Cloning of Human Stat5B RECONSTITUTION OF INTERLEUKIN-2-INDUCED Stat5A AND Stat5B DNA BINDING ACTIVITY IN COS-7 CELLS

Jian-Xin Lin; Judy Mietz; William S. Modi; Susan John; Warren J. Leonard

doi:10.1074/jbc.271.18.10738

Cloning of Human Stat5B

RECONSTITUTION OF INTERLEUKIN-2-INDUCED Stat5A AND Stat5B DNA BINDING ACTIVITY IN COS-7 CELLS (*)

From the (1)Laboratory of Molecular Immunology, NHLBI, National Institutes of Health, Bethesda, Maryland 20892 and the
(2)Biological Carcinogenesis and Development Program, Science Applications International Corp., NCI-Frederick Cancer Research and Development Center, Frederick, Maryland 21702-1201

↵^§ To whom correspondence should be addressed: Lab. of Molecular Immunology, NHLBI, NIH, 9000 Rockville Pike, Bethesda, MD 20892-1674. Tel.: 301-496-0098; Fax: 301-402-0971.

Abstract

We have isolated a second human Stat5 cDNA, Stat5B, and demonstrated that the genes encoding both Stat5A and Stat5B are located at chromosome 17q11.2. Both genes were constitutively transcribed in peripheral blood lymphocytes. By using specific antisera, we demonstrated that both Stat5A and Stat5B are activated by interleukin-2 (IL-2) in peripheral blood lymphocytes, natural killer-like YT leukemia cells, and human T cell lymphotropic virus type I-transformed MT-2 T cells. In COS-7 cells, which constitutively express the Janus family tyrosine kinase Jak1, reconstitution of IL-2-induced Stat5A and Stat5B DNA binding activities was dependent on the coexpression of Jak3 along with the IL-2 receptor β chain and the common cytokine receptor -chain. This IL-2-induced Stat5 activation was dependent on the presence of either of two tyrosines (Tyr-392 or Tyr-510) in the IL-2 receptor β chain, indicating that either of these two tyrosines can serve as a docking site. Moreover, we demonstrated that human Stat5 activation is also dependent on Tyr-694 in Stat5A and Tyr-699 in Stat5B, indicating that these tyrosines are required for dimerization. The COS-7 reconstitution system described herein provides a valuable assay for further elucidation of the IL-2-activated JAK-STAT pathway.

Footnotes

↵* The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore by hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.