Normal Immune Development and Glucocorticoid-Induced Thymocyte Apoptosis in Mice Deficient for the T-Cell Death-Associated Gene 8 Receptor

Abstract

T-cell death-associated gene 8 (TDAG8) is a G-protein-coupled receptor transcriptionally upregulated by glucocorticoids (GCs) and implicated by overexpression studies in psychosine-mediated inhibition of cytokinesis and in GC-induced apoptosis. To examine the physiological function of TDAG8, we generated knockout (KO) mice by homologous recombination. An enhanced green fluorescent protein reporter was knocked into the disrupted tdag8 locus to allow the analysis of TDAG8 expression in living cells. Interestingly, we found that during thymocyte development, TDAG8 expression resembled the dynamic regulation described for known modulators of GC-induced apoptosis, including Bcl-2, Notch1, and GC receptor. TDAG8 was expressed in double-negative cells, was downregulated at the double-positive transition, and was upregulated in single-positive thymocytes. However, despite this striking expression pattern, maturation and selection of thymocytes, as well as major immune functions, were not affected in TDAG8 KO mice. In contrast to previous overexpression results, TDAG8 was dispensable for psychosine-induced formation of multinucleated cells. Furthermore, TDAG8 KO thymocytes showed normal apoptosis following in vivo and in vitro GC treatment. These results, while establishing a useful reporter strain to study T-lymphocyte maturation, argue against a critical role for TDAG8 in immune development, psychosine-mediated inhibition of cytokinesis, and GC-induced cell death.

We thank Shirley Quan for outstanding technical assistance; LaKeisha Perkins, Mathew Au, and Rosalyn Taijeron for maintaining the mouse colony; and Renee L. H. Lim for designing the TDAG8 targeting construct. We also thank Barbara Anderson for excellent preparation of the manuscript and Chengyi J. Shu for help with CCD imaging. We are indebted to Yongwon Choi (The Rockefeller University) for advice and support during the initial phases of this project.

O.N.W. is an Investigator of the Howard Hughes Medical Institute. C.G.R. was supported by a Cancer Research Institute Fellowship during a portion of these studies. A.N. was supported by the Howard Hughes Medical Institute Research Training Fellowship for Medical Students.