Ty1 sequence with enhancer and mating-type-dependent regulatory activities.

Area of Science:

• Molecular Biology
• Genetics
• Yeast Biology

Background:

• Insertion mutations can alter gene expression in Saccharomyces cerevisiae.
• The CYC7-H2 mutation involves a Ty1 insertion upstream of the iso-2-cytochrome c gene (CYC7).
• This mutation leads to increased CYC7 expression in haploid yeast cells, with repression in diploid cells.

Purpose of the Study:

• To investigate the regulatory elements responsible for Ty1 insertion-mediated gene activation in yeast.
• To characterize the DNA sequences involved in the differential regulation of CYC7 expression between haploid and diploid yeast cells.

Main Methods:

• Computer analysis to identify homologous sequences in the Ty1 activator region.
• Functional analysis using DNA fragments and synthetic oligonucleotides to test regulatory activity.
• Site-directed mutagenesis to pinpoint critical elements within the regulatory sequence.

Main Results:

• A 112-base-pair (bp) DNA fragment from the Ty1 insertion acts as a gene activator.
• A shorter, 28-bp synthetic oligonucleotide containing a homologous sequence also functions as an activator.
• A single base pair mutation in this element significantly reduces its activation capacity.
• The 112-bp fragment also mediates repression of gene expression in diploid yeast cells.

Conclusions:

• A specific DNA sequence within the Ty1 insertion functions as a potent enhancer of adjacent gene expression in haploid yeast.
• This regulatory element is subject to repression in diploid yeast cells, indicating cell-type-specific regulation.
• The identified sequence is a key component of the CYC7-H2 activator and demonstrates enhancer-like properties.