A new type of radiation-sensitive mutant of S. cerevisiae is described. The recessive radH mutation sensitizes to the lethal effect of UV radiations haploids in the G1 but not in the G2 mitotic phase. Homozygous diploids are as sensitive as G1 haploids. The UV-induced mutagenesis is depressed, while the induction of gene conversion is increased. The mutation is believed to channel the repair of lesions engaged in the mutagenic pathway into a recombination process, successful if the events involve sister-chromatids but lethal if they involve homologous chromosomes. The sequence of the RADH gene reveals that it may code for a DNA helicase, with a Mr of 134 kDa. All the consensus domains of known DNA helicases are present. Besides these consensus regions, strong homologies with the Rep and UvrD helicases of E. coli were found. The RadH putative helicase appears to belong to the set of proteins involved in the error-prone repair mechanism, at least for UV-induced lesions, and could act in coordination with the Rev3 error-prone DNA polymerase.

Aims:To investigate the functional role of surface layer proteins (S-layer) in probiotic strain Lactobacillus acidophilus M92, especially its influence on adhesiveness to mouse ileal epithelial cells. Methods and Results: Sodium dodecyl sulphate polyacrylamide gel electrophoresis of cell surface proteins revealed the presence of potential surface layer (S-layer) proteins, ca at 45 kDa in L. acidophilus M92. Southern blot with pBK1 plasmid, containing slpA gene, gave a positive signal, suggesting that L. acidophilus M92 has a slpA gene coding for the S-layer proteins. S-layer proteins of this strain are present during all phases of growth. The S-layer proteins appeared when cells treated with 5 mol l )1 LiCl were allowed to grow again. Removal of the S-layer proteins reduced adhesion of L. acidophilus M92 to mouse ileal epithelial cells. Furthermore, the viability of cells without S-layer were reduced in simulated gastric juice at low pH range (2, 2AE5, 3) and simulated pancreatic juice with bile salts (1AE5 and 3 g l )1 ). S-layer proteins of L. acidophilus M92 were resistant to pepsin and pancreatin, in contrast, the treatment with proteinase K led to a significant proteolysis of the S-layer proteins.Conclusions: These results demonstrated functional role of S-layer; it is responsible for adhesiveness of Lactobacillus acidophilus M92 to mouse ileal epithelial cells and has a protective role for this strain. Significance and Impact of the Study: S-layer proteins have an important role in the establishment of probiotic strain Lactobacillus acidophilus M92 in the gastrointestinal tract.

The synbiotic effect of the oral treatment of Swiss albino mice with milk-based diets supplemented with Lactobacillus helveticus M92 and various kinds of prebiotics was investigated. Survival, competition, adhesion and colonization, as well as, immunomodulating capability of Lb. helveticus M92, in synbiotic combination, in the gastrointestinal tract (GIT) of mice, were monitored. After the mice were fed with synbiotics, the lactic acid bacteria (LAB) counts in faeces were increased and reduction of enterobacteria and sulphite-reducing clostridia was observed. Similar results were obtained in homogenates of small and large intestine of mice on the 1st and 14th day, after feeding with synbiotics. After the mice were orally given viable Lb. helveticus M92 cells, alone or in combination with prebiotic, the concentration of faecal SIgA and total serum IgA antibodies from all immunized mice were higher compared with the control. The specific humoral immune response was not evoked after oral administration, therefore their synbiotic application is suitable. Among inulin, lactulose and raffinose, Lb. helveticus M92 in combination with inulin, has shown the best synbiotic effect on intestinal and faecal microflora and immune system of mice.

Palindromic sequences are important DNA motifs involved in the regulation of different cellular processes, but are also a potential source of genetic instability. In order to initiate a systematic study of palindromes at the whole genome level, we developed a computer program that can identify, locate and count palindromes in a given sequence in a strictly defined way. All palindromes, defined as identical inverted repeats without spacer DNA, can be analyzed and sorted according to their size, frequency, GC content or alphabetically. This program was then used to prepare a catalog of all palindromes present in the chromosomal DNA of the yeast Saccharomyces cerevisiae. For each palindrome size, the observed palindrome counts were significantly different from those in the randomly generated equivalents of the yeast genome. However, while the short palindromes (2-12 bp) were under-represented, the palindromes longer than 12 bp were over-represented, AT-rich and preferentially located in the intergenic regions. The 44-bp palindrome found between the genes CDC53 and LYS21 on chromosome IV was the longest palindrome identified and contained only two C-G base pairs. Avoidance of coding regions was also observed for palindromes of 4-12 bp, but was less pronounced. Dinucleotide analysis indicated a strong bias against palindromic dinucleotides that could explain the observed short palindrome avoidance. We discuss some possible mechanisms that may influence the evolutionary dynamics of palindromic sequences in the yeast genome.

We investigated the influence of short terminal heterologies on recombination between transforming linear DNA fragments and the yeast Saccharomyces cerevisiae genome. The efficiency of plasmid integration to the CYC1 locus (ends-in assay) was decreased more than five-fold when the size of terminal heterology exceeded 28 nucleotides (nt) and a similar inhibitory effect was also observed in the ends-out assay (replacement of the ura3-52 allele by the URA3 gene). Plasmid integration occurred almost exclusively in the target homology and was accompanied by excessive degradation of the heterologous termini. Illegitimate integrations were much more frequent in the ends-out transformation in both the absence (8.9%) and the presence (23.7%) of 45/46 heterologous nucleotides at the ends of the transforming fragment. Interestingly, only about 60% of transformants arose by simple gene replacement, regardless of the presence of heterologous ends, whereas more complex interactions resulted in gene or whole chromosome duplications. Our results warn that different genetic alterations may be introduced in the host strain during ends-out transformation but also indicate possible mechanisms for formation of duplications in the genome.