Physical localization of yeast CYS3, a gene whose product resembles the rat γ‐cystathionase and Escherichia coli cystathionine γ‐synthase enzymes

Abstract

We have cloned, sequenced and physically mapped the CYS3 gene of Saccharomyces cerevisiae. This gene can complement the cys3-1 allele, and disruptions at this locus lead to cysteine auxotrophy. The predicted CYS3 product is closely related (46% identical) to the rat cystathionine gamma-lyase (Erickson et al., 1990), but differs in lacking cysteine residues. These results provide further evidence that the S288C strain of yeast resembles mammals in synthesizing cysteine solely via a trans-sulfuration pathway. The CYS3 product was found to have strong homology to three other enzymes involved in cysteine metabolism: the Escherichia coli metB and metC products and the S. cerevisiae MET25 gene product. The trans-sulfuration enzymes appear to form a diverged family and carry out related functions from bacteria to mammals.

Telomeric Silencing of an Open Reading Frame in Saccharomyces cerevisiae

“…Four measurements were carried out on each band and the average 6 the standard error is shown. CYS3 RNA showed virtually constant levels relative to template RNA added in all strains examined and was used as a loading control (Barton et al 1993). RNA copies per cell are calculated from the amount of the YAR073W/IMD1 RNA level measured in the sir3 strain by blot hybridization (one copy/cell).…”

“…This previously observed species (Barton et al 1997) does not appear to be affected by sir3 and is probably transcribed from the additional orthologous copies of this ORF found on chromosomes XII and XIII. A quantitative RT-PCR assay specific for YAR073W/IMD1 (Barton et al 1993) was used to show that sir3 mutant strains contained approximately sevenfold more YAR073W/ IMD1 RNA than the wild-type controls ( Figure 1C). The controls also show that this transcript could not be detected when chromosome I sequences were deleted.…”

“…Lanes: 1, DK425-14A; 2, DK421-1Asir3; 3, DK421-2D; 4, DK421-1A. YAR073W/IMD1 transcript abundance (RNA copies per cell) was determined by RT-PCR using simultaneously amplified CYS3 RNA as a loading control (Barton et al 1993). Nucleic acid isolation and manipulation were as described (Sambrook et al 1989).…”

Yeast cys3 and gsh1 mutant cells display overlapping but non-identical symptoms of oxidative stress with regard to subcellular protein localization and CDP-DAG metabolism

“…The serine residue is highly conserved in pyridoxal-phosphatedependent cystathionine c-lyases and -synthases from bacteria to man. It is located three amino acids N-terminal to the lysine residue to which the cofactor is attached (Barton et al 1993;Appel et al 1994). The mutant protein may therefore be inactive because of an inability to bind pyridoxal-phosphate.…”

“…A complementing plasmid (2l-CYS3, Fig. 2A) was identi®ed and found to contain a 2.7-kb fragment of chromosome I (nucleotide 129954± 132702) with a single complete gene (nucleotides 130,798 to 131,979), STR1/CYS3 (Cherest and SurdinKerjan 1992; Ono et al 1992; Barton et al 1993;Yamagata et al 1993). CYS3 codes for the pyridoxal phosphate-dependent cystathionine-c-lyase (EC 4.4.1.1.).…”