Yasufumi Murakami - Academia.edu (original) (raw)
Papers by Yasufumi Murakami
<b>Copyright information:</b>Taken from "Loss of affects gene expression profile... more <b>Copyright information:</b>Taken from "Loss of affects gene expression profile in a genome-wide manner in ES cells and liver cells"http://www.biomedcentral.com/1471-2164/8/227BMC Genomics 2007;8():227-227.Published online 10 Jul 2007PMCID:PMC1959195.nes (A) & (B). Horizontal and vertical axes represent expression levels normalized for an individual gene. Each point represents normalized expression data for an individual gene. The genes that showed standard deviation greater than 2.0 in the normalized data of both genotypes (A) were excluded and gene lists were constructed with < 0.05 (B). Fig. 1D–F in the original article [1] remains unchanged and is presented as (C) – (E), respectively.
Transcriptional activation and repression are a key step in the regulation of all cellular activi... more Transcriptional activation and repression are a key step in the regulation of all cellular activities. The development of comprehensive analysis methods such as DNA microarray has advanced our understanding of the correlation between the regulation of transcription and that of cellular mechanisms. However, DNA microarray analysis based on steady-state mRNA (total mRNA) does not always correspond to transcriptional activation or repression. To comprehend these transcriptional regulations, the detection of nascent RNAs is more informative. Although the nuclear run-on assay can detect nascent RNAs, it has not been fully applied to DNA microarray analysis. In this study, we have developed a highly efficient method for isolating bromouridine-labeled nascent RNAs that can be successfully applied to DNA microarray analysis. This method can linearly amplify small amounts of mRNAs with little bias. Furthermore, we have applied this method to DNA microarray analysis from mouse G2-arrested cel...
The Molecular Biology Society of Japan, 2016
Genomic DNA contains not only information of DNA sequence, but also epigenetic information that i... more Genomic DNA contains not only information of DNA sequence, but also epigenetic information that is the direct DNA modification by methylation (the addition of methyl group to the 5th carbon of pyrimidine ring of cytosine) and histone modifications (acetylation, methylation, etc). Epigenetic information is closely related to regulation of gene expression. If a methyl group is dislocated to position 5 of the pyrimidine ring of cytosine, the hydrogen bond between complementary GC bases will not be inhibited, but this methyl group is positioned so as to be exposed in the major groove of the double-helix structure of DNA, and according to the genome region/sequence undergoing modification of methylation, gene expression is inhibited by the interaction between the genome and DNAbinding molecules.
Genome Informatics, 1994
The DNA sequence of chromosome VI (270kb) of the budding yeast S. cerevisiae, has been determined... more The DNA sequence of chromosome VI (270kb) of the budding yeast S. cerevisiae, has been determined. The sequence data were then analyzed with the software GENETYX (Software Development Co.) to nd out the candidates of the coding region. As there is little number of intron in budding yeast genome, it is relatively easy to nd the candidates of the coding region. Open reading frames identi ed by the software were generally the same as the coding sequence with little exception. To carry out further analysis of the open reading frames, we tried to automate the process required for similarity search. The processes automated are
The Molecular Biology Society of Japan, 2016
SUMMARYHepatocytes can be used to study the pathogenesis of liver diseases and drug discovery res... more SUMMARYHepatocytes can be used to study the pathogenesis of liver diseases and drug discovery research. Human hepatocytes are, however, hardly expandable in vitro making it difficult to secure large numbers of cells from one donor. In this study, we aimed to establish an in vitro long-term culture system that enables stable proliferation and maintenance of human hepatocytes to ensure a constant supply. We purified human hepatocytes by selection with cytocidal puromycin, and cultured them for more than 60 population doublings over a span of more than 350 days. These results show that this simple culture system with usage of the cytocidal antibiotics enables efficient hepatocyte proliferation and is an effective method for generating a stable supply of hepatocytes for drug discovery research at a significant cost reduction.
Journal of Biological Chemistry
International Journal of Molecular Sciences
Cell division is essential for the maintenance of life and involves chromosome segregation and su... more Cell division is essential for the maintenance of life and involves chromosome segregation and subsequent cytokinesis. The processes are tightly regulated at both the spatial and temporal level by various genes, and failures in this regulation are associated with oncogenesis. Here, we investigated the gene responsible for defects in cell division by using murine temperature-sensitive (ts) mutant strains, tsFT101 and tsFT50 cells. The ts mutants normally grow in a low temperature environment (32 °C) but fail to divide in a high temperature environment (39 °C). Exome sequencing and over-expression analyses identified Diaph3, a member of the formin family, as the cause of the temperature sensitivity observed in tsFT101 and tsFT50 cells. Interestingly, Diaph3 knockout cells showed abnormality in cytokinesis at 39 °C, and the phenotype was rescued by re-expression of Diaph3 WT, but not Diaph1 and Diaph2, other members of the formin family. Furthermore, Diaph3 knockout cells cultured at 3...
Japan Agricultural Research Quarterly: JARQ
To compare the seed productivity of germplasm in India (Tamil Nadu) and Japan (Tsukuba), a total ... more To compare the seed productivity of germplasm in India (Tamil Nadu) and Japan (Tsukuba), a total of 105 accessions from the Japanese National Agriculture and Food Research Organization (NARO) sorghum core collection were cultivated. The comparative cultivation studies were conducted at two locations in India and one in Japan. Differences in cultivation environments, including day length, temperature, and rainfall were evident in all the studies, and accordingly, seed production varied. The accessions grown in Japan yielded the highest number of grains per panicle. However, 11 accessions cultivated in Japan produced no harvest. Conversely, all accessions grown in parts of India, such as in Coimbatore, produced seeds. Therefore, although seed production in Japan was superior, there were benefits to cultivating the crops in India, including longer cultivation periods and the ability to overcome difficulties with seed multiplication found in Japan. Comparative cultivation projects involving international collaborative research are essential to reveal seed productivity in genetically diverse resources of sorghum. This study provides data about international sorghum production and information about differences in available accessions.
Journal of Virology
A soluble system was developed that could support DNA replication in simian virus 40 (SV40) chrom... more A soluble system was developed that could support DNA replication in simian virus 40 (SV40) chromosomes. DNA synthesis in this system required the presence of purified SV40 large tumor antigen, SV40 chromosomes prepared from virus-infected monkey cells, a crude extract from HeLa cells, and several low-molecular-weight components. In comparison to the replication of purified SV40 form I DNA, the rate of DNA synthesis was 15 to 20% in this system. DNA synthesis started near the replication origin of SV40 and proceeded bidirectionally in a semiconservative manner. Micrococcal nuclease digestion experiments revealed that the replicated DNA produced in this system became organized into a regularly spaced array of nucleosome core particles when an appropriate amount of purified HeLa core histones was added to the reaction mixture. SV40 form I DNA replicating under the same conditions was also assembled into nucleosomes, which were arranged in a rather dispersed manner and formed an aberra...
Cancer Research
Poly(ADP-ribose) glycohydrolase (PARG) is the main enzyme responsible for catabolism of poly(ADP-... more Poly(ADP-ribose) glycohydrolase (PARG) is the main enzyme responsible for catabolism of poly(ADP-ribose) (PAR), synthesized by PARP. PARG dysfunction sensitizes certain cancer cells to alkylating agents and cisplatin by perturbing the DNA damage response. The gene mutations that sensitize cancer cells to PARG dysfunction-induced death remain to be identified. Here, we performed a comprehensive analysis of synthetic lethal genes using inducible PARG knockdown cells and identified dual specificity phosphatase 22 (DUSP22) as a novel synthetic lethal gene related to PARG dysfunction. DUSP22 is considered a tumor suppressor and its mutation has been frequently reported in lung, colon, and other tumors. In the absence of DNA damage, dual depletion of PARG and DUSP22 in HeLa and lung cancer A549 cells reduced survival compared to single-knockdown counterparts. Dual depletion of PARG and DUSP22 increased the apoptotic sub-G1 fraction and upregulated PUMA in lung cancer A549, PC14, and SBC5 cells, and inhibited the PI3K/AKT/mTOR pathway in A549 cells, suggesting that dual depletion of PARG and DUSP22 induced apoptosis by upregulating PUMA and suppressing the PI3K/AKT/mTOR pathway. Consistently, the growth of tumors derived from double knockdown A549 cells was slower compared with those derived from control siRNA transfected cells. Taken together, these results indicate that DUSP22 deficiency exerts a synthetic lethal effect when combined with PARG dysfunction, suggesting that DUSP22 dysfunction could be a useful biomarker for cancer therapy using PARG inhibitors.
Japan Agricultural Research Quarterly: JARQ
Molecular and Cellular Biology
Identification of a novel mouse nuclear protein termed activator of basal transcription 1 (mABT1)... more Identification of a novel mouse nuclear protein termed activator of basal transcription 1 (mABT1) that associates with the TATA-binding protein (TBP) and enhances basal transcription activity of class II promoters is described. We also identify mABT1 homologous counterparts in Caenorhabditis elegans and Saccharomyces cerevisiae and show the homologous yeast gene to be essential for growth. The mABT1 associated with TBP in HeLa nuclear extracts and with purified mouse TBP in vitro. In addition, ectopically expressed mABT1 was coimmunoprecipitated with endogenous TBP in transfected cells. More importantly, mABT1 significantly enhanced transcription from an adenovirus major late promoter in a reconstituted cell-free system. We furthermore demonstrate that mABT1 consistently enhanced transcription from a reporter gene with a minimal core promoter as well as from reporter genes with various enhancer elements in a cotransfection assay. Taken together, these results suggest that mABT1 is a...
Nucleic Acids Symposium Series
Current Protein & Peptide Science, 2016
Poly(ADP-ribose) polymerases (PARPs) family proteins catalyze poly(ADP-ribosylation) (PARylation)... more Poly(ADP-ribose) polymerases (PARPs) family proteins catalyze poly(ADP-ribosylation) (PARylation) by conjugating ADP-ribose residues repeatedly on amino acid residues using nicotinamide adenine dinucleotide as a substrate. The inhibitors of PARP widely block DNA repair processes and are currently examined in clinical trials of cancer therapy. Poly(ADP-ribose) glycohydrolase (PARG) is the main nuclear enzyme, which digests poly(ADP-ribose) into ADP-ribose. PARG inhibitor could also be considered as a chemotherapeutic agent for cancer, because of its involvement in DNA repair. Various PARG inhibitors with IC50 value of micromolar to submicromolar range have been reported. However, for most of these chemicals, the specificity of inhibition has not been fully evaluated. PARG functional inhibition models in various organisms have been developed. Here, inducible PARG knockdown system was developed in HeLa cells and the cell line will be useful for identifying the synthetic lethal genes or affecting genes for PARG inhibitor treatment and also for functional elucidation of PARP superfamily molecules.
Biochemical and Biophysical Research Communications, 2010
<b>Copyright information:</b>Taken from "Loss of affects gene expression profile... more <b>Copyright information:</b>Taken from "Loss of affects gene expression profile in a genome-wide manner in ES cells and liver cells"http://www.biomedcentral.com/1471-2164/8/227BMC Genomics 2007;8():227-227.Published online 10 Jul 2007PMCID:PMC1959195.nes (A) & (B). Horizontal and vertical axes represent expression levels normalized for an individual gene. Each point represents normalized expression data for an individual gene. The genes that showed standard deviation greater than 2.0 in the normalized data of both genotypes (A) were excluded and gene lists were constructed with < 0.05 (B). Fig. 1D–F in the original article [1] remains unchanged and is presented as (C) – (E), respectively.
Transcriptional activation and repression are a key step in the regulation of all cellular activi... more Transcriptional activation and repression are a key step in the regulation of all cellular activities. The development of comprehensive analysis methods such as DNA microarray has advanced our understanding of the correlation between the regulation of transcription and that of cellular mechanisms. However, DNA microarray analysis based on steady-state mRNA (total mRNA) does not always correspond to transcriptional activation or repression. To comprehend these transcriptional regulations, the detection of nascent RNAs is more informative. Although the nuclear run-on assay can detect nascent RNAs, it has not been fully applied to DNA microarray analysis. In this study, we have developed a highly efficient method for isolating bromouridine-labeled nascent RNAs that can be successfully applied to DNA microarray analysis. This method can linearly amplify small amounts of mRNAs with little bias. Furthermore, we have applied this method to DNA microarray analysis from mouse G2-arrested cel...
The Molecular Biology Society of Japan, 2016
Genomic DNA contains not only information of DNA sequence, but also epigenetic information that i... more Genomic DNA contains not only information of DNA sequence, but also epigenetic information that is the direct DNA modification by methylation (the addition of methyl group to the 5th carbon of pyrimidine ring of cytosine) and histone modifications (acetylation, methylation, etc). Epigenetic information is closely related to regulation of gene expression. If a methyl group is dislocated to position 5 of the pyrimidine ring of cytosine, the hydrogen bond between complementary GC bases will not be inhibited, but this methyl group is positioned so as to be exposed in the major groove of the double-helix structure of DNA, and according to the genome region/sequence undergoing modification of methylation, gene expression is inhibited by the interaction between the genome and DNAbinding molecules.
Genome Informatics, 1994
The DNA sequence of chromosome VI (270kb) of the budding yeast S. cerevisiae, has been determined... more The DNA sequence of chromosome VI (270kb) of the budding yeast S. cerevisiae, has been determined. The sequence data were then analyzed with the software GENETYX (Software Development Co.) to nd out the candidates of the coding region. As there is little number of intron in budding yeast genome, it is relatively easy to nd the candidates of the coding region. Open reading frames identi ed by the software were generally the same as the coding sequence with little exception. To carry out further analysis of the open reading frames, we tried to automate the process required for similarity search. The processes automated are
The Molecular Biology Society of Japan, 2016
SUMMARYHepatocytes can be used to study the pathogenesis of liver diseases and drug discovery res... more SUMMARYHepatocytes can be used to study the pathogenesis of liver diseases and drug discovery research. Human hepatocytes are, however, hardly expandable in vitro making it difficult to secure large numbers of cells from one donor. In this study, we aimed to establish an in vitro long-term culture system that enables stable proliferation and maintenance of human hepatocytes to ensure a constant supply. We purified human hepatocytes by selection with cytocidal puromycin, and cultured them for more than 60 population doublings over a span of more than 350 days. These results show that this simple culture system with usage of the cytocidal antibiotics enables efficient hepatocyte proliferation and is an effective method for generating a stable supply of hepatocytes for drug discovery research at a significant cost reduction.
Journal of Biological Chemistry
International Journal of Molecular Sciences
Cell division is essential for the maintenance of life and involves chromosome segregation and su... more Cell division is essential for the maintenance of life and involves chromosome segregation and subsequent cytokinesis. The processes are tightly regulated at both the spatial and temporal level by various genes, and failures in this regulation are associated with oncogenesis. Here, we investigated the gene responsible for defects in cell division by using murine temperature-sensitive (ts) mutant strains, tsFT101 and tsFT50 cells. The ts mutants normally grow in a low temperature environment (32 °C) but fail to divide in a high temperature environment (39 °C). Exome sequencing and over-expression analyses identified Diaph3, a member of the formin family, as the cause of the temperature sensitivity observed in tsFT101 and tsFT50 cells. Interestingly, Diaph3 knockout cells showed abnormality in cytokinesis at 39 °C, and the phenotype was rescued by re-expression of Diaph3 WT, but not Diaph1 and Diaph2, other members of the formin family. Furthermore, Diaph3 knockout cells cultured at 3...
Japan Agricultural Research Quarterly: JARQ
To compare the seed productivity of germplasm in India (Tamil Nadu) and Japan (Tsukuba), a total ... more To compare the seed productivity of germplasm in India (Tamil Nadu) and Japan (Tsukuba), a total of 105 accessions from the Japanese National Agriculture and Food Research Organization (NARO) sorghum core collection were cultivated. The comparative cultivation studies were conducted at two locations in India and one in Japan. Differences in cultivation environments, including day length, temperature, and rainfall were evident in all the studies, and accordingly, seed production varied. The accessions grown in Japan yielded the highest number of grains per panicle. However, 11 accessions cultivated in Japan produced no harvest. Conversely, all accessions grown in parts of India, such as in Coimbatore, produced seeds. Therefore, although seed production in Japan was superior, there were benefits to cultivating the crops in India, including longer cultivation periods and the ability to overcome difficulties with seed multiplication found in Japan. Comparative cultivation projects involving international collaborative research are essential to reveal seed productivity in genetically diverse resources of sorghum. This study provides data about international sorghum production and information about differences in available accessions.
Journal of Virology
A soluble system was developed that could support DNA replication in simian virus 40 (SV40) chrom... more A soluble system was developed that could support DNA replication in simian virus 40 (SV40) chromosomes. DNA synthesis in this system required the presence of purified SV40 large tumor antigen, SV40 chromosomes prepared from virus-infected monkey cells, a crude extract from HeLa cells, and several low-molecular-weight components. In comparison to the replication of purified SV40 form I DNA, the rate of DNA synthesis was 15 to 20% in this system. DNA synthesis started near the replication origin of SV40 and proceeded bidirectionally in a semiconservative manner. Micrococcal nuclease digestion experiments revealed that the replicated DNA produced in this system became organized into a regularly spaced array of nucleosome core particles when an appropriate amount of purified HeLa core histones was added to the reaction mixture. SV40 form I DNA replicating under the same conditions was also assembled into nucleosomes, which were arranged in a rather dispersed manner and formed an aberra...
Cancer Research
Poly(ADP-ribose) glycohydrolase (PARG) is the main enzyme responsible for catabolism of poly(ADP-... more Poly(ADP-ribose) glycohydrolase (PARG) is the main enzyme responsible for catabolism of poly(ADP-ribose) (PAR), synthesized by PARP. PARG dysfunction sensitizes certain cancer cells to alkylating agents and cisplatin by perturbing the DNA damage response. The gene mutations that sensitize cancer cells to PARG dysfunction-induced death remain to be identified. Here, we performed a comprehensive analysis of synthetic lethal genes using inducible PARG knockdown cells and identified dual specificity phosphatase 22 (DUSP22) as a novel synthetic lethal gene related to PARG dysfunction. DUSP22 is considered a tumor suppressor and its mutation has been frequently reported in lung, colon, and other tumors. In the absence of DNA damage, dual depletion of PARG and DUSP22 in HeLa and lung cancer A549 cells reduced survival compared to single-knockdown counterparts. Dual depletion of PARG and DUSP22 increased the apoptotic sub-G1 fraction and upregulated PUMA in lung cancer A549, PC14, and SBC5 cells, and inhibited the PI3K/AKT/mTOR pathway in A549 cells, suggesting that dual depletion of PARG and DUSP22 induced apoptosis by upregulating PUMA and suppressing the PI3K/AKT/mTOR pathway. Consistently, the growth of tumors derived from double knockdown A549 cells was slower compared with those derived from control siRNA transfected cells. Taken together, these results indicate that DUSP22 deficiency exerts a synthetic lethal effect when combined with PARG dysfunction, suggesting that DUSP22 dysfunction could be a useful biomarker for cancer therapy using PARG inhibitors.
Japan Agricultural Research Quarterly: JARQ
Molecular and Cellular Biology
Identification of a novel mouse nuclear protein termed activator of basal transcription 1 (mABT1)... more Identification of a novel mouse nuclear protein termed activator of basal transcription 1 (mABT1) that associates with the TATA-binding protein (TBP) and enhances basal transcription activity of class II promoters is described. We also identify mABT1 homologous counterparts in Caenorhabditis elegans and Saccharomyces cerevisiae and show the homologous yeast gene to be essential for growth. The mABT1 associated with TBP in HeLa nuclear extracts and with purified mouse TBP in vitro. In addition, ectopically expressed mABT1 was coimmunoprecipitated with endogenous TBP in transfected cells. More importantly, mABT1 significantly enhanced transcription from an adenovirus major late promoter in a reconstituted cell-free system. We furthermore demonstrate that mABT1 consistently enhanced transcription from a reporter gene with a minimal core promoter as well as from reporter genes with various enhancer elements in a cotransfection assay. Taken together, these results suggest that mABT1 is a...
Nucleic Acids Symposium Series
Current Protein & Peptide Science, 2016
Poly(ADP-ribose) polymerases (PARPs) family proteins catalyze poly(ADP-ribosylation) (PARylation)... more Poly(ADP-ribose) polymerases (PARPs) family proteins catalyze poly(ADP-ribosylation) (PARylation) by conjugating ADP-ribose residues repeatedly on amino acid residues using nicotinamide adenine dinucleotide as a substrate. The inhibitors of PARP widely block DNA repair processes and are currently examined in clinical trials of cancer therapy. Poly(ADP-ribose) glycohydrolase (PARG) is the main nuclear enzyme, which digests poly(ADP-ribose) into ADP-ribose. PARG inhibitor could also be considered as a chemotherapeutic agent for cancer, because of its involvement in DNA repair. Various PARG inhibitors with IC50 value of micromolar to submicromolar range have been reported. However, for most of these chemicals, the specificity of inhibition has not been fully evaluated. PARG functional inhibition models in various organisms have been developed. Here, inducible PARG knockdown system was developed in HeLa cells and the cell line will be useful for identifying the synthetic lethal genes or affecting genes for PARG inhibitor treatment and also for functional elucidation of PARP superfamily molecules.
Biochemical and Biophysical Research Communications, 2010