We have previously identified 67 exons on a yeast artificial chromosome contig spanning 1.5 Mb around the multidrug resistance 1 gene region of human chromosome 7q21.1. In this study, we identified three novel cytoplasmic variants (MDC2-γ γ γ γ, MDC2-δ δ δ δ, and MDC2-ε ε ε ε) of the human metalloprotease-like disintegrin-like cysteine-rich protein 2 (MDC2) among these exons by screening a human brain cDNA library and also by using a reverse transcription polymerase chain reaction. Genomic sequence analysis strongly supported the idea that the variations in the cytoplasmic domain were generated by alternative splicing. The expression of MDC2 variant forms in human brain tissue and gliomas was examined by reverse transcription polymerase chain reaction and RNase protection assay. MDC2-ε ε ε ε was expressed only in the cortical and hippocampal regions in human brain, but not in gliomas. In contrast, MDC2-γ γ γ γ was a major form expressed in human gliomas. Specific expression of these cytoplasmic variants of MDC2 in human brain and its malignancies is discussed.
Key words: MDC -ADAM -Alternative splicing -MDR regionThe mammalian ADAM (a disintegrin and metalloprotease) family is a recently identified gene family encoding membrane proteins, and has an extensive sequence similarity to snake venom disintegrin and metalloprotease.
1)Snake venom disintegrins are a family of anticoagulant peptides with a high cysteine content. 1) Typical ADAMs are cell-surface proteins that have multiple domains; pro, metalloprotease-like, disintegrin-like, cysteine-rich, epidermal growth factor-like, transmembrane, and cytoplasmic domains. The ADAM family is expected to be involved in cell-cell interactions. For example, fertilin, the first ADAM described, has been implicated in integrin-mediated sperm-egg binding, 2, 3) and meltrin is required for myotube formation.4) In contrast, some members of this ADAM protein family have metalloprotease-like domains that are catalytically active and degrade specific substrates. An example of this type of ADAM is human tumor necrosis factor-α converting enzyme, which releases soluble tumor necrosis factor-α by proteolysis.5, 6) Because cellcell interaction and protease activity have important roles in metastasis and invasion of cancer cells, the ADAM family could be associated with malignant phenotype in cancer cells.We previously assembled a contig of 21 non-chimeric yeast artificial chromosomes (YAC) across 1.5 Mb of the multidrug resistance gene (PGY1 and PGY3) region on human chromosome 7q21.1. 7) Exon-trapping directly on the YACs resulted in an exon collection that includes 21 exons identical to the cDNA sequences of PGY1, PGY3, and sorcin, and 43 exons homologous or similar to human cDNA sequences. 8) In these 43 exons, nine exons were homologous to a cDNA coding a human metalloprotease/ disintegrin-like cysteine-rich protein (MDC), which is a member of the ADAM family.9, 10) These exons specifically detected a prominent 10 kb mRNA in the brain among various human tissues examined. 8) Sagan...