Mitochondrial DNA in yeast recombination and subsequent modification following mating between a grande and a suppressive petite

Summary The fluorinated pyrimidines 5-fluorouracil (5FU) and 5-fluorocytosine (5FC) induce the cytoplasmic petite mutation in the yeastSaccharomyces cerevisiae with high efficiency. It was found that in order to induce the mutation, 5FC must first be deaminated to 5FU. However, mutagenesis does not depend on the further conversion of 5FU to its deoxyriboside (5FUDR) and subsequent blockade of intracellular thymidine synthesis, since 5FUDR itself was found not to be mutagenic, and 5FU-induced mutagenesis was not antagonised by supplying thymidine monophosphate (dTMP) to a dTMP permeable strain. In any case, observations of the molecular changes accompanying petite induction in log phase cells ruled out the possibility that mutagenesis resulted simply from the dilution out of replication-blocked mitDNA molecules, since the appearance of mutants coincided with the synthesis of altered mitDNA molecules. In different strains, the resulting defective molecules were either maintained, giving rise to suppressive ^– petites, or completely degraded, to give pure clones of neutral ⁰ mutants. It is suggested that this degradative process was a consequence of the incorporation of 5FU into RNA.     

Sex differences in food acquisition and aggression in captive common marmosets (Callithrix jacchus)

Competition for food within the social group has been postulated as an important factor affecting primate social organization. This study examined how factors such as sex, aggression, dispersion of food, and amount of difficulty involved in obtaining food affect the distribution of food in common marmoset (Callithrix jacchus) social groups. Mated pairs of adultC. jacchus were presented with food that was either dispersed or concentrated, and either difficult or easy, to obtain. The number of food pieces obtained, aggressive displays, incidents of physical aggression, and amount of time spent searching for food were recorded for each animal. Neither dispersion nor difficulty in obtaining food had significant effects on the distribution of food or any of the other behaviors examined. The primary factors affecting food distribution were aggression and amount of time spent searching. Females obtained more food than males in all situations because they were more aggressive and apparently more motivated to search for food than were males.     

Trypanosomes contain two highly different isoforms of peroxin PEX13 involved in glycosome biogenesis

We previously identified the peroxin PEX13 in Trypanosoma brucei. Although lacking some features considered typical of PEX13s, it appeared functional in the biogenesis of glycosomes, the peroxisome-like organelles of trypanosomatids. Here we report the identification of a very different trypanosomatid PEX13, not containing the commonly encountered PEX13 SH3 domain but having other typical features. It is readily detected with the jackhmmer database search program, but not with PSI-BLAST. This is the first time different PEX13 isoforms are reported in a single organism. We show that this PEX13.2, like the PEX13.1 previously described, is associated with glycosomes and that its depletion by RNA interference affects the biogenesis of the organelles and viability of trypanosomes. The features considered typical of PEX13s are discussed.     

The frequency of hereditary defective mismatch repair in a prospective series of unselected colorectal carcinomas

A comprehensive analysis of somatic and germline mutations related to DNA mismatch-repair (MMR) genes can clarify the prevalence and mechanism of inactivation in colorectal carcinoma (CRC). In the present study, 257 unselected patients referred for CRC resection were examined for evidence of defective DNA MMR. In particular, we sought to determine the frequency of hereditary defects in DNA MMR in this cohort of patients. MMR status was assessed by testing of tumors for the presence or absence of hMLH1, hMSH2, and hMSH6 protein expression and for microsatellite instability (MSI). Of the 257 patients, 51 (20%) had evidence of defective MMR, demonstrating high levels of MSI (MSI-H) and an absence of either hMLH1 (n=48) or hMSH2 (n=3). All three patients lacking hMSH2, as well as one patient lacking hMLH1, also demonstrated an absence of hMSH6. DNA sequence analysis of the 51 patients with defective MMR revealed seven germline mutations-four in hMLH1 (two truncating and two missense) and three in hMSH2 (all truncating). A detailed family history was available for 225 of the 257 patients. Of the seven patients with germline mutations, only three had family histories consistent with hereditary nonpolyposis colorectal cancer. Of the remaining patients who had tumors with defective MMR, eight had somatic mutations in hMLH1. In addition, hypermethylation of the hMLH1 gene promoter was present in 37 (88%) of the 42 hMLH1-negative cases available for study and in all MSI-H tumors that showed loss of hMLH1 expression but no detectable hMLH1 mutations. Our results suggest that, although defective DNA MMR occurs in approximately 20% of unselected patients presenting for CRC resection, hereditary CRC due to mutations in the MMR pathway account for only a small proportion of patients. Of the 257 patients, only 5 (1.9%) appear to have unequivocal evidence of hereditary defects in MMR. The epigenetic (nonhereditary) mechanism of hMLH1 promoter hypermethylation appears to be responsible for the majority of the remaining patients whose tumors are characterized by defective DNA MMR.     

The Naturally Occurring Alleles of Mal1 in Saccharomyces Species Evolved by Various Mutagenic Processes Including Chromosomal Rearrangement

In order for a yeast strain to ferment maltose it must contain any one of the five dominant MAL loci. Each dominant MAL locus thus far analyzed contains three genes: GENE 1, encoding maltose permease, GENE 2 encoding maltase and GENE 3 encoding a positive trans-acting regulatory protein. In addition to these dominant MAL loci, several naturally occurring, partially functional alleles of MAL1 and MAL3 have been identified. Here, we present genetic and molecular analysis of the three partially functional alleles of MAL1: the MAL1p allele which can express only the MAL activator; the MAL1 g allele which can express both a maltose permease and maltase; and the mal1(0) allele which can express only maltase. Based on our results, we propose that the MAL1p, MAL1g and mal1(0) alleles evolved from the dominant MAL1 locus by a series of rearrangements and/or deletions of this yeast telomere-associated locus as well as by other mutagenic processes of gene inactivation. One surprising finding is that the MAL1g-encoded maltose permease exhibits little sequence homology to the MAL1-encoded maltose permease though they appear to be functionally homologous.     

Separation of carvedilol enantiomers in very small volumes of human plasma by capillary electrophoresis with laser-induced fluorescence

A sensitive capillary electrophoretic method for the determination of carvedilol enantiomers in 100 μl of human plasma has been developed and validated. Carvedilol and the internal standard carazolol are isolated from plasma samples by liquid–liquid extraction using diethylether. A sensitive and selective detection is provided by helium–cadmium laser-induced fluorescence. The total analysis time is 17.5 min, about 30 min are needed for the sample preparation. The linearity of the assay ranges from 1.56 to 50 ng/ml per carvedilol enantiomer. The limits of quantification (LOQ) for the carvedilol enantiomers in 100 μl of human plasma are 1.56 ng/ml. The inter-day accuracy for R-carvedilol is between 95.8 and 103% (104% at LOQ) and for S-carvedilol between 97.1 and 103% (107% at LOQ); the inter-day precision values are between 3.81 and 8.64% (10.9% at LOQ) and between 5.47 and 7.86% (7.91% at LOQ) for R- and S-carvedilol, respectively. The small sample volume needed is especially advantageous for the application in clinical studies in pediatric patients. As an application of the assay concentration/time profiles of the carvedilol enantiomers in a 5-year-old patient receiving a test dose of 0.09 mg/kg carvedilol are reported.