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IgM H- and L-chains artificially and excessively accumulated in HDAC2(-/-) DT40 mutants are gradually and dramatically reduced in distinct ways in individual mutant clones through various generations during continuous cultivation
http://hdl.handle.net/10458/6504
http://hdl.handle.net/10458/6504573f01a4-2b57-47f8-961e-0a78f40595e9
| Item type | その他 / Others(1) | |||||
|---|---|---|---|---|---|---|
| 公開日 | 2020-06-21 | |||||
| タイトル | ||||||
| タイトル | IgM H- and L-chains artificially and excessively accumulated in HDAC2(-/-) DT40 mutants are gradually and dramatically reduced in distinct ways in individual mutant clones through various generations during continuous cultivation | |||||
| 言語 | en | |||||
| 言語 | ||||||
| 言語 | eng | |||||
| キーワード | ||||||
| 言語 | en | |||||
| 主題Scheme | Other | |||||
| 主題 | Gene targeting techniques, HDAC2-deficient DT40 mutants, continuous cultivation, decreases in IgM H/L chain protein/mRNA levels, changes in mRNA levels of transcription factors in individual mutant clones, changes in mRNA levels of transcription factors, gene targeting techniques, ChIP assay, changes in acetylation levels of K9/H3 surrounding Pax5 5'-upstream chromatin region | |||||
| 資源タイプ | ||||||
| 資源タイプ識別子 | http://purl.org/coar/resource_type/c_1843 | |||||
| 資源タイプ | other | |||||
| 著者 |
中山, 雅美
× 中山, 雅美× 中山, 建男 |
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| 抄録 | ||||||
| 内容記述タイプ | Abstract | |||||
| 内容記述 | We newly generated histone deacetylase-2 （HDAC2）-deficient DT40 mutants HDAC2（-/-） and analyzed their characteristics at various intervals during continuous cultivation. In six independent clones cl.2-1, cl.2-2, cl.2-3, cl.2-4, cl.2-5 and cl.2-6 of HDAC2（-/-） mutants, proteins and mRNAs of IgM H- and L-chains were dramatically increased at the early stage of cultivation, and thereafter decreased in almost similar changing pattern and at the later stage reached comparable levels as in DT40 cells. By contrast, mRNAs of various transcription factors and chromatin-modifying enzymes showed distinct changing patterns in these six HDAC2（-/-） clones during cultivation. In clone cl.2-1, mRNAs of Pax5, Aiolos and EBF1 were almost completely decreased at the early stage and remained unchanged during cultivation, but that of OBF1 was dramatically decreased until the later stage. In clones cl.2-2, cl.2-3, cl.2-4 and cl.2-5, mRNAs of Pax5, Aiolos and EBF1 were almost completely decreased at the early stage, and thereafter mRNAs of Pax5 and Aiolos were increased until the later stage but that of EBF1 remained unchanged during cultivation. In clone cl.2-6, mRNAs of Pax5, Aiolos and EBF1 were almost completely decreased at the early stage and thereafter dramatically increased during cultivation. These findings suggested that three distinct ways of gene expressions of IgM H- and L-chains exist at the later stage in these six individual HDAC2（-/-） clones; i.e., clone cl.2-1 seems the OBF1-dependent type, clones cl.2-2, cl.2-3, cl.2-4 and cl.2-5 seem the Pax5- and Aiolos-dependent type, and clone cl.2-6 seems the Pax5-, Aiolos- and EBF1-dependent type. Based on these results, we proposed a hypothesis that individual clones of HDAC2（-/-） mutants possess the ability to gain new cell function to exclude excessively accumulated IgM H- and L-chains （as an abnormal and uncomfortable environment change） based on their decreased gene expressions through distinct ways depending on alterations in gene expressions of Pax5, Aiolos, EBF1, OBF1 and others through various generations during continuous cultivation. | |||||
| 言語 | en | |||||
| 抄録 | ||||||
| 内容記述タイプ | Abstract | |||||
| 内容記述 | We analyzed histone deacetylase-2 (HDAC2)-deficient DT40 mutant cells, HDAC2(-/-), which were cultivated for varying periods, by two-dimensional polyacrylamide gel electrophoresis (2D-PAGE), Western blotting, reverse transcription-polymerase chain reaction (RT-PCR) and immuno-electron microscopy. Protein and mRNA levels of IgM H- and L-chains were increased at the early stage of cultivation and thereafter until the later stage reduced to comparable levels as in DT40 cells. The mRNA level of PCAF was increased from the early to later stages, and those of HDAC7 and HDAC9 changed during cultivation. The mRNA level of EBF1 was almost completely decreased at the early stage and thereafter remained unchanged, but that of E2A was slightly increased at the early stage and thereafter remained unchanged. The mRNA level of Pax5, which down-regulates gene expressions of IgM H- and L-chains, was reduced at the early stage and thereafter increased through the middle to later stages in anti-parallel with those of these two immunoglobulin proteins. Bulk acetylation levels of Lys-9, Lys-14, Lys-18, Lys-23 and Lys-27 residues of histone H3 (K9/H3, K14/H3, K18/H3, K23/H3 and K27/H3) were increased during cultivation. Chromatin immuno-precipitation (ChIP) assay suggested that acetylation levels of K9/H3 within some chromatin segments surrounding proximal ~2.0 kb 5'-upstream region of the Pax5 gene were decreased at the early stage in HDAC2(-/-) and thereafter at the later stage increased to comparable levels as in DT40 cells. These results revealed not only that the HDAC2-deficiency induces dramatic increases in mRNA and protein levels of IgM H- and L-chains, and thereafter these accumulated mRNAs and proteins are gradually reduced during cultivation, but also that alterations in gene expressions of IgM H- and L-chains may be coupled with changes in those of Pax5, EBF1, PCAF, HDAC7 and HDAC9. In addition, the changing pattern in acetylation levels of K9/H3 within proximal 5'-upstream chromatin region of the Pax5 gene in HDAC2(-/-) qualitatively agrees with that in the transcription level of the gene. | |||||
| 内容記述 | ||||||
| 内容記述タイプ | Other | |||||
| 内容記述 | The original edition of the revision is available from following URL: http://opac2.lib.miyazaki-u.ac.jp/webopac/TC10145178 | |||||
| 言語 | en | |||||
| 著者版フラグ | ||||||
| 出版タイプ | AM | |||||
| 出版タイプResource | http://purl.org/coar/version/c_ab4af688f83e57aa | |||||
