Revertants of a transcription termination mutant of yeast contain diverse genetic alterations.

Revertants of the cyc1-512 transcription termination mutant of the yeast Saccharomyces cerevisiae were isolated and subjected to a detailed genetic analysis. The cyc1-512 mutation previously was shown to be a 38-base pair deletion that causes only 10% of the normal steady-state levels of CYC1 mRNA and of the CYC1 gene product, iso-1-cytochrome c. Forty-one cyc1-512 revertants were classified by their content of iso-1-cytochrome c and by their genetic properties in meiotic crosses. Many of the revertants contain local genetic changes that either partially or completely restore the level of iso-1-cytochrome c. One revertant was shown to contain an unlinked extragenic suppressor, designated sut1, that causes partial suppression of the transcription termination defect. Four revertants of cyc1-512 contain chromosomal rearrangements with breakpoints that are tightly linked to the CYC1 locus; these include one duplication, one possible inversion, and two reciprocal translocations. Detailed genetic mapping demonstrated that one of the reciprocal translocations is between the right arms of chromosomes X and XII, with a breakpoint mapping 3' to the CYC1 locus. These results indicate that the defect in transcription termination in cyc1-512 can be restored in a variety of ways, including the translocation of different chromosomal regions to the 3' end of the CYC1 locus, local changes presumably at or near the original defect, and by mutation at another locus distinct from CYC1.

A physical map of the bioAB region in the lambda bio transducing phage.

The center of the pBopA promoter-operator region for the bioABFCD operon is located 1.71 kb clockwise from the att lambda site on the Escherichia coli genome, as determined by the position of the p131 (IS1) insertion. The order of several bio endpoints to the right of p131 is lambda bio267, 122, 169, 74, 1, and 69. The endpoints of the two bio deletions, delta 61 in bioA and delta 3h in bioB, were also determined.

Leftward transcription in the Escherichia coli bio operon does not require products of the rightward transcript.

The amount of leftward transcription in the Escherichia coli bio operon, as measured by hybridization and by beta-galactosidase assays in lac-bio fusion strains, was determined in bacteria lysogenic for lambda bio phage carrying different amounts of DNA corresponding to rightward message, and in bacteria with polar or nonpolar bioB mutations. The positions of the bioB endpoints in relation to the pB promoter were determined by electron microscopy of heteroduplexes. Normal rates of leftward transcription were found in all cases, except that the shortest lambda bio (lambda bio showed a 2- to 3-fold increase in leftward transcription, which was not abolished by the presence of a wild-type bio operon in trans. These results indicate that no product of the rightward transcript is needed to turn on leftward transcription.

Requirement for protein synthesis for survival of unmodified bacteriophage T1 in a restricting host.

At high multiplication of infection, a substantial fraction of restricting cells (P1 lysogens) could be productively infected by unmodified coliphage T1 (T1.0) provided that protein synthesis was uninhibited during the first 5 min of infection. Successful infection under restricting conditions was accompanied by more genetic recombination than was seen under nonrestricting host, the recombination frequency declined for markers on T1.0 genomes; no effect was seen on recombination between markers on modified (T1.P) genomes. This suggested that recombination between unmodified genomes may be essential for their survival under conditions of host restriction. In a restricting host, genetic markers on T1.0 could recombine with T1.P even when the rescuing phage was added 6 min after T1.0 infection. However, even marker rescue recombination was diminished when protein synthesis was inhibited during early infection. Since DNA restriction is an early event, protein synthesis may be required soon after infection of a restricting host by T1.0 in order to preserve restriction-damaged DNA in a form that can participate in recombination. Experiments are also described that rule out some possibilities for the role of such a protein(s).

Recombination in bacteriophage T1 in the presence of host restriction.

When unmodified phage T1 infects restricting host cells at high multiplicities of infection, there is an increase in recombination frequency in all regions of the T1 map compared to the level of recombination in standard crosses when short distances are examined. The enhancement of recombination frequency is not uniform for all regions but is greatest for markers near the center of the map and not so great for markers near the ends. Crosses between markers at the extremities of the map show that there is no increase in recombination frequency under restriction conditions. An examination of phage T1 heterozygotes suggests that an increase of ends created by the process of P1 restriction increases recombination. When T1 crosses are done in the absence of host restriction, recombination defects in the host have no effect on phage recombination and we conclude that phage T1 codes for its own recombination genes. Host recombination functions are also dispensable for the recombination occurring during infection of restricting host cells by unmodified phage at high multiplicities of infection.