Browsing by Author "Kramer, W."

With recent progress in surgery and immunosuppression, more and more older men receive a kidney transplant. Thus, it is likely that the incidence of BPH in male transplant recipients is growing in parallel with age. Nonetheless, no data exist about diagnostic parameters for BPH in freshly transplanted male kidney allograft recipients. We evaluated whether established diagnostic and therapeutic criteria for BPH are valid for the evaluation of renal transplant recipients. BPH was diagnosed in 8 of 11 recipients older than 55 years. In all freshly transplanted renal allograft recipients, lower urinary tract symptoms (LUTS) were detected using an international. prostate symptoms score (IPSS). This score was 9.6 +/- 7.1 in patients without BPH, and significantly higher with 21.1 +/- 4.3 in patients with BPH. In receiver-operating characteristics (ROC) curve analysis a cut-off of 15.5 was calculated to distinguish best between BPH and non-BPH giving an accuracy of 90.2%. Acute urinary retention (AUR) was the predominant sign, which occurred in all BPH patients but only in 6.9% in non-BPH patients. Bladder outlet obstruction (BOO) was also common with a reduced uroflow with 9.5 +/- 2.2 ml/sec in non-BPH and 3.0 +/- 1.8 ml/sec in BPH (8/11 BPH-patients developed AUR prior to measurement). By digital rectal examinations, benign prostate enlargement was estimated as minimal in 10 of 11 cases of BPH. In urethrocystoscopy kissing lobes were detected in all cases of BPH. Since medical treatment with a-receptor antagonists was not successful, a surgical procedure using a transurethral resection was performed without any complications in all cases. Symptoms did not recur after resection, and BOO improved with increased uroflow measurements with 12.3 +/- 4.8 ml/sec 8 days after resection. We conclude that LUTS and BOO are common in freshly transplanted renal allograft recipients. The sudden onset of outlet obstruction without the potentiality of adaptation of urinary bladder may effect lower urinary tract symptoms and bladder outlet obstruction. We conclude that an elevated IPSS over 15.5 in combination with AUR and typical urethrocystoscopy results are the best methods to diagnose BPH. Conversely, our results indicate that uroflowmetry and digital rectal examination are neither sensitive nor specific. In addition, once BPH has been diagnosed and treatment with receptor antagonists does not relieve urinary tract symptoms, surgical resection should be considered.

We have characterized the MPH1 gene from Saccharomyces cerevisiae. mph1 mutants display a spontaneous mutator phenotype. Homologs were found in archaea and in the EST libraries of Drosophila, mouse, and man. Mph1 carries the signature motifs of the DEAH family of helicases. Selected motifs were shown to be necessary for MPH1 function by introducing missense mutations. Possible indirect effects on translation and splicing were excluded by demonstrating nuclear localization of the protein and splicing proficiency of the mutant. A mutation spectrum did not show any conspicuous deviations from wild type except for an underrepresentation of frameshift mutations. The mutator phenotype Mras dependent on REV3 and RAD6. The mutant was sensitive to MMS, EMS, 4-NQO, and camptothecin, but not to UV light and X rays. Epistasis analyses were carried out with representative mutants from various repair pathways (msh6, mag1, apn1, rad14, rad52, rad6, mms2, and rev3). No epistatic interactions were found, either for the spontaneous mutator phenotype or for MMS, EMS, and 4-NQO sensitivity. mph1 slightly increased the UV sensitivity of mmse, rad6, and rad14 mutants, but no effect on X-ray sensitivity was observed. These data suggest that MPH1 is not part of a hitherto known repair pathway. Possible functions are discussed.

The MPH1 (mutator pHenotype 1) gene of Saccharomyces cerevisiae was identified on the basis of elevated spontaneous mutation rates of haploid cells deleted for this gene. Further studies showed that MPH1 functions to channel DNA lesions into an error-free DNA repair pathway. The Mph1 protein contains the seven conserved motifs of the superfamily 2 (SF2) family of nucleic acid unwinding enzymes. Genetic analyses have found epistasis of the mph1 deletion with mutations in the RAD52 gene group that mediates homologous recombination and DNA repair by homologous recombination. To begin dissecting the biochemical functions of the MPH1-encoded product, we have expressed it in yeast cells and purified it to near homogeneity. We show that Mph1 has a robust ATPase function that requires single-stranded DNA for activation. Consistent with its homology to members of the SF2 helicase family, we find a DNA helicase activity in Mph1. We present data to demonstrate that the Mph1 DNA helicase activity is fueled by ATP hydrolysis and has a 3' to 5' polarity with respect to the DNA strand on which this protein translocates. The DNA helicase activity of Mph1 is enhanced by the heterotrimeric single-stranded DNA binding protein replication protein A. These results, thus, establish Mph1 as an ATP-dependent DNA helicase, and the availability of purified Mph1 should facilitate efforts at deciphering the role of this protein in homologous recombination and mutation avoidance.

DNA mismatch repair proteins play an important role in maintaining the integrity of the genetic information during replication and homologous recombination. The MutS-homologous (MSH) and MutL-homologous (MLH) proteins are highly conserved among all prokaryotes and eukaryotes. We have isolated two mutS homologous genes from Zea mays, named Mus1 and Mus2. Phylogenetic analysis identifies Mus1 as a member of the MSH2 protein family. Mus2 is an ortholog of the Arabidopsis thaliana MSH7 protein and belongs to a subgroup of MSH proteins that is possibly plant-specific. Mus1 and Mus2 are expressed at very low levels. Mus1 is located on chromosome 7L near locus b32B, and mus2 maps on chromosome 3S.

Thermus thermophilus HB27 is an extremely thermophilic, halotolerant bacterium, which was originally isolated from a natural thermal environment in Japan. This organism has considerable biotechnological potential; many thermostable proteins isolated from members of the genus Thermus are indispensable in research and in industrial applications. We present here the complete genome sequence of T. thermophilus HB27, the first for the genus Thermus. The genome consists of a 1,894,877 base pair chromosome and a 232,605 base pair megaplasmid, designated pTT27. The 2,218 identified putative genes were compared to those of the closest relative sequenced so far, the mesophilic bacterium Deinococcus radiodurans. Both organisms share a similar set of proteins, although their genomes lack extensive synteny. Many new genes of potential interest for biotechnological applications were found in T. thermophilus HB27. Candidates include various proteases and key enzymes of other fundamental biological processes such as DNA replication, DNA repair and RNA maturation.

We have analysed the levels of mRNA transcripts of the mutS- and mutL-homologous genes of the yeast Saccharomyces cerevisiae during the course of meiosis. by quantitative RT-PCR. We found that all mutS homologues (MSH1-6) were induced during meiosis, whereas no evidence for regulation of the mutL homologues (PMS1, MLH1-3) was obtained. Temporal expression patterns indicative of co-regulation were observed for the gene pairs MSH4/MSH5 and MSH2/SPO11. Sequence comparisons of the 5 ' flanking regions revealed similar sequence stretches in the respective gene pairs, which may constitute regulatory elements. Similar sequences were also found in the 5 ' flanking regions of the pairs MSH1/MSH3 and MSH1/MSH6. Upstream of MSH2 three closely spaced sequences similar to UAS(H) elements were found, which - surprisingly - are located within the coding region of SPO21. Deletion of these elements resulted in loss of meiotic induction of MSH2. Genetic analysis of homozygous deletion mutants did not reveal any differences from wild type with respect to genetic distance estimates, aberrant segregation, or suppression of homoeologus recombination in an interspecies cross with Saccharomyces paradoxus.

The MPH1 gene from Saccharomyces cerevisiae, encoding a member of the DEAH family of proteins, had been identified by Virtue of the spontaneous mutator phenotype of respective deletion mutants. Genetic analysis suggested that. MPH1 functions in a previously uncharacterized DNA repair pathway that protects the cells from damage-induced mutations. We have now analyzed genetic interactions of mph1 with a variety of mutants from different repair systems with respect to spontaneous mutation rates and sensitivities to different DNA-damaging agents. The dependence of the mph1 mutator phenotype on REV3 and REV1 and the synergy with mutations in base and nucleotide excision repair suggest an involvement of MPH1 in error-free bypass of lesions. However, although we observed an unexpected partial suppression of the mph1 mutator phenotype by rad5, genetic interactions with other mutations in postreplicative repair imply that MPH1 does not belong to this pathway. Instead, mutations from the homologous recombination pathway were found to be epistatic to mph1 with respect to both spontaneous mutation rates and damage sensitivities. Determination of spontaneous mitotic recombination rates demonstrated that mph1 mutants are not deficient in homologous recombination. On the contrary, in an sgs1 background we found a pronounced hyperrecombination phenotype. Thus, we propose that MPH1 is involved in a branch of homologous recombination that is specifically dedicated to error-free bypass.