PURIFICATION, STRUCTURAL-ANALYSIS, AND FUNCTION OF NATURAL ATAC, A CYTOKINE SECRETED BY CD8(-CELLS() T)

Recently, we identified a novel putative human cytokine expressed by a ctivated CD8(+) T cells, which was designated ATAC (activation-induced , T cell-derived, and chemokine-related; the same molecule has been id entified independently as lymphotactin and single cysteine motif-1). I n this report, we provide evidence that ATAC is a secreted 93-amino ac id protein that is generated from its precursor by proteolytic cleavag e between Gly(21) and Val(22). An estimated 60% of ATAC (Val(22)-Gly(1 14)) is secreted as an unmodified protein with a molecular mass of 10, 271.72 Da (apparent molecular mass of 12 kDa in SDS-polyacrylamide gel electrophoresis) and in which Cys(32) and Cys(69) are linked by a dis ulfide bridge. Unmodified ATAC is a cationic protein with a pI of 11.3 5 and is capable of binding to heparin. Some 40% of ATAC is O-glycosyl ated within 25 min of synthesis, giving rise to the appearance of a ho mogeneous 15-kDa (minor fraction) and a heterogeneous, terminally sial ylated 17-19-kDa (major fraction) protein species in SDS-polyacrylamid e gel electrophoresis. The secretion of all ATAC protein variants is c ompleted within 30-40 min of synthesis. In terms of function, various ATAC protein forms were consistently ineffective in chemotaxis assays. In contrast, both purified natural ATAC and a chemically synthesized aglycosyl analog induced locomotion (chemokinesis) in purified CD4(+) and CD8(+) T cell populations at 400 ng/ml.