Y. Churcher et Se. Moss, MODULATION OF PROTEIN-TYROSINE PHOSPHORYLATION DURING G(1) S TRANSITION IN ACTIVATED HUMAN T-LYMPHOBLASTS/, The Journal of biological chemistry, 268(35), 1993, pp. 26144-26149
We report that in activated human T-lymphoblasts synchronized early in
the G1-phase of the cell cycle, addition of interleukin 2 (IL-2) stim
ulates transition into S-phase in conjunction with specific and reprod
ucible protein tyrosine phosphorylation events. Prominent among these
was de novo phosphorylation of the p56lck tyrosine kinase, which appea
red as a single polypeptide on SDS-polyacrylamide gel electrophoresis
prior to the addition of IL-2, but as a doublet of approximately 56 an
d 59 kDa 3 h after IL-2 addition. Although the lck polypeptide doublet
persisted into S-phase, the 59-kDa form was virtually undetectable in
T-lymphoblasts in log phase growth. In T-lymphoblasts metabolically l
abeled with P-32i, antiphosphotyrosine antisera identified a major 56-
kDa tyrosine-phosphorylated protein, whereas antisera to lck showed th
at phosphorylation of lck occurred on both the 56- and 59-kDa forms. P
hosphoamino acid analyses identified phosphoserine as the major phosph
oamino acid on both of the lck polypeptides, although small amounts of
phosphotyrosine were also detected on the 56-kDa form. Immune complex
kinase assays revealed that only the lower band of the lck doublet ex
hibited autocatalytic activity. Furthermore, the 59-kDa form neither d
isplayed autocatalytic activity, nor was it a substrate for phosphoryl
ation by the 56-kDa form. We conclude that entry into S-phase in activ
ated human T-lymphoblasts is associated with tyrosine phosphorylation
of a limited number of proteins and suggest that p56lck activity is re
gulated during G1/S.