An alternatively spliced form of NQO1 (DT-diaphorase) messenger RNA lacking the putative quinone substrate binding site is present in human normal and tumor tissues.
DT-diaphorase is a ubiquitously expressed flavoenzyme responsible for the two-electron reduction of a number of quinone and other anticancer drugs. The majority of DT-diaphorase enzyme activity in human tissues is the product of the NQO1 gene. We have now identified a novel alternatively spliced form of human NQO1 mRNA lacking exon 4 at levels equal to or exceeding those of wild-type NQO1 mRNA. Exon 4 codes for the putative quinone substrate binding site of DT-diaphorase derived from NQO1 and the recombinant protein from alternatively spliced NQO1 mRNA lacking exon 4 has minimal enzyme activity with quinoid and other known substrates of DT-diaphorase. The physiological substrate of DT-diaphorase is unknown, and it is possible that the protein derived from the alternatively spliced NQO1 mRNA could have enzyme activity with an appropriate substrate. We found full-length DT-diaphorase protein but could not detect expression of an appropriately smaller form of DT-diaphorase in human tissues using polyclonal antibody to DT-diaphorase, suggesting that alternatively spliced NQO1 mRNA lacking exon 4 may not be translated or that the protein product is rapidly degraded. Alternative splicing of NQO1 RNA could provide an important mechanism for regulating NQO1 gene expression.
Gasdaska
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PY
Arizona Cancer Center, University of Arizona, Tucson 85724, USA.
Fisher
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Powis
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D000230:Adenocarcinoma / Q000201:enzymology*; D017398:Alternative Splicing; D001483:Base Sequence; D001665:Binding Sites; D015152:Blotting, Northern; D002460:Cell Line; D003106:Colon / Q000201:enzymology*; D003110:Colonic Neoplasms / Q000201:enzymology*; D017931:DNA Primers; D018076:DNA, Complementary; D006801:Humans; D008168:Lung / Q000201:enzymology*; D008175:Lung Neoplasms / Q000201:enzymology*; D008969:Molecular Sequence Data; D016660:NAD(P)H Dehydrogenase (Quinone) / Q000096:biosynthesis* / Q000378:metabolism; D016133:Polymerase Chain Reaction; D011809:Quinones / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism*; D012016:Reference Values; D013379:Substrate Specificity; D014407:Tumor Cells, Cultured
D017931:DNA Primers; D018076:DNA, Complementary; D011809:Quinones; D012333:RNA, Messenger; D016660:NAD(P)H Dehydrogenase (Quinone)