Biomedical subjects
Publications and source records attributed to P Munz.
The evaluation of three-point crosses at the tetrad and random spore level leads to the conclusion that both chiasma and chromatid interference are absent in the fission yeast Schizosaccharomyces pombe.
Hybrid DNA with mismatched base pairs is a central intermediate of meiotic recombination. Mismatch repair leads either to restoration or conversion, while failure of repair results in postmeiotic segregation (PMS). The behavior of three G to C transversions in one-factor crosses with the wild-type alleles is studied in Schizosaccharomyces pombe. They lead to C/C and G/G mismatches and are compared with closely linked mutations yielding other mismatches. A method is presented for the detection of PMS in random spores. The procedure yields accurate PMS frequencies as shown by comparison with tetrad data. A scheme is presented for the calculation of the frequency of hybrid DNA formation and the efficiency of mismatch repair. The efficiency of C/C repair in S. pombe is calculated to be about 70%. Other mismatches are repaired with close to 100% efficiency. These results are compared with data published on mutations in Saccharomyces cerevisiae and Ascobolus immersus. This study forms the basis for the detailed analysis of the marker effects caused by G to C transversions in two-factor crosses.
The effect of the strong promoter from the alcohol dehydrogenase gene on mitotic and meiotic intragenic recombination has been studied at the ade6 locus of the fission yeast Schizosaccharomyces pombe. A 700-bp fragment containing the functional adh1 promoter was used to replace the weak wild-type promoter of the ade6 gene. Analysis of mRNA showed that strains with this ade6::adh1 fusion construct had strongly elevated ade6-specific mRNA levels during vegetative growth as well as in meiosis. These increased levels of mRNA correlated with a 20- to 25-fold stimulation of intragenic recombination in meiosis and a 7-fold increased prototroph formation during vegetative growth. Analysis of flanking marker configurations of prototrophic recombinants indicated that simple conversions as well as conversions associated with crossing over were stimulated in meiosis. The strongest stimulation of recombination was observed when the adh1 promoter was homozygous. Studies with heterologous promoter configurations revealed that the highly transcribed allele was the preferred acceptor of genetic information. The effect of the recombinational hot spot mutation ade6-M26 was also investigated in this system. Its effect was only partly additive to the elevated recombination rate generated by the ade6::adh1 fusion construct.
With the help of in vitro constructed intragenic double mutants, we investigated the influence of the recombinational hot spot mutation ade6-M26 on meiotic recombination between two additional ade6 mutations proximal to it. Recombination was stimulated four-fold when M26 was present in a heterozygous condition and ten-fold when homozygous. M26 itself remained unaffected in a substantial number of these events. This indicates that the stimulation can not only be due to a preferred conversion of M26 to wild-type with co-conversion of the second mutation in cis. A model is proposed in which M26 acts as an "entry site" for recombinational enzymes.
Intergenic conversion is a mechanism for the concerted evolution of repeated DNA sequences. A new approach for the isolation of intergenic convertants of serine tRNA genes in the yeast Schizosaccharomyces pombe is described. Contrary to a previous scheme, the intergenic conversion events studied in this case need not result in functional tRNA genes. The procedure utilizes crosses of strains that are homozygous for an active UGA suppressor tRNA gene, and the resulting progeny spores are screened for loss of suppressor activity. In this way, intergenic convertants of a tRNA gene are identified that inherit varying stretches of DNA sequence from either of two other tRNA genes. The information transferred between genes includes anticodon and intron sequences. Two of the three tRNA genes involved in these information transfers are located on different chromosomes. The results indicate that intergenic conversion is a conservative process. No infidelity is observed in the nucleotide sequence transfers. This provides further evidence for the hypothesis that intergenic conversion and allelic conversion are the result of the same molecular mechanism. The screening procedure for intergenic revertants also yields spontaneous mutations that inactivate the suppressor tRNA gene. Point mutations and insertions of A occur at various sites at low frequency. In contrast, A insertions at one specific site occur with high frequency in each of the three tRNA genes. This new type of mutation hot spot is found also in vegetative cells.
Successive rounds of mutagenesis of a Schizosaccharomyces pombe strain bearing the UGA-reading sup3 tRNASer suppressor have been carried out for two cycles of inactivation and reactivation of the suppressor. The suppressor phenotype at each stage was found to involve different combinations of three mutations, A30, A53, and A67, in the sup3-UGA gene. Single mutations A30 and A53 inactivate the suppressor as does the presence of all three mutations. A67 by itself is phenotypically neutral, but in combination with either A30 or A53 suppressor function is restored. The frequency with which these and other complementation events occur in S. pombe demonstrates a significant potential for nucleotide sequence evolution in tRNA. Differential expression of the S. pombe genes in Saccharomyces cerevisiae suggests that the two yeasts have diverged at the transcriptional and RNA processing level. Processing of the mutant tRNA precursors in S. cerevisiae reveals a hierarchy of structural domains within the tRNA that vary in their importance for RNase P cleavage.
In many cases the multiple genes coding for one specific tRNA are dispersed throughout the genome. The members of such a gene family nevertheless maintain a common nucleotide sequence during evolution. A major mechanism contributing to this concerted evolution is intergenic conversion. Here we show that it occurs between three tRNA genes of related sequence residing on different chromosomes of Schizosaccharomyces pombe. Sequence analysis of converted genes indicates that blocks of a minimal length of 18-33 bp and of a maximal length of 190 bp can be transferred from one gene to the other. During meiosis the frequency of these transfers lies in the order of 10(-5) per progeny spore. Information transfer between any two members of the gene family occurs in both directions.
Explore the source record for details and available documents.
We have studied the temporal and functional relationships between DNA replication and recombination in adenovirus-infected cells by using Southern blot hybridization to detect recombinant products among intracellular viral genomes. The data show that recombination can be detected soon after DNA replication has commenced and that the proportion of recombinant products increases thereafter. To determine the functional relationship between DNA replication and recombination, replication was blocked with the protein synthesis inhibitor anisomycin, the replication inhibitor cytosine arabinoside, and conditionally lethal mutations in either the virus-specified DNA-binding protein or the DNA polymerase. All treatments that directly or indirectly blocked DNA replication caused a delay in the appearance of recombinant products and a marked decline in their abundance relative to products of parental genotype. These data strongly suggest that DNA replication and recombination are interrelated, either because both processes share functions or because DNA structures produced by replication are suitable substrates for recombination. In addition, we have shown that some recombination function(s) is intrinsically thermolabile at 40.9 degrees C, even in wild-type crosses, since the appearance of recombinant products is delayed and their extent is reduced compared with that from crosses performed at 39.9 degrees C.
Explore the source record for details and available documents.
Explore the source record for details and available documents.
Explore the source record for details and available documents.
Explore the source record for details and available documents.
Explore the source record for details and available documents.
Explore the source record for details and available documents.
Explore the source record for details and available documents.
Explore the source record for details and available documents.
Explore the source record for details and available documents.