Posttranscriptional regulation of the expression of MET2 gene of Saccharomyces cerevisiae.
The first step of the specific pathway for methionine biosynthesis in the yeast Saccharomyces cerevisiae is catalyzed by the enzyme L-homoserine-O-acetyltransferase (HSTase) (EC 2.3.1.31), encoded by the MET2 gene. In order to ascertain whether there is a posttranscriptional control on the MET2 gene expression, as suggested by previous results on the expression of the cloned gene, systems for high inducible expression of MET2 gene were constructed. In these constructs the MET2 gene was cloned in yeast expression vectors under the control of an inducible yeast GAL promoter element so that the MET2 was transcribed at very high levels under induced conditions. Measurements of the specific mRNA levels showed a strong stimulation of MET2 gene transcription in yeast transformants grown on galactose as carbon source, corresponding to 50-100-fold the repressed conditions, while only a 2-fold increase of the enzymatic activity was observed. In addition, no evidence of a strong induced polypeptide of appropriate size on two dimensional gel electrophoresis was obtained. To understand the functional role of the non-coding 5' region of MET2 mRNA, we performed either a partial and a complete deletion of the 5' leader sequence, but even with these constructs an elevated mRNA level was not associated to a marked increase of the HSTase activity. These data support the idea of a posttranscriptional regulation of MET2 gene expression and show that the untranslated region of the specific mRNA is not involved in this regulatory mechanism.
Forlani
N
N
Dipartimento di Fisiologia e Biochimica Generali, Sezione di Biochimica Comparata, Milano, Italy.
Martegani
E
E
Alberghina
L
L
D000123:Acetyltransferases / Q000235:genetics*; D001483:Base Sequence; D015152:Blotting, Northern; D002872:Chromosome Deletion; D015180:Electrophoresis, Gel, Two-Dimensional; D005656:Fungal Proteins / Q000235:genetics; D015966:Gene Expression Regulation, Fungal; D008715:Methionine / Q000096:biosynthesis*; D008969:Molecular Sequence Data; D010957:Plasmids; D014176:Protein Biosynthesis; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D014158:Transcription, Genetic
D005656:Fungal Proteins; D008715:Methionine; D000123:Acetyltransferases; C021369:homoserine O-acetyltransferase