The characterization of the monoclonal antibody Th‐10a, specific for a nuclear protein appearing in the S phase of the cell cycle in normal thymocytes and its unregulated expression in lymphoma cell lines

Cell Proliferation - Tập 28 Số 12 - Trang 645-657 - 1995
Masahiro Muto1, Masanori Utsuyama2, Taiga Horiguchi3, Eiko Kubo4, Toshihiko Sado4, Katsuiku Hirokawa5
1Division of Biology and Oncology, National Institute of Radiological Science, Chiba, Japan.
2Department of Pathology, Tokyo Metropolitan Institute of Gerontology, Tokyo
3Department of Biological Science, Tokyo Institute of Technology, Tokyo
4Division of Biology and Oncology, National Institute of Radiological Science, Chiba
5Department of Pathology, School of Medicine, Tokyo Medical and Dental University, Tokyo, Japan

Tóm tắt

AbstractA monoclonal antibody (Th‐10a) specific for the nuclear protein appearing in the S phase of the cell cycle in normal mouse thymocytes was derived by immunizing Wistar rats with a murine thymic lymphoma (TIGN), and its isotype was rat IgG2a and had κ light chain. Immunohistochemical staining of frozen sections of B10.Thy1.1 newborn thymus and embryonic intestine revealed that this monoclonal antibody reacted strongly with the nuclear proteins of subcortical thymocytes and the basal layer of the mucosa, where many cells were dividing, but not with that of the thymic medullary area. To evaluate the expression of the nuclear proteins during the cell cycle in detail, the results of an immunofluorescence analysis of the thymocytes from hydroxyurea‐treated B10 mice using Th‐10a monoclonal antibody were compared with those of DNA synthesis of these cells with the use of the FITC‐conjugated anti‐BrdUrd monoclonal antibody. The results indicated that the nuclear protein detected by Th‐10a monoclonal antibody was highly expressed in the S phase of normal thymocytes, while the cells in G1, G2 and M phases exhibited a low level of the expression. Moreover, the variations in expression of the nuclear proteins in the thymocytes at different times after hydroxyurea treatment were observed to correspond with the frequency of DNA synthesizing cells. In contrast, the high level and unregulated expression of the nuclear protein detected by Th‐10a monoclonal antibody was observed throughout the cell cycle of the mouse lymphoma cell lines examined. Since Th‐10a monoclonal antibody does not react with the nuclear proteins derived from human, hamster or rat proliferating cells, this antibody may recognize a murine specific epitope of the nuclear protein.To further characteize the nuclear proteins, we extracted them from normal thymocytes or thymic lymphomas, and analysed them by immunoblotting or***

Từ khóa


Tài liệu tham khảo

10.1002/j.1460-2075.1993.tb05678.x

10.1002/j.1460-2075.1992.tb05270.x

Bianchi V, 1986, Changes in deoxynucleoside triphosphate pools induced by hydroxyurea and their relation to DNA synthesis, J. Biol. Chem., 261, 16037, 10.1016/S0021-9258(18)66672-4

10.1016/0022-1759(82)90178-8

10.1111/j.1365-2184.1992.tb01435.x

10.1073/pnas.82.10.3262

Danova M, 1990, Cell cycle‐related proteins and flow cytometry. (Review), Haematologica, 75, 252

10.1073/pnas.80.18.5573

Dou QP, 1993, Cyclin E and cyclin A as candidates for the restriction point protein, Cancer Res., 53, 1493

Dou QP, 1994, G1/S‐regulated E2F‐containing protein complexes bind to the mouse thymidine kinase gene promoter, J. Biological Chemistry, 269, 1306, 10.1016/S0021-9258(17)42259-9

10.1111/j.1365-2184.1994.tb01421.x

10.1016/0092-8674(89)90839-8

10.1016/0145-2126(91)90101-X

10.1128/MCB.6.5.1579

Hirokawa K, 1986, Analysis of the thymic microenvironment by monoclonal antibodies with special reference to thymic nurse cells, Thymus, 8, 349

Hodgson GS, 1975, Recovery of proliferating haemopoietic progenitor cells after killing by hydroxyurea, Cell Tissue Kinet, 8, 51

Johnstone A, 1987, Immunochemistry in Practice, 183

Juven T, 1993, Wild type p53 can mediate sequence‐specific transactivation of an internal promoter within the mdm2 gene, Oncogene, 8, 3411

Kaczmarek L, 1986, Protooncogene expression during the cell cycle, Lab. Invest., 15, 965

10.1073/pnas.90.3.1112

10.1016/0008-8749(91)90067-L

10.1016/0092-8674(92)90644-R

Muto M, 1990, Phenotypic characterization of thymic prelymphoma cells of B10 mice treated with split‐dose irradiation, J. Immunol., 144, 849, 10.4049/jimmunol.144.3.849

10.1016/0014-4827(87)90020-6

Ogata K, 1985, Purification and N‐terminal amino acid sequence of proliferating cell nuclear antigen (PCNA)/cyclin and development of ELISA for anti‐PCNA antibodies, J. Immunol., 135, 2623, 10.4049/jimmunol.135.4.2623

10.1002/jcb.240540403

Penit C, 1988, Sequential events in thymocyte differentiation and thymus regeneration revealed by a combination of bromodeoxyuridine DNA labeling and antimitotic drug treatment, J. Immunol., 140, 3315, 10.4049/jimmunol.140.10.3315

10.1111/j.1365-2184.1994.tb01474.x

10.1111/j.1432-1033.1984.tb08440.x

10.1016/0022-1759(86)90319-4

Thông tin
Thông tin xuất bản

Cell Proliferation

Tập 28 Số 12

645-657

Thông tin tác giả