80 -

68 -48 -

30 -25 -

kDa 118 -

IP: IgG VHL

HA-Ub + + +Hypoxia + - +MG132 + + +

68 -

48 -

30 -

25 -

IB: a-HA

blot: VHL

Ub-

VH

L17 - p19 VHL

p25 VHL

blot: HA

Supplemental Figure 1

786-O G7F cells were transfected with an HA-Ubiquitin expression vector. Twenty four hours later, transfected cells were treated with MG-132 10 M and with hypoxia or normoxia for 8 hours. Cells were lysed with NP40 buffer and immunoprecipitation was performed with anti-VHL (Ig32) antibody and western blot was further performed with anti-HA (Y11) (top panel) or anti-VHL (Ig32) (bottom panel), respectively.