FIGURE 6.
The USP10-knockdown A549 cells display a reduced MMR activity compared with the control cells. DNA substrate (left) used in this study contains a G-T mismatch (red) placed into two overlapping restriction endonucleases and a strand break 172-bp 3′ to the mismatch. Restriction enzymes used for scoring the repair are shown. MMR activity of A549 cells with or without shUSP10 was determined by incubating the circular DNA heteroduplex with 75 μg of whole cell extracts. Repair products were identified by Southern blot analysis using a 32P-labeled oligonucleotide probe (red bar) complementary to the nicked strand at the indicated location. MMR activities in HeLa extracts and heat-inactivated HeLa extracts (HI-HeLa) were used as a positive control and a negative control, respectively. The repaired bands were quantified by densitometry.