Analysis of mechanisms of the differentiation inhibition in erythroid cells induced by Ets transcription factor PU. I

Ets转录因子PU诱导红系细胞分化抑制机制分析。

• 批准号: 12671015
• 负责人: YAMADA Toshiyuki
• 金额: $ 1.73万
• 依托单位: Sasaki Institute
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2000
• 资助国家: 日本
• 起止时间: 2000 至 2001
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-12671015/
• 关键词: PU.l; murine erythroleukemia (MEL); differentiation inhibition; lineage switch; CKLiK gene; CKLiK遺伝子

To identify a gene(s) involved in the differentiation inhibition of murine erythroleukemia (MEL) cells induced by PU. 1, we had screened the genes whose expression is up- or downregulated in MEL cells after overexpression of PU. 1 by using the mRNA differential display (DD) strategy. Based on the results of the screening, we have extended the study as follows.(1) Because DD had shown that the expression of some of myelomonocyte-specific genes is up-regulated, we expanded analysis and demonstrated that expression of a variety of myelomonocyte-specific genes is up-regulated. Furthermore, following overexpression of PU. 1, MEL cells became adherent and phagocytic. On the other hand, expression of myelomonocyte-specific genes was not induced when a mutant PU. 1, with part of the activation domain deleted, which also inhibits erythroid differentiation of MEL cells was expressed. These results indicate that PU. 1 induces lineage switch in MEL cells toward myelomonocytic cells and suggest that the pathway of lineage switch is distinct from that of inhibition of the erythroid differentiation of the cells.(2) One of the novel genes whose expression is up-regulated in MEL cells after overexpression of PU. 1 was found to be expressed normally in T cells and embryonal carcinoma cells. We have cloned this gene. As an open reading frame was identified with homology to human calcium-calmodelin-dependent kinase I-like kinase (CKLiK) gene, the novel gene was thought to be the mouse homologue of human CKLiK gene. However, the nucleotide sequence of 3 portion of the open reading frame was diverged from that of human CKLiK gene Two kinds of transcripts showmg difference in their 3 ends, which is presumably due to alternative splicing, were present We are now planning to investigate the functional role of the novel gene in the PU. 1-mediated differentiation inhibition of MEL cells and involvement of PU.1 in transcriptional regulation of the gene.

目的：鉴定一个参与PU诱导小鼠红细胞白血病（MEL）细胞分化抑制的基因。1、筛选过表达PU后MEL细胞中表达上调或下调的基因。1通过mRNA差异显示（DD）策略。根据筛选结果，我们将研究扩展如下：(1)由于DD已经表明部分髓单核细胞特异性基因的表达上调，我们扩大分析，证实多种髓单核细胞特异性基因的表达上调。此外，PU过表达后。1、MEL细胞贴壁并吞噬。另一方面，当PU突变时，不诱导骨髓单核细胞特异性基因的表达。1，部分激活域缺失，也抑制了MEL细胞的红系分化。这些结果表明，PU。1诱导MEL细胞向髓系细胞的谱系转换，提示谱系转换的途径不同于抑制细胞红细胞分化的途径。(2) MEL细胞中PU过表达后表达上调的新基因之一。1在T细胞和胚胎癌细胞中正常表达。我们已经克隆了这个基因。由于一个开放阅读框与人钙-calmodelin依赖性激酶i样激酶（CKLiK）基因同源，该新基因被认为是人类CKLiK基因的小鼠同源基因。然而，开放阅读框的3个部分的核苷酸序列与人类的CKLiK基因不同，两种转录本在它们的3端显示差异，这可能是由于选择性剪接，我们现在计划研究新基因在PU中的功能作用。1介导的MEL细胞分化抑制和PU.1参与该基因的转录调控。

项目成果

Kihara-Negichi F, Yamada T. et al.: "In vivo complex formation of PU.1 with HDAC1 associated with PU.1-mediated transcriptional repression"Oncogene. 20. 6039-6047 (2001)

Kihara-Negichi F、Yamada T.等人：“PU.1与HDAC1的体内复合物形成与PU.1介导的转录抑制有关”癌基因。

Yamada T. et al.: "Lineage switch induced by overexpression of Ets family transcription factor PU.1 in murine erythroleukemia cells"Blood. 97. 2300-2307 (2001)

Yamada T. 等人：“小鼠红白血病细胞中 Ets 家族转录因子 PU.1 过度表达诱导的谱系转换”血液。

Oikawa T, Yamada T.et al.: "Extinction of expression of the genes encoding hematopoietic cell-restricted transcription factors in T lymphoma × fibroblast cell hybrids"Immunology. 104. 164-167 (2001)

Oikawa T、Yamada T. 等人：“T 淋巴瘤 × 成纤维细胞杂种中编码造血细胞限制性转录因子的基因的表达消失”《免疫学》104. 164-167 (2001)。

Kihara-Negishi F, Yamada T.et al.: "In vivo complex formation of PU.1 with HDAC1 associated with PU.1-mediated transcriptional repression"Oncogene. 20. 6039-6047 (2001)

Kihara-Negishi F、Yamada T.等人：“PU.1 与 HDAC1 的体内复合物形成与 PU.1 介导的转录抑制有关”癌基因。

Yamada,T., et al.: "Lineage switch by overexpression of Ets family transcription factor PU.1 in murine erythroleukemia cells"Blood. (印刷中).

Yamada, T. 等人：“通过在小鼠红白血病细胞中过度表达 Ets 家族转录因子 PU.1 进行谱系转换”（正在出版）。