Histone H3 phosphorylation is required for the initiation, but not maintenance, of mammalian chromosome condensation

1998 ◽  
Vol 111 (23) ◽  
pp. 3497-3506 ◽  
Author(s):  
A. Van Hooser ◽  
D.W. Goodrich ◽  
C.D. Allis ◽  
B.R. Brinkley ◽  
M.A. Mancini
Keyword(s):  
Cell Cycle Progression ◽  
Histone H3 ◽  
Chromosome Morphology ◽  
Chromosome Condensation ◽  
S Phase ◽  
Condensed State ◽  
Cycle Progression ◽  
H3 Phosphorylation ◽  
Excess Substrate ◽  
Histone H3 Phosphorylation

The temporal and spatial patterns of histone H3 phosphorylation implicate a specific role for this modification in mammalian chromosome condensation. Cells arrest in late G2 when H3 phosphorylation is competitively inhibited by microinjecting excess substrate at mid-S-phase, suggesting a requirement for activity of the kinase that phosphorylates H3 during the initiation of chromosome condensation and entry into mitosis. Basal levels of phosphorylated H3 increase primarily in late-replicating/early-condensing heterochromatin both during G2 and when premature chromosome condensation is induced. The prematurely condensed state induced by okadaic acid treatment during S-phase culminates with H3 phosphorylation throughout the chromatin, but in an absence of mitotic chromosome morphology, indicating that the phosphorylation of H3 is not sufficient for complete condensation. Mild hypotonic treatment of cells arrested in mitosis results in the dephosphorylation of H3 without a cytological loss of chromosome compaction. Hypotonic-treated cells, however, complete mitosis only when H3 is phosphorylated. These observations suggest that H3 phosphorylation is required for cell cycle progression and specifically for the changes in chromatin structure incurred during chromosome condensation.

scholarly journals Cyclic AMP‐induced loss of histone H3 phosphorylation and disruption of cell cycle progression mediated through a novel cAMP binding protein

2006 ◽  
Vol 20 (5) ◽  
Author(s):  
Rebecca Claire Chiffer ◽  
Sara K. Snyder ◽  
Pedro Rodriguez ◽  
Eric Anderson ◽  
Catharine L. Smith
Keyword(s):  
Cell Cycle ◽  
Cyclic Amp ◽  
Cell Cycle Progression ◽  
Binding Protein ◽  
Histone H3 ◽  
Cycle Progression ◽  
H3 Phosphorylation ◽  
Histone H3 Phosphorylation

scholarly journals FOXD1 regulates cell division in clear cell renal cell carcinoma

BMC Cancer ◽  
2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Kyle H. Bond ◽  
Jennifer L. Fetting ◽  
Christine W. Lary ◽  
Ivette F. Emery ◽  
Leif Oxburgh
Keyword(s):  
Cell Cycle ◽  
Cell Cycle Progression ◽  
Patient Survival ◽  
Histone H3 ◽  
Tumor Formation ◽  
Cycle Progression ◽  
H3 Phosphorylation ◽  
Histone H3 Phosphorylation

Abstract Background Forkhead transcription factors control cell growth in multiple cancer types. Foxd1 is essential for kidney development and mitochondrial metabolism, but its significance in renal cell carcinoma (ccRCC) has not been reported. Methods Transcriptome data from the TCGA database was used to correlate FOXD1 expression with patient survival. FOXD1 was knocked out in the 786-O cell line and known targets were analyzed. Reduced cell growth was observed and investigated in vitro using growth rate and Seahorse XF metabolic assays and in vivo using a xenograft model. Cell cycle characteristics were determined by flow cytometry and immunoblotting. Immunostaining for TUNEL and γH2AX was used to measure DNA damage. Association of the FOXD1 pathway with cell cycle progression was investigated through correlation analysis using the TCGA database. Results FOXD1 expression level in ccRCC correlated inversely with patient survival. Knockout of FOXD1 in 786-O cells altered expression of FOXD1 targets, particularly genes involved in metabolism (MICU1) and cell cycle progression. Investigation of metabolic state revealed significant alterations in mitochondrial metabolism and glycolysis, but no net change in energy production. In vitro growth rate assays showed a significant reduction in growth of 786-OFOXD1null. In vivo, xenografted 786-OFOXD1null showed reduced capacity for tumor formation and reduced tumor size. Cell cycle analysis showed that 786-OFOXD1null had an extended G2/M phase. Investigation of mitosis revealed a deficiency in phosphorylation of histone H3 in 786-OFOXD1null, and increased DNA damage. Genes correlate with FOXD1 in the TCGA dataset associate with several aspects of mitosis, including histone H3 phosphorylation. Conclusions We show that FOXD1 regulates the cell cycle in ccRCC cells by control of histone H3 phosphorylation, and that FOXD1 expression governs tumor formation and tumor growth. Transcriptome analysis supports this role for FOXD1 in ccRCC patient tumors and provides an explanation for the inverse correlation between tumor expression of FOXD1 and patient survival. Our findings reveal an important role for FOXD1 in maintaining chromatin stability and promoting cell cycle progression and provide a new tool with which to study the biology of FOXD1 in ccRCC.

scholarly journals Chromatin remodeling in somatic cells injected into mature pig oocytes

Reproduction ◽  
2006 ◽  
Vol 131 (6) ◽  
pp. 1037-1049 ◽  
Author(s):  
Hong-Thuy Bui ◽  
Nguyen Van Thuan ◽  
Teruhiko Wakayama ◽  
Takashi Miyano
Keyword(s):  
Somatic Cell ◽  
Somatic Cells ◽  
Histone H3 ◽  
Chromosome Morphology ◽  
Chromosome Condensation ◽  
Cell Nuclei ◽  
H3 Phosphorylation ◽  
Histone H3 Methylation ◽  
Histone H3 Phosphorylation

We examined the involvement of histone H3 modifications in the chromosome condensation and decondensation of somatic cell nuclei injected into mature pig oocytes. Nuclei of pig granulosa cells were transferred into in vitro matured intact pig oocytes, and histone H3 phosphorylation, acetylation, and methylation were examined by immunostaining with specific antibodies in relation to changes in chromosome morphology. In the condensed chromosomes of pig oocytes at metaphase II, histone H3 was phosphorylated at serine 10 (H3–S10) and serine 28 (H3–S28), and methylated at lysine 9 (H3–K9), but was not acetylated at lysine 9, 14 and 18 (H3–K9, H3–K14 and H3–K18). During the first 2 h after nuclear transfer, a series of events were observed in the somatic nuclei: nuclear membrane disassembly; chromosome condensation to form a metaphase-like configuration; an increase in histone H3 phosphorylation levels (H3–S10 and H3–S28). Next, pig oocytes injected with nuclei of somatic cells were electroactivated and the chromosome morphology of oocytes and somatic cells was examined along with histone modifications. Generally, chromosomes of the somatic cells showed a similar progression of cell cycle stage to that of oocytes, through anaphase II- and telophase II-like stages then formed pronucleus-like structures, although the morphology of the spindles differed from that of oocyte spindles. The chromosomes of somatic cells also showed changes in histone H3 dephosphorylation and reacetylation, similar to oocytes. In contrast, histone H3 methylation (H3–K9) of somatic cell nuclei did not show any significant change after injection and electroactivation of the oocytes. These results suggest that nuclear remodeling including histone H3 phosphorylation and acetylation of injected somatic nuclei took place in the oocytes under regulation by the oocyte cytoplasm.

scholarly journals Cell Cycle-dependent Expression and Nucleolar Localization of hCAP-H

2001 ◽  
Vol 12 (11) ◽  
pp. 3527-3537 ◽  
Author(s):  
Olga A. Cabello ◽  
Elena Eliseeva ◽  
WeiGong He ◽  
Hagop Youssoufian ◽  
Sharon E. Plon ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Histone H3 ◽  
Chromosome Condensation ◽  
Nucleolar Localization ◽  
Mitotic Chromosomes ◽  
Proliferating Cells ◽  
Protein Levels ◽  
Sister Chromatids ◽  
H3 Phosphorylation ◽  
Histone H3 Phosphorylation

Condensin is a conserved 13S heteropentamer composed of two nonidentical structural maintenance of chromosome (SMC) family proteins, in Xenopus XCAP-C and XCAP-E, and three regulatory subunits, XCAP-D2, XCAP-G, and XCAP-H. Both biochemical and genetic analyses have demonstrated an essential role for the 13S condensin complex in mitotic chromosome condensation. Further, a potential requirement for condensin in completion of chromatid arm separation in early anaphase is demonstrated by the mutational phenotypes of the Drosophila homologues ofXCAP-H, barren and XCAP-C,DmSMC4. In this study we have investigated the expression and subcellular distribution of hCAP-H, the human homolog of XCAP-H, in order to better understand its cellular functions. Transcription of hCAP-H was restricted to proliferating cells with highest expression during the G2 phase of the cell cycle. In contrast, cellular hCAP-H protein levels were constant throughout the cell cycle. hCAP-H was found to be associated with mitotic chromosomes exhibiting a nonuniform but symmetric distribution along sister chromatids. The symmetry of hCAP-H association with sister chromatids suggests that there are sequence-dependent domains of condensin aggregation. During interphase hCAP-H, -C, and -E, have distinct punctate nucleolar localization, suggesting that condensin may associate with and modulate the conformation and function of rDNA. hCAP-H association with condensed chromatin was not observed in the early phase of chromosome condensation when histone H3 phosphorylation has already taken place. This finding is consistent with the hypothesis that histone H3 phosphorylation precedes condensin-mediated condensation.

scholarly journals 62SPINDLE MORPHOGENESIS AND THE MORPHOLOGY OF CHROMOSOMES IN MOUSE NUCLEAR TRANSFER: AN ABNORMAL START IN CLONING OF MICE

2004 ◽  
Vol 16 (2) ◽  
pp. 153
Author(s):  
V.T. Nguyen ◽  
S. Wakayama ◽  
S. Kishigami ◽  
T. Wakayama
Keyword(s):  
Success Rate ◽  
Nuclear Membrane ◽  
Nuclear Transfer ◽  
Histone H3 ◽  
Donor Cell ◽  
Chromosome Morphology ◽  
H3 Phosphorylation ◽  
Monopolar Spindle ◽  
Histone H3 Phosphorylation ◽  
Bipolar Spindles

Cloning of mammals by somatic nuclear transfer (NT) was achieved more than 6 years ago. Nonetheless, the success rate is very low. Although spindles and nuclear mitotic apparatus protein (NuMA) are thought to play important roles in the cell cycle and maybe reprogramming after NT, the spindle and NuMA morphogenesis after NT is not well understood. The aim of this study was to examine the spindle, NuMA morphogenesis and chromosome morphology in mouse NT. Maturated mouse oocytes were collected from mature B6D2F1 females 16h after hCG injection. The donor nuclei used in this study were cumulus cells. A donor nucleus was injected into an oocyte by means of a piezo pulse system. For double labeling, DNA was stained with PI (red) or Hoechst 33342 (blue); tubulin, NuMA, and phosphorylated histone H3 were stained with FITC or alexa 488 (green). All samples were visualized with a BioRad Radiance 2100 confocal scanning laser microscope. In this study, the spindle, NuMA morphogenesis, chromosome morphology, and histone H3 phosphorylation were identified using more than 3000 maturated mouse oocytes in five experimental studies to examine (1) the morphological changes of spindle and NuMA in NT oocytes at 10, 30, 60, 90, 120, 180, and 360min after injection of somatic cell nuclei into intact or enucleated maturated oocytes; (2) the localization and morphology of spindles and NuMA of NT embryo followed by activation at 1, 2, and 6h after somatic nuclear injection; (3) the effects of the timing of activation and donor cell membrane on the spindle morphogenesis; (4) the effects of intact nuclear membrane or its breakdown by micropipette on spindle morphogenesis; and (5) the correlation between donor cell chromosome condensation and histone H3 phosphorylation after injection into enucleated maturated oocytes. As control, the NuMA morphogenesis during the early pronuclear stages of NT was compared with those of ICSI oocytes. We consistently observed that abnormal spindle morphogenesis occurred during the early stages of mouse NT. Most of the NT oocytes began with monopolar spindle and monopolar NuMA (90–95%), and this phenomenon occurred 10min after NT. Only 5–10% of NT oocytes started with bipolar spindles. However, monopolar spindles were transformed into bipolar spindles during 30–60min after NT by bipolarization of centrosomal NuMA. After activation, NuMA was localized in the perispindle region, and finally concentrated inside the pronuclei of embryo. This spindle morphogenesis was independent of the presence or absence of metaphase II [not like in rhesus monkeys, whereas meiotic spindle removal depletes the ooplasm of NuMA and HSET, both vital for mitotic spindle pole formation (Lanza R et al., 2003 Science, 300, 297)]. The breakdown of the donor nuclear membrane by micropipette resulted in the increase of bipolar spindle formation in NT. The initiation of chromosome condensation and histone H3 phosphorylation were observed 30 min after NT and the maximum of histone H3 phosphorylation was occurred 60 to 90min after NT. Future studies are required to elucidate the mechanism of monopolar spindle formation, chromosomal abnormalities in NT, and the relation of the monopolar phenomenon to the success rate in mammalian cloning.

scholarly journals Drosophila Aurora B Kinase Is Required for Histone H3 Phosphorylation and Condensin Recruitment during Chromosome Condensation and to Organize the Central Spindle during Cytokinesis

2001 ◽  
Vol 152 (4) ◽  
pp. 669-682 ◽  
Author(s):  
Régis Giet ◽  
David M. Glover
Keyword(s):  
Histone H3 ◽  
Chromosome Condensation ◽  
Aurora B ◽  
Aurora B Kinase ◽  
Kinase Gene ◽  
Central Spindle ◽  
H3 Phosphorylation ◽  
Founding Family ◽  
Reduced Density ◽  
Histone H3 Phosphorylation

Aurora/Ipl1-related kinases are a conserved family of enzymes that have multiple functions during mitotic progression. Although it has been possible to use conventional genetic analysis to dissect the function of aurora, the founding family member in Drosophila (Glover, D.M., M.H. Leibowitz, D.A. McLean, and H. Parry. 1995. Cell. 81:95–105), the lack of mutations in a second aurora-like kinase gene, aurora B, precluded this approach. We now show that depleting Aurora B kinase using double-stranded RNA interference in cultured Drosophila cells results in polyploidy. aurora B encodes a passenger protein that associates first with condensing chromatin, concentrates at centromeres, and then relocates onto the central spindle at anaphase. Cells depleted of the Aurora B kinase show only partial chromosome condensation at mitosis. This is associated with a reduction in levels of the serine 10 phosphorylated form of histone H3 and a failure to recruit the Barren condensin protein onto chromosomes. These defects are associated with abnormal segregation resulting from lagging chromatids and extensive chromatin bridging at anaphase, similar to the phenotype of barren mutants (Bhat, M.A., A.V. Philp, D.M. Glover, and H.J. Bellen. 1996. Cell. 87:1103–1114.). The majority of treated cells also fail to undertake cytokinesis and show a reduced density of microtubules in the central region of the spindle. This is accompanied by a failure to correctly localize the Pavarotti kinesin-like protein, essential for this process. We discuss these conserved functions of Aurora B kinase in chromosome transmission and cytokinesis.

scholarly journals 309CHROMOSOME CONDENSATION IS CORRELATED WITH HISTONE H3 PHOSPHORYLATION WITHOUT CDC2 KINASE AND MAP KINASE ACTIVITIES IN PIG OOCYTES

2004 ◽  
Vol 16 (2) ◽  
pp. 273
Author(s):  
T. Bui Hong ◽  
L.G. Villa-Diaz ◽  
E. Yamaoka ◽  
T. Miyano
Keyword(s):  
Map Kinase ◽  
Kinase Activity ◽  
Histone H3 ◽  
Germinal Vesicle ◽  
Chromosome Condensation ◽  
Kinase Assay ◽  
Cdc2 Kinase ◽  
H3 Phosphorylation ◽  
Diakinesis Stage ◽  
Histone H3 Phosphorylation

Chromosome condensation is the first step of oocyte maturation. When the oocytes resume meiosis, chromosomes start to condense and Cdc2 kinase becomes activated. However, recent findings show that the chromosome condensation does not always correlate with Cdc2 kinase activity in pig oocytes. The objectives of this study were to examine (1) the correlation between chromosome condensation and histone H3 phosphorylation at serine 10 (Ser10) during meiotic maturation of pig oocytes, and (2) the effects of protein phosphatase 1/2A (PP1/PP2A) inhibitors on the chromosome condensation and the involvement of Cdc2 kinase, MAP kinase and histone H3 kinase in this process. Oocyte-cumulus-granulosa cell complexes (OCGCs) were collected from follicles of 4–6mm in diameter. OCGCs were cultured in modified TCM 199 for different periods of time to obtain oocytes at the germinal vesicle (GV, 0h), diakinesis (18h), metaphase I (24–27h), anaphase I to telophase I (30–33h), and metaphase II (42h) stages. To examine the effects of PP1/PP2A inhibitors on the chromosome condensation, oocyte-cumulus-complexes (OCCs) were cultured in modified TCM 199 with either 2.5μM okadaic acid (OA) or 50nM calyculin A (CL-A) for 0.5, 1, 2, 3, 4 and 6h. To inhibit the MAP kinase activity in the oocytes treated with the PP1/PP2A inhibitor, OCCs were cultured in medium containing CL-A and the MEK inhibitor, U0126 (0.1mM). Morphology of the chromosome and nuclear membrane, and phosphorylation of histone H3 were examined by the immunofluorescent microscopy. In each group 30 oocytes were examined for OA or CL-A and 60 oocytes for CL-A+U0126 treatments. Activities of Cdc2 kinase, MAP kinase and histone H3 kinase were also examined. Phosphorylation of histone H3 (Ser10) was not detected in the oocytes at the GV stage. The phosphorylation was first detected in the clump of condensed chromosomes at the diakinesis stage of prophase I and maintained until metaphase II. The kinase assay also showed that histone H3 kinase activity was low in GV oocytes, increased at the diakinesis stage, and then maintained high activity until metaphase II. PP1/PP2A inhibitors induced rapid chromosome condensation in pig oocytes. Histone H3 phosphorylation (Ser10) became detectable together with the chromosome condensation in the treated oocytes after 2h. After 6h, oocytes had highly condensed chromosomes with phosphorylated histone H3 (81% in CL-A- and 71% in OA-treated oocytes). Both histone H3 kinase and MAP kinase were activated in the treated oocytes, although Cdc2 kinase was not activated. In the oocytes treated with CL-A and U0126, neither Cdc2 kinase nor MAP kinase were activated, although histone H3 kinase was still activated and chromosomes condensed. These results suggest that phosphorylation of histone H3 (Ser10) occurs in condensed chromosomes during maturation in pig oocytes. Futhermore, the chromosome condensation is correlated with histone H3 kinase activity, but not with Cdc2 kinase and MAP kinase activities.

Sign in / Sign up

Export Citation Format

Share Document