The former annotated human pseudogene dihydrofolate reductase-like 1 ( DHFRL1 ) is expressed and functional

Abstract

Human dihydrofolate reductase (DHFR) was previously thought to be the only enzyme capable of the reduction of dihydrofolate to tetrahydrofolate; an essential reaction necessary to ensure a continuous supply of biologically active folate. DHFR has been studied extensively from a number of perspectives because of its role in health and disease. Although the presence of a number of intronless DHFR pseudogenes has been known since the 1980s, it was assumed that none of these were expressed or functional. We show that humans do have a second dihydrofolate reductase enzyme encoded by the former pseudogene DHFRP4, located on chromosome 3. We demonstrate that the DHFRP4, or dihydrofolate reductase-like 1 (DHFRL1), gene is expressed and shares some commonalities with DHFR. Recombinant DHFRL1 can complement a DHFR-negative phenotype in bacterial and mammalian cells but has a lower specific activity than DHFR. The K m for NADPH is similar for both enzymes but DHFRL1 has a higher K m for dihydrofolate when compared to DHFR. The need for a second reductase with lowered affinity for its substrate may fulfill a specific cellular requirement. The localization of DHFRL1 to the mitochondria, as demonstrated by confocal microscopy, indicates that mitochondrial dihydrofolate reductase activity may be optimal with a lowered affinity for dihydrofolate. We also found that DHFRL1 is capable of the same translational autoregulation as DHFR by binding to its own mRNA; with each enzyme also capable of replacing the other. The identification of DHFRL1 will have implications for previous research involving DHFR.

The 1,2-Diaminocyclohexane Carrier Ligand in Oxaliplatin Induces p53-Dependent Transcriptional Repression of Factors Involved in Thymidylate Biosynthesis

“…5A and B). DHFR catalyzes the reduction of dihydrofolate to tetrahydrofolate, which is an essential cofactor for thymidylate biosynthesis by TS (35), and TK1 is an essential kinase of the salvage pathway for thymidylate biosynthesis (33). Consistent with this, following oxaliplatin treatment we observed a significant decrease in the level of dTTP in the dNTP pool (Fig.…”

“…In addition, the salvage thymidylate biosynthesis pathway, which is initiated by TK1-mediated phosphorylation of thymidine, is another route for dTTP biosynthesis (33). We focused on three genes encoding enzymes of the thymidylate biosynthesis pathway, DHFR, TK1, and TS, whose promoters have architectures similar to those of genes involved in the G 1 -S transition, including DUT (34), and whose transcript levels during cell-cycle progression are regulated by Sp1 and E2F1 (35)(36)(37). The expression of DHFR (Fig.…”

A-to-I RNA Editing Up-regulates Human Dihydrofolate Reductase in Breast Cancer

“…21), indicating that the contribution of ADAR1 to resistance is not different between MCF-7 and A549 cells. It has been reported that dihydrofolate reductase like 1, whose gene is known as pseudogene DHFRP4, is expressed and has same function as DHFR in folate metabolism (McEntee et al, 2011). It is possible that this enzyme may complement the decreased expression of DHFR by ADAR1 knockdown, leading to residual resistance to methotrexate in these cell lines.…”

The Dihydrofolate Reductase 19 bp Polymorphism Is Not Associated with Biomarkers of Folate Status in Healthy Young Adults, Irrespective of Folic Acid Intake

“…Recently, a second DHFR-like enzyme, DHFRL1, has been identified (28,29), but it has much lower enzyme activity than DHFR (28) and is unlikely to account for the reported differences in enzyme activity between individuals (25). This points toward common polymorphisms that control the regulation and expression of the DHFR gene.…”