Characterization of the Formate (for) Locus, Which Encodes the Cytosolic Serine Hydroxymethyltransferase of Neurospora crassa

Abstract

Serine hydroxymethyltransferase (SHMT) occupies a central position in one-carbon (C₁) metabolism, catalyzing the reaction of serine and tetrahydrofolate to yield glycine and 5,10-methylenetetrahydrofolate. Methylenetetrahydrofolate serves as a donor of C₁ units for the synthesis of numerous compounds, including purines, thymidylate, lipids, and methionine. We provide evidence that the formate (for) locus of Neurospora crassa encodes cytosolic SHMT. The for⁺ gene was localized to a 2.8-kb BglII fragment by complementation (restoration to formate-independent growth) of a strain carrying a recessive for allele, which confers a growth requirement for formate. The for⁺ gene encodes a polypeptide of 479 amino acids which shows significant similarity to amino acid sequences of SHMT from bacterial and mammalian sources (47 and 60% amino acid identity, respectively). The for⁺ mRNA has several different start and stop sites. The abundance of for⁺ mRNA increased in response to amino acid imbalance induced by glycine supplementation, suggesting regulation by the N. crassa cross-pathway control system, which is analogous to general amino acid control in Saccharomyces cerevisiae. This was confirmed by documenting that for⁺ expression increased in response to histidine limitation (induced by 3-amino-l,2,4-triazole) and that this response was dependent on the presence of a functional cross-pathway control-1 (cpc-1) gene, which encodes CPC1, a positively acting transcription factor. There are at least five potential CPC1 binding sites upstream of the for⁺ transcriptional start, as well as one that exactly matches the consensus CPC1 binding site in the first intron of the for⁺ gene.