Cloning and characterization of human ubiquitin-processing protease-43 from terminally differentiated human melanoma cells using a rapid subtraction hybridization protocol RaSH

Defects in growth control and differentiation occur frequently in human can cers. In the case of human melanoma cells, treatment with a combination of fibroblast interferon (IFN-beta) and the protein kinase C activator mezerei n (MEZ) results in an irreversible loss of proliferative potential and tumo rigenic properties with a concomitant induction of terminal differentiation . These changes in cellular properties are associated with an induction and suppression in specific subsets of genes that occur in a temporal manner. To identify the complete repertoire of gene changes occurring during melano ma reversion to a more differentiated state a number of molecular approache s are being used. These include, subtraction hybridization using temporally spaced cDNA libraries, random cDNA isolation and evaluation by reverse Nor thern blotting and high throughput microarray analysis of subtracted cDNA c lones. In the present study we have used a novel approach, rapid subtractio n hybridization (RaSH), to identify and clone an additional gene of potenti al relevance to cancer growth control and terminal cell differentiation. Ra SH has identified a human ubiquitin-processing protease gene, HuUBP43, that is differentially expressed in melanoma cells as a function of treatment w ith IFN-beta or IFN-beta + MEZ. HuUBP43 is a type I interferon inducible ge ne that is upregulated in a diverse panel of normal and tumor cells when tr eated with IFN-beta via the JAK/STAT kinase pathway. This gene may contribu te to the phenotypic changes induced by IFN-beta during growth arrest and d ifferentiation in human melanoma cells and other cell types as well as the antiviral and growth inhibitory effects of interferon. (C) 2001 Elsevier Sc ience B.V. All rights reserved.