Fig. S4.

(Related to Fig. 5) Depletion of p97 by shRNA blocks glutamine-induced GS disassembly. HEK293 cells stably expressing doxycycline (Dox)-inducible shRNA targeting p97 were mock-treated or were induced with doxycycline (1 µg/mL) for 48 h, starved of glutamine for 24 h, and then pretreated with MG132 (10 μM) for 30 min, followed by the addition (or not) of 4 mM glutamine for 6 h. Cell lysates were fractionated on a Superdex 200 gel filtration column. Individual fractions were concentrated by TCA precipitation and analyzed by SDS/PAGE and immunoblotting with the indicated antibodies.