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

Some insertion mutations in Saccharomyces cerevisiae activate the expression of adjacent structural genes. The CYC7-H2 mutation is a Ty1 insertion 5' to the iso-2-cytochrome c coding region of CYC7. The Ty1 insertion causes a 20-fold increase in CYC7 expression in a and alpha haploid cell types of S. cerevisiae. This activation is repressed in the a/alpha diploid cell type. Previous computer analysis of the CYC7-H2 Ty1 activator region identified two related sequences with homology both to mammalian enhancers and to a yeast a/alpha control site. A 112-base-pair (bp) DNA fragment encompassing one of these blocks of homology functioned as one component of the Ty1 activator. A 28-bp synthetic oligonucleotide with the wild-type homology block sequence was also functional. A single base pair mutation within the enhancer core of the synthetic 28-bp regulatory element reduced its activation ability to near background amounts. In addition, the 112-bp Ty1 fragment by itself functioned as a target for repression of adjacent gene expression in a/alpha diploid cells.     

A Ty1 cell-type-specific regulatory sequence is a recognition element for a constitutive binding factor.

Ty transposable-element insertion mutations of Saccharomyces cerevisiae can cause cell-type-dependent activation of adjacent-gene expression. Several cis-acting regulatory regions within Ty1 are responsible for the effect of Ty1 on adjacent-gene expression. One of these is the block II sequence that was defined by its homology to mammalian enhancers and to the yeast a1-alpha 2 control site. Tandem copies of a 57-base-pair region encompassing block II caused an additive increase in expression of the CYC7 reporter gene in the absence of other Ty1 sequences. The activation of gene expression by the multiple repeats was abolished in a/alpha diploid cells. A specific complex between a constitutive factor in whole-cell extracts and the DNA regulatory element was observed. The protein-binding site for the constitutive factor coincided with the block II element. Base-pair substitutions within the binding site abolished the ability of the block II element to function as a component of the Ty1 activator and to form the factor-DNA complex. The correlation between complex formation and reporter gene expression indicates that factor binding to the cis-acting element is essential for this element to function as a component of the Ty1 activator.     

A yeast-Escherichia coli shuttle vector containing the M13 origin of replication

A yeast-Escherichia coli shuttle vector containing the M13 origin of replication has been constructed. This vector allows selection and replication in both Saccharomyces cerevisiae and E. coli, as well as single-stranded packaging from E. coli upon infection with a helper phage. The presence of a polylinker with various unique restriction sites facilitates the cloning of desired genes.     

Phylogeny of Greya (Lepidoptera: Prodoxidae), based on nucleotide sequence variation in mitochondrial cytochrome oxidase I and II: congruence with morphological data

The phylogeny of Greya Busck (Lepidoptera: Prodoxidae) was inferred from nucleotide sequence variation across a 765-bp region in the cytochrome oxidase I and II genes of the mitochondrial genome. Most parsimonious relationships of 25 haplotypes from 16 Greya species and two outgroup genera (Tetragma and Prodoxus) showed substantial congruence with the species relationships indicated by morphological variation. Differences between mitochondrial and morphological trees were found primarily in the positions of two species, G. variabilis and G. pectinifera, and in the branching order of the three major species groups in the genus. Conflicts between the data sets were examined by comparing levels of homoplasy in characters supporting alternative hypotheses. The phylogeny of Greya species suggests that host-plant association at the family level and larval feeding mode are conservative characters. Transition/transversion ratios estimated by reconstruction of nucleotide substitutions on the phylogeny had a range of 2.0-9.3, when different subsets of the phylogeny were used. The decline of this ratio with the increase in maximum sequence divergence among taxa indicates that transitions are masked by transversions along deeper internodes or long branches of the phylogeny. Among transitions, substitutions of A-->G and T-->C outnumbered their reciprocal substitutions by 2-6 times, presumably because of the approximately 4:1 (77%) A+T-bias in nucleotide base composition. Of all transversions, 73%-80% were A<-->T substitutions, 85% of which occurred at third positions of codons; these estimates did not decrease with an increase in maximum sequence divergence of taxa included in the analysis. The high frequency of A<-->T substitutions is either a reflection or an explanation of the 92% A+T bias at third codon positions.