Evidence for small ubiquitin-like modifier-dependent nuclear import of the thymidylate biosynthesis pathway.
Perturbations in folate-mediated one-carbon metabolism increase rates of uracil misincorporation into DNA during replication, impair cellular methylation reactions, and increase risk for neural tube defects and cancer. One-carbon metabolism is compromised by folate deficiency and common genetic polymorphisms. In this study, the mechanism for the preferential partitioning of cytoplasmic serine hydroxymethyltransferase (cSHMT)-derived methylenetetrahydrofolate to de novo thymidylate biosynthesis was investigated. The cSHMT enzyme was shown to interact with UBC9 and was a substrate for UBC9-catalyzed small ubiquitin-like modifier (SUMO) modification in vitro. SUMOylated cSHMT was detected in extracts from S phase MCF-7 cells, and cSHMT was shown to localize to the nucleus and nuclear periphery during the S and G(2)/M phases of the cell cycle. A common single nucleotide polymorphism (L474F-cSHMT) impaired the UBC9-cSHMT interaction and inhibited cSHMT SUMOylation in vitro. The three folate-dependent enzymes that constitute the de novo thymidylate biosynthesis pathway, cSHMT, thymidylate synthase, and dihydrofolate reductase, all contain SUMO modification consensus sequences. Compartmentation of the folate-dependent de novo thymidylate biosynthesis pathway in the nucleus accounts for the preferential partitioning of cSHMT-derived folate-activated one-carbon units into thymidylate biosynthesis; the efficiency of nuclear folate metabolism is likely to be modified by the cSHMT L474F polymorphism.
Woeller
Collynn F
CF
Division of Nutritional Sciences, Cornell University, Ithaca, New York 14853, USA.
Anderson
Donald D
DD
Szebenyi
Doletha M E
DM
Stover
Patrick J
PJ
D021581:Active Transport, Cell Nucleus / Q000502:physiology*; D000595:Amino Acid Sequence; D000818:Animals; D002244:Carbon / Q000378:metabolism; D002453:Cell Cycle / Q000502:physiology; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism*; D005492:Folic Acid / Q000378:metabolism; D012696:Glycine Hydroxymethyltransferase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011110:Polymorphism, Genetic; D011487:Protein Conformation; D016415:Sequence Alignment; D025841:Small Ubiquitin-Related Modifier Proteins / Q000235:genetics / Q000378:metabolism; D013938:Thymidine Monophosphate / Q000096:biosynthesis*; D020798:Two-Hybrid System Techniques; D044763:Ubiquitin-Conjugating Enzymes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D020931:ran GTP-Binding Protein / Q000235:genetics / Q000378:metabolism; D000098843:Ubiquitin-Conjugating Enzyme UBC9
D025841:Small Ubiquitin-Related Modifier Proteins; D013938:Thymidine Monophosphate; D002244:Carbon; D005492:Folic Acid; D012696:Glycine Hydroxymethyltransferase; C520541:SHMT protein, human; D044763:Ubiquitin-Conjugating Enzymes; D020931:ran GTP-Binding Protein; D000098843:Ubiquitin-Conjugating Enzyme UBC9