Translational Regulation of the Synthesis of Dihydrofolate Reductase

Ercikan, Emine; Banerjee, Debabrata; Waltham, Mark; Schnieders, Barbara; Scotto, Kathleen W.; Bertino, Joseph R.

doi:10.1007/978-1-4615-2960-6_109

Cited by 34 publications

(29 citation statements)

References 8 publications

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“…This binding is significant, particularly if the enzymes have different affinities for antifolate drugs such as methotrexate. The initial response of cells to methotrexate is to upregulate protein levels through disruption of the DHFR∶mRNA complex (29). Our finding that DHFRL1 can also prevent DHFR mRNA translation (and vice versa) indicates that this autoregulation mechanism and response to methotrexate needs to be reconsidered.…”

Section: Discussionmentioning

confidence: 90%

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

McEntee

Minguzzi

O’Brien

et al. 2011

Proc. Natl. Acad. Sci. U.S.A.

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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.

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Section: Discussionmentioning

confidence: 90%

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

McEntee

Minguzzi

O’Brien

et al. 2011

Proc. Natl. Acad. Sci. U.S.A.

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“…This has been attributed to a phenomenon in which the DHFR message binds to DHFR, thus regulating its availability for translation. In the presence of an antifolate, the message is released, translation is enhanced, and synthesis of protein is increased (Chu et al, 1993;Ercikan et al, 1993;Ercikan-Abali et al, 1997;Schmitz et al, 2001). Regulation of DHFR expression by this mechanism was absent in the malaria parasite and suggested as a factor in the utility of antifolates in the treatment of this disease (Zhang and Rathod, 2002).…”

Section: Alterations In Dhfrmentioning

confidence: 99%

Resistance to antifolates

2003

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“…Based on studies demonstrating an interaction between DHFR protein and its cognate mRNA, a model was proposed to account for increases in DHFR protein levels in response to MTX exposure, namely that the binding of human DHFR protein to its cognate mRNA results in decreased translation and that the addition of MTX disrupts this autoregulation (Chu et al, 1993;Ercikan et al, 1993). Using an in vitro translation assay, our laboratory (Ercikan et al, 1993) and Chu et al (1993) demonstrated that addition of exogenous DHFR protein in a rabbit reticulocyte system inhibited translation of DHFR mRNA. using both RNA gel-shift assays and UV cross-linking competition studies, the DHFR binding site was localized to a 100-base pair region in the coding region between nucleotides 380 and 480 (Ercikan-Abali et al, 1997).…”

mentioning

confidence: 99%

Species-Specific Differences in Translational Regulation of Dihydrofolate Reductase

Hsieh¹,

Skacel²,

Bansal³

et al. 2009

Mol Pharmacol

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We have observed that rodent cell lines (mouse, hamster) contain approximately 10 times the levels of dihydrofolate reductase as human cell lines, yet the sensitivity to methotrexate (ED 50 ), the folate antagonist that targets this enzyme, is similar. Our previous studies showed that dihydrofolate reductase protein levels increased after methotrexate exposure, and we proposed that this increase was due to the relief of feedback inhibition of translation as a consequence of methotrexate binding to dihydrofolate reductase. In the current report, we show that unlike what was observed in human cells, dihydrofolate reductase (DHFR) levels do not increase in hamster cells after methotrexate exposure. We provide evidence to show that although there are differences in the putative mRNA structure between hamster and human mRNA in the dihydrofolate reductase binding region previously identified, "hamsterization" of this region in human dihydrofolate reductase mRNA did not change the level of the enzyme or its induction by methotrexate. Further experiments showed that human dihydrofolate reductase is a promiscuous enzyme and that it is the difference between the hamster and human dihydrofolate reductase protein, rather than the DHFR mRNA, that determines the response to methotrexate exposure. We also present evidence to suggest that the translational up-regulation of dihydrofolate reductase by methotrexate in tumor cells is an adaptive mechanism that decreases sensitivity to this drug.

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Translational Regulation of the Synthesis of Dihydrofolate Reductase

Cited by 34 publications

References 8 publications

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

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

Resistance to antifolates

Species-Specific Differences in Translational Regulation of Dihydrofolate Reductase

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