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. 2011 Jul 5;286(34):29951–29963. doi: 10.1074/jbc.M111.244715

FIGURE 5.

FIGURE 5.

Requirement for MAGE-11 in human AR transactivation. A, LAPC-4 cells were untreated (lane 1) or transduced with lentivirus MAGE-11 shRNA-169, -827, -947, and -964 (lanes 2–5); human AR shRNA-5 (lane 6); empty vector (NS1; lane 7); or 18-bp-spacer nonspecific shRNA (NS2; lane 8). Cell extracts (100 μg of protein/lane) separated on transblots were probed using AR32 and AR52 antibodies combined, MAGE-Ab-94–108 and FLAG-MAGE-11 antibodies combined, and β-actin antibody. B, quantitative real time RT-PCR analysis of MAGE-11 mRNA from LAPC-4 cells was performed as described under “Experimental Procedures” using cells not treated with lentivirus (−) or transduced with lentivirus MAGE-11 shRNA-169, -827, -947, and -964 (M); human AR shRNA-5 (AR); and 18-bp nonspecific shRNA-2 (NS). Cells were incubated for 24 h with 10 nm DHT. MAGE-11 mRNA levels were normalized to peptidylprolyl isomerase A (PPIA) mRNA. C, quantitative real time RT-PCR analysis of PSA mRNA was performed in LAPC-4 cells as in B except that cells were incubated for 24 h with and without 10 nm DHT. Results indicate PSA mRNA -fold induction normalized to peptidylprolyl isomerase A assayed in triplicate ±S.E. relative to activity without DHT.