Identification And Analysis Of Novel Alternative Splice Variant Of DHRS4LI Gene In HepG2 Cell Line

All-trans retinoic acid (atRA)is a hormone-like and physiological active substance in mammals, including humans. In vivo, either too high or too low concentration of the atRA may lead to various disease and cancer. atRA is derived from vitamin A, which also known as retinol, through a two-step oxidative metabolism, and retinol as intermediate metabolite between the two metabolisms, has a narrow physiological dose range in vivo, both too high or too low will also exert negative effects on organism. Therefore, the metabolism of both atRA and retinol are subject to strict and subtle regulation.In recent years, our group has been committed to the study on an short-chain dehydrogenase /reductas(eSDR)which is called NADP(H)-dependent retinol dehydrogenase/ reductase (NRDR). NRDR identified from rabbit liver cytosol firstly is a rate-limiting enzyme during metabolism of atRA. It has high retinol oxidation and retinal reduction activity, and is widely distributed in various mammalian tissues.Compared with other members of SDR superfamily, NRDR exhibits higher aldehyde reductase activity in retinoic acid (RA) metabolism. Therefore , NRDR may paly an important role in keeping the metabolism balance between atRA and retinal in vivo.We focused on the cDNA that deposited in Genebank (accession number:AB045133) and annotated as the coding gene of NRDR, which is called DHRS4. There are three copies for human DHRS4 gene cluster, including DHRS4, DHRS4L2 and DHRS4L1. The identity between the former two copies is 97.5%, suggesting that they are segmental duplication (90%-98%;≥1 kb). In the study on gene expression and regulation of DHRS4, we found there are sophisticated ways of transcription and alternative splicing, and obtained many alternatively spliced variants. The different expression of these spliced variants in different tissues may have important clinical significance. In this study, we identified an alternatively spliced variant from Hep-G2, detected its expression in different tissues and cells, as well as its clinical significance.In this study, we cloned the full-length cDNA of this alternatively spliced variant in Hep-G2 cell line by Rapid amplification of cDNA ends(RACE), it's about 1117bp. Sequence alignment analysis shows that the cDNA of this alternatively spliced variant named DHRS4L1A1 has highly homology with DHRS4L1. Human genome Blast analysis shows that the structure of DHRS4L1A1 is e1-e2-insertion-e8-e9-e10 with exon 3 to exon 7 deletion. We discovered an open reading frame of 699bp, and it may could be translated a protein contained 232 amino acids. Finally, we found DHRS4L1A1 could express in many different cell lines , and its expression in tumor cells ,specifically in ovarian cancer cell line SKOV3, is higher than normal cells, suggesting the enzyme might has close relationship with metabolic pathways of reproductive system.