ETH-1, THE NEUROSPORA-CRASSA LOCUS ENCODING S-ADENOSYLMETHIONINE SYNTHETASE - MOLECULAR-CLONING, SEQUENCE-ANALYSIS AND IN-VIVO OVEREXPRESSION

Intense biochemical and genetic research on the eth-1' mutant of Neuro spora crassa suggested that this locus might encode S-adenosylmethioni ne synthetase (S-Adomet synthetase). We have used protoplast transform ation and phenotypic rescue of a thermosensitive phenotype associated with the eth-1' mutation to clone the locus. Nucleotide sequence analy sis demonstrated that it encodes S-Adomet synthetase. Homology analyse s of prokaryotic, fungal and higher eukaryotic S-Adomet synthetase pol ypeptide sequences show a remarkable evolutionary conservation of the enzyme. N. crassa strains carrying S-Adomet synthetase coding sequence s fused to a strong heterologous promoter were constructed to assess t he phenotypic consequences of in vivo S-Adomet synthetase overexpressi on. Studies of growth rates and microscopic examination of vegetative development revealed that normal growth and morphogenesis take place i n N. crassa even at abnormally high levels of cellular S-Adomet. The d egree of cytosine methylation of a naturally methylated genomic region was dependent on the cellular levels of S-Adomet. We conclude that va riation in S-Adomet levels in N. crassa cells, which in addition to th e status of genomic DNA methylation could modify the flux of other S-A domet-dependent metabolic pathways, does not affect growth rate or mor phogenesis.