostatin A; CHX; cycloheximide; SIRT1; sirtuin (silent mating type information regulator 2 homolog 1); PARP-1; poly(ADP-ribose) polymerase 1; H3K4me3; histone H3 trimethylated at lysine residue 4; SAMe; S-adenosyl-methionine; SAH; S-adenosyl-homocysteineCircadian; mper1; Acute response; Dexamethasone; Nicotinamide; Histone H3 lysine 4 trimethylation
Ldb1 regulates carbonic anhydrase 1 during erythroid differentiation by Sang-Hyun Song; AeRi Kim; Ryan Dale; Ann Dean (pp. 885-891).
Carbonic anhydrase 1 (Car1), an early specific marker of the erythroid differentiation, has been used to distinguish fetal and adult erythroid cells since its production closely follows the γ- to β-globin transition, but the molecular mechanism underlying transcriptional regulation of Car1 is unclear. Here, we show that Car1 mRNA decreases significantly when erythroid differentiation is induced in MEL cells. The Ldb1 protein complex including GATA1/SCL/LMO2 binds to the Car1 promoter in uninduced cells and reduced enrichment of the complex during differentiation correlates with loss of Car1 expression. Knockdown of Ldb1 results in a reduction of Ser2 phosphorylated RNA Pol II and Cdk9 at the Car1 promoter region, suggesting that Ldb1 is required for recruitment of Pol II as well as the transcription regulator P-TEFb to enhance elongation of Car1 transcripts. Taken together, these data show that Ldb1 forms a regulatory complex to maintain Car1 expression in erythroid cells.► Ldb1 is essential for embryonic and definitive erythropoiesis. ► The Ldb1/GATA1/SCL/LMO2 complex directly regulates transcription of Car1. ► Ldb1 is required to recruit the Ldb1 complex to the promoter of the Car1 gene.
Keywords: Abbreviations; CA; carbonic anhydrase; Epb4.2; erythrocyte membrane protein band 4.2; Gypa; glycophorin A; DMSO; dimethyl sulfoxide; qPCR; quantitative real-time polymerase chain reactionLdb1; Car1; Car2; Erythroid differentiation
Ascending the nucleosome face: Recognition and function of structured domains in the histone H2A–H2B dimer by John J. Wyrick; McKenna N.M. Kyriss; William B. Davis (pp. 892-901).
Research over the past decade has greatly expanded our understanding of the nucleosome's role as a dynamic hub that is specifically recognized by many regulatory proteins involved in transcription, silencing, replication, repair, and chromosome segregation. While many of these nucleosome interactions are mediated by post-translational modifications in the disordered histone tails, it is becoming increasingly apparent that structured regions of the nucleosome, including the histone fold domains, are also recognized by numerous regulatory proteins. This review will focus on the recognition of structured domains in the histone H2A–H2B dimer, including the acidic patch, the H2A docking domain, the H2B α3-αC helices, and the HAR/HBR domains, and will survey the known biological functions of histone residues within these domains. Novel post-translational modifications and trans-histone regulatory pathways involving structured regions of the H2A–H2B dimer will be highlighted, along with the role of intrinsic disorder in the recognition of structured nucleosome regions.► Reviews protein recognition of structural features in the histone H2A-H2B dimer. ► Describes novel post-translational modifications and trans-histone regulatory pathways involving the histone H2A-H2B dimer. ► Discusses the role of intrinsically disordered regions in recognizing structured histone regions.
Keywords: Histone H2A and H2B; Acidic patch; Nucleosome recognition; Sir3; Post-translational modification
Efficient metal-specific transcription activation by Drosophila MTF-1 requires conserved cysteine residues in the carboxy-terminal domain by Sharon K. Marr; Katie L. Pennington; Michael T. Marr (pp. 902-912).
MTF-1 is a sequence-specific DNA binding protein that activates the transcription of metal responsive genes. The extent of activation is dependent on the nature of the metal challenge. Here we identify separate regions within the Drosophila MTF-1 (dMTF-1) protein that are required for efficient copper- versus cadmium-induced transcription. dMTF-1 contains a number of potential metal binding regions that might allow metal discrimination including a DNA binding domain containing six zinc fingers and a highly conserved cysteine-rich C-terminus. We find that four of the zinc fingers in the DNA binding domain are essential for function but the DNA binding domain does not contribute to the metal discrimination by dMTF-1. We find that the conserved C-terminus of the cysteine-rich domain provides cadmium specificity while copper specificity maps to the previously described copper-binding region (Chen et al.). In addition, both metal specific domains are autorepressive in the absence of metal and contribute to the low level of basal transcription from metal inducible promoters.► Separate domains of dMTF-1 confer metal specificity. ► The metal-binding domains are autorepressive in the absence of metal. ► Mutations in the conserved CTD affect cadmium- but not copper-induced transcription. ► Mutations in the copper cluster have little effect on cadmium-induced transcription. ► Metal specific conformational changes of dMTF-1
Keywords: MTF-1; Transcription; Coactivator; Metal; Conformation; Metallothionein