Figure 4

P62 and c-Myc mediate the metabolic effect of the proteasome inhibitors in UOK262 cells. (A) UOK262 cells were transiently silenced for p62 or c-Myc using pooled siRNA. Viability was assessed by luminescence 72 hours post-transfection; (B) Relative mRNA expression of MYC, SQSTM1 (p62) and LDHA were measured by RT- PCR 48 hours after transfection with siRNA targeting p62 (SQSTM1) or MYC; (C) MYC expression levels were evaluated after 24 hours treatment with bortezomib, marizomib and carfilzomib (30 nM). Transient silencing of MYC was used as a control. Transfection of MYC expression plasmid in UOK262 cells was shown to result in significant overexpression of MYC; (D) MYC overexpression partially rescues the viability of UOK262 24 hours after bortezomib, marizomib and carfilzomib treatments (30 nM); (E) Transient silencing of p62 and c-Myc significantly decreased ECAR in UOK262 cells. Measurements were performed 48 hours after transfection; (F) Overexpression of MYC negates the effect of proteasome inhibitors on ECAR; (G) Overexpression of MYC abrogates the effect of marizomib on the lactate secretion and glucose consumption of UOK262 cells. *p < 0.05; Bort: bortezomib; Mar: marizomib; Carf: carfilzomib; Control: DMSO control; ECAR: extra-cellular acidification rate.