JAK3 FERM mutations cause increased phosphorylation of STAT5A in reconstituted IL-2 signaling in 293T cells. (A) Western blots show phospho-(P-)STAT5A (92 kDa), STAT5A, and JAK3. Lane 1 shows whole-cell lysate of the HuT-102 cell line. STAT5A phosphorylation requires JAK3 and IL-2 (100 U/mL). (B) Graph shows quantification of Western blots; P-STAT5A protein per unit of expressed JAK3 protein. Mutant JAK3s were compared with WT JAK3 in 4 independent cotransfection experiments and showed consistently higher P-STAT5A levels (mean ± SEM; top panel). P values were generated from the Student t test (2-tailed). The bottom panel shows P-STAT5A protein levels in cotransfection experiments with various other JAK3 mutants normalized to endogenous levels in HuT-102. The K855A mutant causes loss of kinase activity in JAK3. The Y100C mutation was found in a SCID patient and P132T is a SNP.