摘要：

Methylenetetrahydrofolate reductase (MTHFR), a key enzyme in the folate cycle, catalyzes the reduction of 5,10-methylenetetrahydrofolate to 5-methyltetrahydrofolate, a co-substrate for homocysteine remethylation to methionine. Methionine serves as the precursor of the active methyl donor S-adenosylmethionine, which provides methyl groups for many biological methylations. It has been reported that MTHFR is highly phosphorylated under unperturbed conditions and T34 is the priming phosphorylation site. In this report, we generated a phospho-specific antibody that recognized T34-phosphorylated form of MTHFR and revealed that MTHFR was phosphorylated at T34 in vivo and this phosphorylation peaked during mitosis. We further demonstrated that the cyclin-dependent kinase 1 (CDK1)/Cyclin B1 complex is the kinase that mediates MTHFR phosphorylation at T34 and the MTHFR immunocomplex purified from mitotic cells exhibited lower enzymatic activity. Inhibition of MTHFR expression resulted in a decrease of H3K9me3 levels, and an increase of transcription of the centromeric heterochromatin markers. Taken together, our results demonstrated that CDK1/Cyclin B1 phosphorylates MTHFR on T34 and MTHFR plays a role in the heterochromatin maintenance at the centromeric region.Article Outline1. Introduction2. Materials and methods2.1. Cell lines, plasmids, siRNA oligos and antibodies2.2. Cell synchronization2.3. CDK1/Cyclin B1 kinase assay2.4. NADPH-menadione oxidoductase assay2.5. Real-time PCR assay3. Results3.1. T34 is the major phosphorylation site of MTHFR3.2. MTHFR T34 is phosphorylated by the CDK1/Cyclin B1 kinase complex3.3. The enzymatic activity of MTHFR decreases during mitosis3.4. MTHFR plays an important role in maintaining the heterochromatin structure at the centromeric region4. DiscussionAcknowledgmentsReferences

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