[Show abstract][Hide abstract] ABSTRACT: The gene for the INK4 family Cdk inhibitor p15 (INK4B) is frequently deleted or inactivated in multiple types of human cancers, indicating that p15 is a tumor suppressor. p15RS is a ubiquitously expressed nuclear protein that is positively regulated by p15 and, in turn, inhibits the expression of cyclin D and cyclin E. To determine whether p15RS has malignancy inhibitory functions in addition to its inhibitory effects on cell cycle entry, we ectopically expressed p15RS in metastatic melanoma A375 cells, in which p15 gene is deleted and p15RS expression is dramatically downregulated, and examined the effect on various malignant phenotypes. Here, we report that while the p15RS expression had little effect on cell growth in monolayer cultures, it dramatically inhibited anchorage-independent cell growth in soft agar, a hallmark for malignancy. p15RS expression also inhibited cell migration and invasion, which are key determinants of metastasis. At molecular levels, p15RS expression specifically downregulates the expression of cathepsin B and MMP-9 at RNA levels, which are known to promote cell invasion through degrading extracellular matrix proteins. These results indicate that p15RS has malignancy inhibitory functions independent of cell cycle inhibition and provide novel insights on the role of p15 in tumor inhibition.