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Gary S Stein - One of the best experts on this subject based on the ideXlab platform.
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higher order genomic organization and regulatory compartmentalization for cell cycle control at the g1 s phase transition
Journal of Cellular Physiology, 2018Co-Authors: Prachi N Ghule, Janet L Stein, Jane B Lian, Andre J. Van Wijnen, Andrew J Fritz, Joseph R Boyd, David J Seward, Gary S SteinAbstract:Fidelity of Histone Gene regulation, and ultimately of Histone protein biosynthesis, is obligatory for packaging of newly replicated DNA into chromatin. Control of Histone Gene expression within the 3-dimensional context of nuclear organization is reflected by two well documented observations. DNA replication-dependent Histone mRNAs are synthesized at specialized subnuclear domains designated Histone locus bodies (HLBs), in response to activation of the growth factor dependent Cyclin E/CDK2/HINFP/NPAT pathway at the G1/S transition in mammalian cells. Complete loss of the Histone Gene regulatory factors HINFP or NPAT disrupts HLB integrity that is necessary for coordinate control of DNA replication and Histone Gene transcription. Here we review the molecular Histone-related requirements for G1/S-phase progression during the cell cycle. Recently developed experimental strategies, now enable us to explore mechanisms involved in dynamic control of Histone Gene expression in the context of the temporal (cell cycle) and spatial (HLBs) remodeling of the Histone Gene loci.
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epiGenetic control of cell cycle dependent Histone Gene expression is a principal component of the abbreviated pluripotent cell cycle
Molecular and Cellular Biology, 2012Co-Authors: Ricardo F Medina, Janet L Stein, Gary S Stein, Andre J. Van Wijnen, Prachi N Ghule, Fernando Cruzat, Rasim A Barutcu, Martin MontecinoAbstract:Self-renewal of human pluripotent embryonic stem cells proceeds via an abbreviated cell cycle with a shortened G(1) phase. We examined which Genes are modulated in this abbreviated period and the epiGenetic mechanisms that control their expression. Accelerated upregulation of Genes encoding Histone proteins that support DNA replication is the most prominent Gene regulatory program at the G(1)/S-phase transition in pluripotent cells. Expedited expression of Histone Genes is mediated by a unique chromatin architecture reflected by major nuclease hypersensitive sites, atypical distribution of epiGenetic Histone marks, and a region devoid of Histone octamers. We observed remarkable differences in chromatin structure--hypersensitivity and Histone protein modifications--between human embryonic stem (hES) and normal diploid cells. Cell cycle-dependent transcription factor binding permits dynamic three-dimensional interactions between transcript initiating and processing factors at 5' and 3' regions of the Gene. Thus, progression through the abbreviated G(1) phase involves cell cycle stage-specific chromatin-remodeling events and rapid assembly of subnuclear microenvironments that activate Histone Gene transcription to promote nucleosomal packaging of newly replicated DNA during stem cell renewal.
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cyclin d2 and the cdk substrate p220npat are required for self renewal of human embryonic stem cells
Journal of Cellular Physiology, 2010Co-Authors: Klaus A Becker, Janet L Stein, Jane B Lian, Andre J. Van Wijnen, Prachi N Ghule, Gary S SteinAbstract:Self-renewal of pluripotent human embryonic stem (hES) cells utilizes an abbreviated cell cycle that bypasses E2F/pRB-dependent growth control. We investigated whether self-renewal is alternatively regulated by cyclin/CDK phosphorylation of the p220(NPAT)/HiNF-P complex to activate Histone Gene expression at the G1/S phase transition. We show that cyclin D2 is prominently expressed in pluripotent hES cells, but cyclin D1 eclipses cyclin D2 during differentiation. Depletion of cyclin D2 or p220(NPAT) causes a cell cycle defect in G1 reflected by diminished phosphorylation of p220(NPAT), decreased cell cycle dependent Histone H4 expression and reduced S phase progression. Thus, cyclin D2 and p220(NPAT) are principal cell cycle regulators that determine competency for self-renewal in pluripotent hES cells. While pRB/E2F checkpoint control is relinquished in human ES cells, fidelity of physiological regulation is secured by cyclin D2 dependent activation of the p220(NPAT)/HiNF-P mechanism that may explain perpetual proliferation of hES cells without transformation or tumoriGenesis.
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the Histone Gene cell cycle regulator hinf p is a unique zinc finger transcription factor with a novel conserved auxiliary dna binding motif
Biochemistry, 2008Co-Authors: Ricardo F Medina, Janet L Stein, Jane B Lian, Andre J. Van Wijnen, Sayyed K Zaidi, Timothy Buck, Angela Mielechamberland, Gary S SteinAbstract:Accumulation of Histone proteins is necessary for packaging of replicated DNA during the S phase of the cell cycle. Different mechanisms operate to regulate Histone protein levels, and induction of human Histone Gene expression at the G1-S phase transition plays a critical role. The zinc finger HiNF-P and coactivator p220 (NPAT) proteins are key regulators of Histone Gene expression. Here, we describe a novel HiNF-P-specific conserved region (PSCR) located within the C-terminus that is present in HiNF-P homologues of all metazoan species that have been examined. The PSCR motif is required for activation of Histone H4 Gene transcription and contributes to DNA binding of HiNF-P. Thus, the PSCR module represents an auxiliary DNA-binding determinant that plays a critical role in mediating Histone Gene expression during the cell cycle and defines HiNF-P as a unique cell cycle regulatory member of the zinc finger transcription factor family.
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cell cycle dependent phosphorylation and subnuclear organization of the Histone Gene regulator p220npat in human embryonic stem cells
Journal of Cellular Physiology, 2007Co-Authors: Prachi N Ghule, Janet L Stein, Klaus A Becker, Jane B Lian, Andre J Van Wijnen, Wade J Harper, Gary S SteinAbstract:Human embryonic stem (ES) cells have an expedited cell cycle (∼15 h) due to an abbreviated G1 phase (∼2.5 h) relative to somatic cells. One principal regulatory event during cell cycle progression is the G1/S phase induction of Histone biosynthesis to package newly replicated DNA. In somatic cells, Histone H4 Gene expression is controlled by CDK2 phosphorylation of p220NPAT and localization of HiNF-P/p220NPAT complexes with Histone Genes at Cajal body related subnuclear foci. Here we show that this ‘S point’ pathway is operative in situ in human ES cells (H9 cells; NIH-designated WA09). Immunofluorescence microscopy shows an increase in p220NPAT foci in G1 reflecting the assembly of Histone Gene regulatory complexes in situ. In contrast to somatic cells where duplication of p220NPAT foci is evident in S phase, the increase in the number of p220NPAT foci in ES cells appears to precede the onset of DNA synthesis as measured by BrdU incorporation. Phosphorylation of p220NPAT at CDK dependent epitopes is most pronounced in S phase when cells exhibit elevated levels of cyclins E and A. Our data indicate that subnuclear organization of the HiNF-P/p220NPAT pathway is rapidly established as ES cells emerge from mitosis and that p220NPAT is subsequently phosphorylated in situ. Our findings establish that the HiNF-P/p220NPAT Gene regulatory pathway operates in a cell cycle dependent microenvironment that supports expression of DNA replication-linked Histone Genes and chromatin assembly to accommodate human stem cell self-renewal. J. Cell. Physiol. 213: 9–17, 2007. © 2007 Wiley-Liss, Inc.
Robert E Maxson - One of the best experts on this subject based on the ideXlab platform.
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direct induction of dna hypermethylation in sea urchin embryos by microinjection of 5 methyl dctp stimulates early Histone Gene expression and leads to developmental arrest
Developmental Biology, 1993Co-Authors: Jeannie Chen, Robert E Maxson, Peter A JonesAbstract:Abstract The role of DNA methylation in Gene inactivation has been studied extensively in vertebrates but it is not clear whether it serves similar functions in other organisms. We devised a novel approach to induce hypermethylation of both endogenous and injected DNA in the sea urchin Lytechinus pictus in order to study the effect of DNA methylation on Gene expression in this invertebrate. By injecting 5-methyl dCTP either alone or together with a cloned DNA construct into fertilized sea urchin eggs, replicating DNA became hypermethylated from the random incorporation of the methylated nucleotide in place of cytosine during DNA synthesis. During subsequent rounds of replication, the injected 5-methyl dCTP became depleted but methylation at CpG sites was still elevated presumably due to the action of a methyltransferase enzyme. Using this approach, we studied the effect of hypermethylation on two members of the sea urchin multiGene family, the early H2B and the late H2B Genes. De novo methylation was shown to occur at known cis-regulatory regions of the Genes. The effect of methylation on Gene activity was probed using RNase protection assay. Methylation resulted in increased early H2B Histone Gene expression but had no effect on late H2B Histone Gene expression. These results demonstrate that methylation does not necessarily inactivate Genes in the sea urchins as previously thought. Interestingly, the development of embryos injected with 5-methyl dCTP typically was arrested at the blastula stage, and analysis of the genomic DNA extracted from injected embryos showed a significant increase in the endogenous methylation content. These data suggest that perturbation of methylation patterns in developing sea urchin embryos may be responsible for the developmental arrest through altering the Gene expression pattern.
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an octamer element is required for the expression of the alpha h2b Histone Gene during the early development of the sea urchin
Developmental Biology, 1992Co-Authors: Jeffrey Bell, Bharat Char, Robert E MaxsonAbstract:Abstract Early (alpha) Histone Genes are one of several Histone Gene families in the sea urchin genome. They are expressed at high levels in blastula-stage embryos and are inactivated by the early gastrula stage. By microinjecting mutant early H2B Genes into sea urchin zygotes and monitoring their transcriptional activity in blastula- and gastrula-stage embryos, we sought to identify the cis-regulatory elements responsible for this dramatic change in early H2B Gene activity. We found that deletion of DNA 5′ of −71 and 3′ of +591 did not affect the timing or magnitude of early H2B Gene expression. Neither was early H2B Gene expression affected by the replacement of sequences downstream of −36 with the corresponding region of the L1 late H2B Gene, expressed after the peak transcription of the early H2B Gene. Further deletion of early H2B promoter sequences from −71 to −56, removing a conserved octamer element, resulted in near-complete inactivation of the early H2B Gene in both blastula- and gastrula-stage embryos. Also inactivating early H2B Gene expression were an internal deletion of the octamer element and a base substitution mutation that altered its sequence. This base substitution mutation also caused a parallel reduction in the ability of the octamer element to bind a factor present in nuclear extracts of sea urchin blastulae. These data strongly suggest that the proper expression of the early H2B Gene in cleavage- and blastula-stage embryos depends on the octamer element and a factor with which it interacts.
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activation of the l1 late h2b Histone Gene in blastula stage sea urchin embryos by an antennapedia class homeoprotein
Mechanisms of Development, 1991Co-Authors: Allan Zijian Zhao, Gordon Vansant, Jeffrey R Bell, Tom Humphreys, Robert E MaxsonAbstract:The L1 late H2B Histone Gene of the sea urchin Strongylocentrotus purpuratus is transcriptionally activated in late blastula stage embryos by a mechanism that depends on an enhancer element located 3' of the Gene (Zhao et al., 1990). A protein factor, designated H2B abp 1, binds this element at a site that resembles the consensus recognition sequence of Antennapedia-class homeodomain proteins. We demonstrate here that Antennapedia (Antp) and Hbox4 proteins, members of the Antennapedia class of homeoproteins from Drosophila and sea urchin respectively, bind the L1 H2B abp 1 site, and that the Drosophila Antp protein acts through this site to trans-activate the L1 H2B Gene, in vivo. In addition, RNA gel blot analysis demonstrated that Hbox4 transcripts accumulate in developing embryos with a time course that closely resembles that of H2B adp 1 DNA binding activity and the activity and the transcription rate of the L1 late H2B Gene. Finally, we show that antibody prepared against the sea urchin Hbox4 protein, a member of the Abd-B subclass of the Antennapedia class, specifically inhibits binding of the H2B abp 1 factor to the L1 H2B enhancer, suggesting that H2B abp 1 is encoded by Hbox4 or a closely related Gene.
Janet L Stein - One of the best experts on this subject based on the ideXlab platform.
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higher order genomic organization and regulatory compartmentalization for cell cycle control at the g1 s phase transition
Journal of Cellular Physiology, 2018Co-Authors: Prachi N Ghule, Janet L Stein, Jane B Lian, Andre J. Van Wijnen, Andrew J Fritz, Joseph R Boyd, David J Seward, Gary S SteinAbstract:Fidelity of Histone Gene regulation, and ultimately of Histone protein biosynthesis, is obligatory for packaging of newly replicated DNA into chromatin. Control of Histone Gene expression within the 3-dimensional context of nuclear organization is reflected by two well documented observations. DNA replication-dependent Histone mRNAs are synthesized at specialized subnuclear domains designated Histone locus bodies (HLBs), in response to activation of the growth factor dependent Cyclin E/CDK2/HINFP/NPAT pathway at the G1/S transition in mammalian cells. Complete loss of the Histone Gene regulatory factors HINFP or NPAT disrupts HLB integrity that is necessary for coordinate control of DNA replication and Histone Gene transcription. Here we review the molecular Histone-related requirements for G1/S-phase progression during the cell cycle. Recently developed experimental strategies, now enable us to explore mechanisms involved in dynamic control of Histone Gene expression in the context of the temporal (cell cycle) and spatial (HLBs) remodeling of the Histone Gene loci.
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epiGenetic control of cell cycle dependent Histone Gene expression is a principal component of the abbreviated pluripotent cell cycle
Molecular and Cellular Biology, 2012Co-Authors: Ricardo F Medina, Janet L Stein, Gary S Stein, Andre J. Van Wijnen, Prachi N Ghule, Fernando Cruzat, Rasim A Barutcu, Martin MontecinoAbstract:Self-renewal of human pluripotent embryonic stem cells proceeds via an abbreviated cell cycle with a shortened G(1) phase. We examined which Genes are modulated in this abbreviated period and the epiGenetic mechanisms that control their expression. Accelerated upregulation of Genes encoding Histone proteins that support DNA replication is the most prominent Gene regulatory program at the G(1)/S-phase transition in pluripotent cells. Expedited expression of Histone Genes is mediated by a unique chromatin architecture reflected by major nuclease hypersensitive sites, atypical distribution of epiGenetic Histone marks, and a region devoid of Histone octamers. We observed remarkable differences in chromatin structure--hypersensitivity and Histone protein modifications--between human embryonic stem (hES) and normal diploid cells. Cell cycle-dependent transcription factor binding permits dynamic three-dimensional interactions between transcript initiating and processing factors at 5' and 3' regions of the Gene. Thus, progression through the abbreviated G(1) phase involves cell cycle stage-specific chromatin-remodeling events and rapid assembly of subnuclear microenvironments that activate Histone Gene transcription to promote nucleosomal packaging of newly replicated DNA during stem cell renewal.
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cyclin d2 and the cdk substrate p220npat are required for self renewal of human embryonic stem cells
Journal of Cellular Physiology, 2010Co-Authors: Klaus A Becker, Janet L Stein, Jane B Lian, Andre J. Van Wijnen, Prachi N Ghule, Gary S SteinAbstract:Self-renewal of pluripotent human embryonic stem (hES) cells utilizes an abbreviated cell cycle that bypasses E2F/pRB-dependent growth control. We investigated whether self-renewal is alternatively regulated by cyclin/CDK phosphorylation of the p220(NPAT)/HiNF-P complex to activate Histone Gene expression at the G1/S phase transition. We show that cyclin D2 is prominently expressed in pluripotent hES cells, but cyclin D1 eclipses cyclin D2 during differentiation. Depletion of cyclin D2 or p220(NPAT) causes a cell cycle defect in G1 reflected by diminished phosphorylation of p220(NPAT), decreased cell cycle dependent Histone H4 expression and reduced S phase progression. Thus, cyclin D2 and p220(NPAT) are principal cell cycle regulators that determine competency for self-renewal in pluripotent hES cells. While pRB/E2F checkpoint control is relinquished in human ES cells, fidelity of physiological regulation is secured by cyclin D2 dependent activation of the p220(NPAT)/HiNF-P mechanism that may explain perpetual proliferation of hES cells without transformation or tumoriGenesis.
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the Histone Gene cell cycle regulator hinf p is a unique zinc finger transcription factor with a novel conserved auxiliary dna binding motif
Biochemistry, 2008Co-Authors: Ricardo F Medina, Janet L Stein, Jane B Lian, Andre J. Van Wijnen, Sayyed K Zaidi, Timothy Buck, Angela Mielechamberland, Gary S SteinAbstract:Accumulation of Histone proteins is necessary for packaging of replicated DNA during the S phase of the cell cycle. Different mechanisms operate to regulate Histone protein levels, and induction of human Histone Gene expression at the G1-S phase transition plays a critical role. The zinc finger HiNF-P and coactivator p220 (NPAT) proteins are key regulators of Histone Gene expression. Here, we describe a novel HiNF-P-specific conserved region (PSCR) located within the C-terminus that is present in HiNF-P homologues of all metazoan species that have been examined. The PSCR motif is required for activation of Histone H4 Gene transcription and contributes to DNA binding of HiNF-P. Thus, the PSCR module represents an auxiliary DNA-binding determinant that plays a critical role in mediating Histone Gene expression during the cell cycle and defines HiNF-P as a unique cell cycle regulatory member of the zinc finger transcription factor family.
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cell cycle dependent phosphorylation and subnuclear organization of the Histone Gene regulator p220npat in human embryonic stem cells
Journal of Cellular Physiology, 2007Co-Authors: Prachi N Ghule, Janet L Stein, Klaus A Becker, Jane B Lian, Andre J Van Wijnen, Wade J Harper, Gary S SteinAbstract:Human embryonic stem (ES) cells have an expedited cell cycle (∼15 h) due to an abbreviated G1 phase (∼2.5 h) relative to somatic cells. One principal regulatory event during cell cycle progression is the G1/S phase induction of Histone biosynthesis to package newly replicated DNA. In somatic cells, Histone H4 Gene expression is controlled by CDK2 phosphorylation of p220NPAT and localization of HiNF-P/p220NPAT complexes with Histone Genes at Cajal body related subnuclear foci. Here we show that this ‘S point’ pathway is operative in situ in human ES cells (H9 cells; NIH-designated WA09). Immunofluorescence microscopy shows an increase in p220NPAT foci in G1 reflecting the assembly of Histone Gene regulatory complexes in situ. In contrast to somatic cells where duplication of p220NPAT foci is evident in S phase, the increase in the number of p220NPAT foci in ES cells appears to precede the onset of DNA synthesis as measured by BrdU incorporation. Phosphorylation of p220NPAT at CDK dependent epitopes is most pronounced in S phase when cells exhibit elevated levels of cyclins E and A. Our data indicate that subnuclear organization of the HiNF-P/p220NPAT pathway is rapidly established as ES cells emerge from mitosis and that p220NPAT is subsequently phosphorylated in situ. Our findings establish that the HiNF-P/p220NPAT Gene regulatory pathway operates in a cell cycle dependent microenvironment that supports expression of DNA replication-linked Histone Genes and chromatin assembly to accommodate human stem cell self-renewal. J. Cell. Physiol. 213: 9–17, 2007. © 2007 Wiley-Liss, Inc.
Linda D Strausbaugh - One of the best experts on this subject based on the ideXlab platform.
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drosophila virilis has atypical kinds and arrangements of Histone repeats
Chromosoma, 1998Co-Authors: John Schienman, Elena R. Lozovskaya, Linda D StrausbaughAbstract:Genomic and P1 clone DNAs of Drosophila virilis were analyzed to determine the structure and organization of Histone Genes in this species. The species contains unique and variable repeat types, in comparison with the related species Drosophila melanogaster, with quartet repeats lacking the H1 Gene and multi-length variant quintet repeats containing the H1 Gene. Unexpectedly, the H1-containing repeats are highly polymorphic in length, and thus not in a strict tandem arrangement, while the H1-less repeats are very uniform and tandemly reiterated. Despite such differences, the relative positions and transcriptional polarities of the Histone Gene subtypes of one subcloned quintet are similar to the major Histone repeat type of D. melanogaster. For the first time, the Histone H1 Gene has been shown to be associated with other Histone Gene subtypes and is present at both chromosomal loci. DNA sequence variants of the H1 Gene have been mapped to individual P1 clones and found to be in a partitioned organization. The P1 cloning system has proved useful in completely retrieving a complex repetitive locus in vitro and in examining the structure and organization of the Histone Genes of D. virilis.
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low codon bias and high rates of synonymous substitution in drosophila hydei and d melanogaster Histone Genes
Molecular Biology and Evolution, 1993Co-Authors: David H A Fitch, Linda D StrausbaughAbstract:: We have evaluated codon usage bias in Drosophila Histone Genes and have obtained the nucleotide sequence of a 5,161-bp D. hydei Histone Gene repeat unit. This repeat contains Genes for all five Histone proteins (H1, H2a, H2b, H3, and H4) and differs from the previously reported one by a second EcoRI site. These D. hydei repeats have been aligned to each other and to the 5.0-kb (i.e., long) and 4.8-kb (i.e., short) Histone repeat types from D. melanogaster. In each species, base composition at synonymous sites is similar to the average genomic composition and approaches that in the small intergenic spacers of the Histone Gene repeats. Accumulation of synonymous changes at synonymous sites after the species diverged is quite high. Both of these features are consistent with the relatively low codon usage bias observed in these Genes when compared with other Drosophila Genes. Thus, the Generalization that abundantly expressed Genes in Drosophila have high codon bias and low rates of silent substitution does not hold for the Histone Genes.
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low codon bias and high rates of synonymous substitution in drosophila hydei and d melanogaster Histone Genes
Molecular Biology and Evolution, 1993Co-Authors: David H A Fitch, Linda D StrausbaughAbstract:We have evaluated codon usage bias in Drosophila Histone Genes and have obtained the nucleotide sequence of a 5,161-bp D. hydei Histone Gene repeat unit. This repeat contains Genes for all five Histone proteins (H1, H2a, H2b, H3, and H4) and differs from the previously reported one by a second EcoRI site. These D. hydei repeats have been aligned to each other and to the 5.0-kb (i.e., long) and 4.8-kb (i.e., short) Histone repeat types from D. melanogaster. In each species, base composition at synonymous sites is similar to the average genomic composition and approaches that in the small intergenic spacers of the Histone Gene repeats. Accumulation of synonymous changes at synonymous sites after the species diverged is quite high. Both of these features are consistent with the relatively low codon usage bias observed in these Genes when compared with other Drosophila Genes. Thus, the Generalization that abundantly expressed Genes in Drosophila have high codon bias and low rates of silent substitution does not hold for the Histone Genes.
Jane B Lian - One of the best experts on this subject based on the ideXlab platform.
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higher order genomic organization and regulatory compartmentalization for cell cycle control at the g1 s phase transition
Journal of Cellular Physiology, 2018Co-Authors: Prachi N Ghule, Janet L Stein, Jane B Lian, Andre J. Van Wijnen, Andrew J Fritz, Joseph R Boyd, David J Seward, Gary S SteinAbstract:Fidelity of Histone Gene regulation, and ultimately of Histone protein biosynthesis, is obligatory for packaging of newly replicated DNA into chromatin. Control of Histone Gene expression within the 3-dimensional context of nuclear organization is reflected by two well documented observations. DNA replication-dependent Histone mRNAs are synthesized at specialized subnuclear domains designated Histone locus bodies (HLBs), in response to activation of the growth factor dependent Cyclin E/CDK2/HINFP/NPAT pathway at the G1/S transition in mammalian cells. Complete loss of the Histone Gene regulatory factors HINFP or NPAT disrupts HLB integrity that is necessary for coordinate control of DNA replication and Histone Gene transcription. Here we review the molecular Histone-related requirements for G1/S-phase progression during the cell cycle. Recently developed experimental strategies, now enable us to explore mechanisms involved in dynamic control of Histone Gene expression in the context of the temporal (cell cycle) and spatial (HLBs) remodeling of the Histone Gene loci.
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cyclin d2 and the cdk substrate p220npat are required for self renewal of human embryonic stem cells
Journal of Cellular Physiology, 2010Co-Authors: Klaus A Becker, Janet L Stein, Jane B Lian, Andre J. Van Wijnen, Prachi N Ghule, Gary S SteinAbstract:Self-renewal of pluripotent human embryonic stem (hES) cells utilizes an abbreviated cell cycle that bypasses E2F/pRB-dependent growth control. We investigated whether self-renewal is alternatively regulated by cyclin/CDK phosphorylation of the p220(NPAT)/HiNF-P complex to activate Histone Gene expression at the G1/S phase transition. We show that cyclin D2 is prominently expressed in pluripotent hES cells, but cyclin D1 eclipses cyclin D2 during differentiation. Depletion of cyclin D2 or p220(NPAT) causes a cell cycle defect in G1 reflected by diminished phosphorylation of p220(NPAT), decreased cell cycle dependent Histone H4 expression and reduced S phase progression. Thus, cyclin D2 and p220(NPAT) are principal cell cycle regulators that determine competency for self-renewal in pluripotent hES cells. While pRB/E2F checkpoint control is relinquished in human ES cells, fidelity of physiological regulation is secured by cyclin D2 dependent activation of the p220(NPAT)/HiNF-P mechanism that may explain perpetual proliferation of hES cells without transformation or tumoriGenesis.
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the Histone Gene cell cycle regulator hinf p is a unique zinc finger transcription factor with a novel conserved auxiliary dna binding motif
Biochemistry, 2008Co-Authors: Ricardo F Medina, Janet L Stein, Jane B Lian, Andre J. Van Wijnen, Sayyed K Zaidi, Timothy Buck, Angela Mielechamberland, Gary S SteinAbstract:Accumulation of Histone proteins is necessary for packaging of replicated DNA during the S phase of the cell cycle. Different mechanisms operate to regulate Histone protein levels, and induction of human Histone Gene expression at the G1-S phase transition plays a critical role. The zinc finger HiNF-P and coactivator p220 (NPAT) proteins are key regulators of Histone Gene expression. Here, we describe a novel HiNF-P-specific conserved region (PSCR) located within the C-terminus that is present in HiNF-P homologues of all metazoan species that have been examined. The PSCR motif is required for activation of Histone H4 Gene transcription and contributes to DNA binding of HiNF-P. Thus, the PSCR module represents an auxiliary DNA-binding determinant that plays a critical role in mediating Histone Gene expression during the cell cycle and defines HiNF-P as a unique cell cycle regulatory member of the zinc finger transcription factor family.
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cell cycle dependent phosphorylation and subnuclear organization of the Histone Gene regulator p220npat in human embryonic stem cells
Journal of Cellular Physiology, 2007Co-Authors: Prachi N Ghule, Janet L Stein, Klaus A Becker, Jane B Lian, Andre J Van Wijnen, Wade J Harper, Gary S SteinAbstract:Human embryonic stem (ES) cells have an expedited cell cycle (∼15 h) due to an abbreviated G1 phase (∼2.5 h) relative to somatic cells. One principal regulatory event during cell cycle progression is the G1/S phase induction of Histone biosynthesis to package newly replicated DNA. In somatic cells, Histone H4 Gene expression is controlled by CDK2 phosphorylation of p220NPAT and localization of HiNF-P/p220NPAT complexes with Histone Genes at Cajal body related subnuclear foci. Here we show that this ‘S point’ pathway is operative in situ in human ES cells (H9 cells; NIH-designated WA09). Immunofluorescence microscopy shows an increase in p220NPAT foci in G1 reflecting the assembly of Histone Gene regulatory complexes in situ. In contrast to somatic cells where duplication of p220NPAT foci is evident in S phase, the increase in the number of p220NPAT foci in ES cells appears to precede the onset of DNA synthesis as measured by BrdU incorporation. Phosphorylation of p220NPAT at CDK dependent epitopes is most pronounced in S phase when cells exhibit elevated levels of cyclins E and A. Our data indicate that subnuclear organization of the HiNF-P/p220NPAT pathway is rapidly established as ES cells emerge from mitosis and that p220NPAT is subsequently phosphorylated in situ. Our findings establish that the HiNF-P/p220NPAT Gene regulatory pathway operates in a cell cycle dependent microenvironment that supports expression of DNA replication-linked Histone Genes and chromatin assembly to accommodate human stem cell self-renewal. J. Cell. Physiol. 213: 9–17, 2007. © 2007 Wiley-Liss, Inc.
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cell cycle dependent phosphorylation and subnuclear organization of the Histone Gene regulator p220 npat in human embryonic stem cells
Journal of Cellular Physiology, 2007Co-Authors: Prachi N Ghule, Janet L Stein, Klaus A Becker, Jane B Lian, Andre J. Van Wijnen, Wade J Harper, Gary S SteinAbstract:Human embryonic stem (ES) cells have an expedited cell cycle ( approximately 15 h) due to an abbreviated G1 phase ( approximately 2.5 h) relative to somatic cells. One principal regulatory event during cell cycle progression is the G1/S phase induction of Histone biosynthesis to package newly replicated DNA. In somatic cells, Histone H4 Gene expression is controlled by CDK2 phosphorylation of p220(NPAT) and localization of HiNF-P/p220(NPAT) complexes with Histone Genes at Cajal body related subnuclear foci. Here we show that this 'S point' pathway is operative in situ in human ES cells (H9 cells; NIH-designated WA09). Immunofluorescence microscopy shows an increase in p220(NPAT) foci in G1 reflecting the assembly of Histone Gene regulatory complexes in situ. In contrast to somatic cells where duplication of p220(NPAT) foci is evident in S phase, the increase in the number of p220(NPAT) foci in ES cells appears to precede the onset of DNA synthesis as measured by BrdU incorporation. Phosphorylation of p220(NPAT) at CDK dependent epitopes is most pronounced in S phase when cells exhibit elevated levels of cyclins E and A. Our data indicate that subnuclear organization of the HiNF-P/p220(NPAT) pathway is rapidly established as ES cells emerge from mitosis and that p220(NPAT) is subsequently phosphorylated in situ. Our findings establish that the HiNF-P/p220(NPAT) Gene regulatory pathway operates in a cell cycle dependent microenvironment that supports expression of DNA replication-linked Histone Genes and chromatin assembly to accommodate human stem cell self-renewal.