A study of BP1 function in lymphocyte development through a generation of BP1-gene knock-out mice.

通过一代 BP1 基因敲除小鼠研究 BP1 在淋巴细胞发育中的功能。

• 批准号: 06670358
• 负责人: KITAMURA Daisuke
• 金额: $ 1.41万
• 依托单位: SCIENCE UNIVERSITY OF TOKYO (1995)Kyushu University (1994)
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for General Scientific Research (C)
• 财政年份: 1994
• 资助国家: 日本
• 起止时间: 1994 至 1995
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-06670358/
• 关键词: BP1; aminopeptidase A; gene targeting; B cell development; pre-B cell; knock-out mouse; エクトエンザイム; ジーンターゲティング

BP1 is a 140 kD type-II transmembrane glyoprotein expressed as a homodimer on a surface of precursor (pre-) B cells selectively in a lymphoid-cell lineage. Cloning and analysis of a cDNA cording for BP1 revealed its identity as aminopeptidase A (APA), a Zn^<2+>/Ca^<2+>-dependent peptidase present in a broad range of organs like kidney or small intestine. A well known function of APA is to convert angiotensin II into less active angiotensin III, thus involved in regulation of blood pressure. Also, APA is probably involved in digestion of peptides in small intestine. In the present study, a possible role of BP1 in B cell development was assessed by generating and analyzing BP1-gene knock-out mice. Lymphocyte cellularity of bone marrow, thymus, spleen and lymph-nodes in the mutant mice was grossly normal when analyzed by flow cytometry after staining the cells with a panel of monoclonal antibodies against various cell-surface antigens. Proliferative responses to cytokines such as interleu … More kin (IL) -7, IL-3 and GM-CSF of the BP1-deficient bone marrow cells were comparable to normal ones, indicating that development of lymphoid and myeloid progenitors is independent of BP1 protein. Mature B cells from spleens of the mutants also normally responded to anti-IgM antibody, CD40-ligand and LPS, suggesting that functional competernce of such cells are unaffected. Finally, B-cell progenitors of the mutants immigrated into the bone marrow of the recipient mice to the same extent as those of wild-type mice and also observed was equal contribution of the donor cells in the mutant and wild-type recipients in a bone-marrow chimera experiment. Thus, although BP1 is expressed in pre-B cells and bone-marrow stroma cells supporting the growth of the pre-B cells, BP1 is dispensable for the development, homing and maturation of pre-B cells. The BP1-deficient mice were healthy, fertile, grew up normally and no gross deformity of organs were noticed. Therefore roles of BP1 in other organs were not obvious. Genetic redundancy of metalopeptidase may possibly have concealed the overt phenotype of the BP1-deficiency. Alternative reverse genetic approaches such as dominant-negative transgenic or inducible gene knock-out mice as well as an identification of physiological substrates of BP1 would be necessary to disclose a real function of BP1. Less

BP1是一种140 kD的ii型跨膜糖蛋白，在淋巴细胞谱系中选择性地在前体（前）B细胞表面以同型二聚体表达。BP1的cDNA序列克隆和分析表明其为氨基肽酶a (APA)，一种锌<2+>/钙<2+>依赖性肽酶，广泛存在于肾脏或小肠等器官中。众所周知，APA的功能是将血管紧张素II转化为活性较低的血管紧张素III，从而参与血压调节。此外，APA可能参与小肠内多肽的消化。在本研究中，通过生成和分析BP1基因敲除小鼠来评估BP1在B细胞发育中的可能作用。用一组抗各种细胞表面抗原的单克隆抗体染色后，流式细胞术分析突变小鼠的骨髓、胸腺、脾脏和淋巴结的淋巴细胞结构基本正常。BP1缺失的骨髓细胞对interleu、IL -7、IL-3和GM-CSF等细胞因子的增殖反应与正常细胞相当，表明淋巴细胞和髓细胞祖细胞的发育不依赖于BP1蛋白。来自突变体脾脏的成熟B细胞也对抗igm抗体、cd40配体和LPS有正常反应，表明这些细胞的功能能力不受影响。最后，在骨髓嵌合体实验中，突变体的b细胞祖细胞迁移到受体小鼠骨髓的程度与野生型小鼠相同，并且在突变体和野生型受体中也观察到供体细胞的贡献相等。因此，尽管BP1在支持前b细胞生长的前b细胞和骨髓基质细胞中表达，但BP1对于前b细胞的发育、归巢和成熟是必不可少的。bp1缺陷小鼠健康、可育、发育正常，无明显脏器畸形。因此BP1在其他器官中的作用不明显。超肽酶的遗传冗余可能掩盖了bp1缺乏的显性表型。替代的反向遗传方法，如显性阴性转基因或诱导基因敲除小鼠，以及BP1生理底物的鉴定，对于揭示BP1的真实功能是必要的。少

项目成果

Yamanashi,Y.: "Identification of HS1 protein as a major substrate of protein-tyrosine kinase(s) upon B-cell antigen receptor-mediated signaling." Natl.Acad.Sci.USA. 90. 3631-3635 (1993)

Yamanashi,Y.：“根据 B 细胞抗原受体介导的信号，鉴定 HS1 蛋白为蛋白酪氨酸激酶的主要底物。”

Kitamura,D.: "Molecular cloning and characterization of mouse HS1." Biochem.Biophysic.Res.Commun.(in press).

Kitamura,D.：“小鼠 HS1 的分子克隆和表征。”