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Biomedical subjects

Hiroyuki Niida

Publications and source records attributed to Hiroyuki Niida.

8 recordsLinked to original sources
PubMed2006

Genetic instability in cancer cells by impaired cell cycle checkpoints.

Cells continuously encounter DNA damage caused either by damaging agents, including oxygen radicals and DNA replication errors caused by stalled replication forks, or by extracellular environments such as ultraviolet or ionizing irradiation. Such DNA damage poses a great threat to genome stability, potentially leading to loss or amplification of chromosome activity, which may result in cellular senescence, cancer or apoptosis. The DNA damage checkpoints coordinate an arrest in cell cycle progression with the DNA repair process, suppressing either mitotic catastrophe or proliferation of cells with damaged DNA. Numerous key players have been identified in terms of damage sensor proteins, transducer kinases and effectors, but their coordination and interconnectedness in damage control have only recently become evident. In this review, we discuss changes in chromatin structure, recruitment of mediator proteins and activation of transducer kinases in response to DNA damage. These cellular responses are important for determining the potential effects of current cancer therapies in terms of toxicity and efficacy.

Cell Cycle↗
PubMed2005

DNA damage checkpoints in mammals.

DNA damage is a common event and probably leads to mutation or deletion within chromosomal DNA, which may cause cancer or premature aging. DNA damage induces several cellular responses including DNA repair, checkpoint activity and the triggering of apoptotic pathways. DNA damage checkpoints are associated with biochemical pathways that end delay or arrest of cell-cycle progression. These checkpoints engage damage sensor proteins, such as the Rad9-Rad1-Hus1 (9-1-1) complex, and the Rad17-RFC complex, in the detection of DNA damage and transduction of signals to ATM, ATR, Chk1 and Chk2 kinases. Chk1 and Chk2 kinases regulate Cdc25, Wee1 and p53 that ultimately inactivate cyclin-dependent kinases (Cdks) which inhibit cell-cycle progression. In this review, we discuss the molecular mechanisms by which DNA damage is recognized by sensor proteins and signals are transmitted to Cdks. We classify the genes involved in checkpoint signaling into four categories, namely sensors, mediators, transducers and effectors, although their proteins have the broad activity, and thus this classification is for convenience and is not definitive.

Animals↗
PubMed2005

Depletion of Chk1 leads to premature activation of Cdc2-cyclin B and mitotic catastrophe.

Mitotic catastrophe occurs as a result of the uncoupling of the onset of mitosis from the completion of DNA replication, but precisely how the ensuing lethality is regulated or what signals are involved is largely unknown. We demonstrate here the essential role of the ATM/ATR-p53 pathway in mitotic catastrophe from premature mitosis. Chk1 deficiency resulted in a premature onset of mitosis because of abnormal activation of cyclin B-Cdc2 and led to the activation of caspases 3 and 9 triggered by cytoplasmic release of cytochrome c. This deficiency was associated with foci formation by the phosphorylated histone, H2AX (gammaH2AX), specifically at S phase. Ectopic expression of Cdc2AF, a mutant that cannot be phosphorylated at inhibitory sites, also induced premature mitosis and foci formation by gammaH2AX at S phase in both embryonic stem cells and HCT116 cells. Depletion of ATM and ATR protected against cell death from premature mitosis. p53-deficient cells were highly resistant to lethality from premature mitosis as well. Our results therefore suggest that ATM/ATR-p53 is required for mitotic catastrophe that eliminates cells escaping Chk1-dependent mitotic regulation. Loss of this function might be important in mammalian tumorigenesis.

Animals↗
PubMed2004

Human SAD1 kinase is involved in UV-induced DNA damage checkpoint function.

Checkpoint activation by DNA damage during G(2) prevents activation of cyclin B/Cdc2 complexes, and as a consequence, mitotic entry is blocked. Although initiation and maintenance of G(2) arrest are known to be regulated by at least two distinct signaling pathways, including those of p38MAPK and ataxia-telangiectasia-mutated (ATM)- and Rad3-related (ATR)-Chk1 in higher eukaryotes, the actual number of signaling pathways involved in this regulation is still elusive. In the present study, we identified human SAD1 (hsSAD1) by searching a sequence data base. The predicted hsSAD1 protein comprises 778 amino acids and shares significant homology with the fission yeast Cdr2, a mitosis-regulatory kinase, and Caenorhabditis elegans SAD1, a neuronal cell polarity regulator. HsSAD1 transcript was expressed ubiquitously with the highest levels of expression in brain and testis. HsSAD1 specifically phosphorylated Wee1A, Cdc25-C, and -B on Ser-642, Ser-216, and Ser-361 in vitro, respectively. Overexpression of hsSAD1 resulted in an increased phosphorylation of Cdc25C on Ser-216 in vivo. DNA damage induced by UV or methyl methane sulfonate but not by IR enhanced endogenous hsSAD1 kinase activity in a caffeine-sensitive manner and caused translocation of its protein from cytoplasm to nucleus. Overexpression of wild-type hsSAD1 induced G(2)/M arrest in HeLa S2 cells. Furthermore, UV-induced G(2)/M arrest was partially abrogated by the reduced expression of hsSAD1 using small interfering RNA. These results suggest that hsSAD1 acts as checkpoint kinase upon DNA damage induced by UV or methyl methane sulfonate. The identification of this new kinase suggests the existence of an alternative checkpoint pathway other than those of ATR-Chk1 and p38MAPK.

Amino Acid Sequence↗
PubMed2004

Negative regulation of Chk2 expression by p53 is dependent on the CCAAT-binding transcription factor NF-Y.

The kinase Chk2 and tumor suppressor p53 participate in an ill defined regulatory interaction in mammalian cells. The abundance of Chk2 mRNA and protein has now been shown to be decreased by the induction of p53 in Saos2 cells. Ionizing radiation also triggered the phosphorylation and subsequent down-regulation of Chk2 in human colorectal HCT116 (p53(+/+)) cancer cells; irradiation of its isogenic mutant HCT116 (p53(-/-)) cells, which lack functional p53, induced Chk2 phosphorylation but not its down-regulation. In addition, HCT116 (p53(+/+)) cells constitutively expressing a dominant negative p53 (V143A) failed to suppress Chk2 expression after irradiation. Reporter gene assays in HCT116 (p53(+/+)) cells revealed that wild-type p53 repressed, whereas a dominant negative p53 mutant increased, the activity of the human Chk2 gene promoter. Mutational analysis showed that a CCAAT box located between nucleotides -152 and -138 of the promoter was responsible for its negative regulation by p53. Electrophoretic mobility shift assays demonstrated that the transcription factor NF-Y binds to this CCAAT sequence. A dominant negative mutant of NF-YA abolished the effect of p53 on Chk2 promoter activity. These results suggest that p53 negatively regulates Chk2 gene transcription through modulation of NF-Y function and that this regulation may be important for reentry of cells into the cell cycle after DNA damage is repaired.

Base Sequence↗
PubMed2003

Genomic structure and characterization of the promoter region of the human NAK gene.

NAK has been identified as an IkappaB-kinase activating-kinase that plays an important role in NF-kappaB activation in response to several pro-inflammatory cytokines such as TNF-alpha. We describe here the genomic structure of the human NAK gene and analysis of the promoter. The gene spanned 40.5 kb and contained 21 exons with lengths ranging from 39 to 196 bp. Comparison of the phase and position of intron insertions within the human NAK gene with those within IKKalpha, IKKbeta and IKK epsilon indicated that the exon/intron organization of IKK epsilon is more highly conserved than that of IKKalpha or IKKbeta. The transcriptional start site was mapped at a position about 98 bp upstream from the translation start site by means of both an RNase protection assay and a primer extension method. Fluorescence in situ hybridization using full-length human NAK cDNA as a probe showed that the human NAK gene is localized to human chromosome 13q14.2-3, a region in which the loss of heterozygosity is associated with squamous cell carcinoma and leukemia. By using a series of deletion constructs in performing a reporter assay, a minimal 77 bp upstream of the transcriptional initiation site was shown to contribute to the major promoter activity.

5' Flanking Region↗
PubMed2002

G9a histone methyltransferase plays a dominant role in euchromatic histone H3 lysine 9 methylation and is essential for early embryogenesis.

Covalent modification of histone tails is crucial for transcriptional regulation, mitotic chromosomal condensation, and heterochromatin formation. Histone H3 lysine 9 (H3-K9) methylation catalyzed by the Suv39h family proteins is essential for establishing the architecture of pericentric heterochromatin. We recently identified a mammalian histone methyltransferase (HMTase), G9a, which has strong HMTase activity towards H3-K9 in vitro. To investigate the in vivo functions of G9a, we generated G9a-deficient mice and embryonic stem (ES) cells. We found that H3-K9 methylation was drastically decreased in G9a-deficient embryos, which displayed severe growth retardation and early lethality. G9a-deficient ES cells also exhibited reduced H3-K9 methylation compared to wild-type cells, indicating that G9a is a dominant H3-K9 HMTase in vivo. Importantly, the loss of G9a abolished methylated H3-K9 mostly in euchromatic regions. Finally, G9a exerted a transcriptionally suppressive function that depended on its HMTase activity. Our results indicate that euchromatic H3-K9 methylation regulated by G9a is essential for early embryogenesis and is involved in the transcriptional repression of developmental genes.

Acetylation↗