Extended Data Figure 3: The integrity of the SOG complex is not regulated by amino acids or SESN2.
a, SZT2Flag/− cells were either untreated, deprived of amino acids for 60 min, deprived of amino acids for 60 min and stimulated with amino acids for 10 or 30 min, or treated with rapamycin (Rapa) for 60 min. Anti-Flag immunoprecipitates and total cell lysates were analysed by immunoblotting. HEK293T cells stably overexpressing Flag–RFP were used as a control. b, Anti-Flag immunoprecipitates prepared from SZT2Flag/− cells were washed three times with buffer containing sodium chloride, and analysed by immunoblotting. HEK293T cells stably overexpressing Flag–RFP were used as a control. c, The leucine-, arginine- or glutamine-binding activity of Flag–RFP, Flag–SZT2 and Flag–SESN2 was determined. Data represent mean ± s.d. (n = 3, two-tailed unpaired t-test). d, The leucine-binding activity of Flag–SESN2 expressed and purified from control or SZT2-deficient HEK293T cells. Data represent mean ± s.d. (n = 3, two-tailed unpaired t-test). e, Control or SZT2-deficient HEK293T cells stably overexpressing Flag–WDR24 were deprived of amino acids for 1 h and stimulated with amino acids for 10 min where indicated. Anti-Flag immunoprecipitates and total cell lysates were analysed by immunoblotting. f, HEK293T cells were transfected with the indicated constructs. Anti-HA immunoprecipitates and total cell lysates were analysed by immunoblotting. HA, haemagglutinin. Data (a, b, e, f) are representative of two independent experiments.
