Wolfram Siede - Academia.edu (original) (raw)

Papers by Wolfram Siede

Cold Spring Harbor Monograph Archive, 1991

I. INTRODUCTION The general topic of cellular responses to DNA damage in the yeast Saccharomyces ... more I. INTRODUCTION The general topic of cellular responses to DNA damage in the yeast Saccharomyces cerevisiae has been reviewed in several contexts (Lemontt 1980; Haynes and Kunz 1981; Lawrence 1982; Friedberg 1985a,b, 1988; von Borstel and Hastings 1985; Cooper and Kelly 1987; Moustacchi 1987). These reviews have documented the basic genetics and general biology of DNA repair and mutagenesis. In 1991, the essential question is no longer what are the phenomena of DNA repair and mutagenesis in yeast, but, rather, how are these elements of DNA metabolism transacted at the molecular level? Although the answers to this question are far from complete, the emphasis of this chapter is on the molecular biology and biochemistry of cellular responses to DNA damage, in keeping with the intended focus of these volumes. Strictly defined, the term DNA repair refers to cellular events associated with the removal of damaged, inappropriate, or mispaired bases from the genome of living cells (see Friedberg 1985a). In this narrow context, the topic is confined to (1) DNA damage reversal (e.g., monomerization of cyclobutyldipyrimidines [pyrimidine dimers] by DNA photolyase or demethylation of methylated [alkylated] bases by specific DNA methyltransferases); (2) base excision repair, whereby damaged bases (e.g., alkylated) or inappropriate bases (e.g., uracil) are excised by specific classes of repair enzymes designated DNA glycosylases and AP (apurinic/apyrimidinic) endonucleases; (3) nucleotide excision repair, whereby oligonucleotide tracts containing distortive bulky base adducts are excised; and (4) mismatch repair, whereby mismatched bases generated during semiconservative DNA synthesis or during recombination are excised.

The Internet journal of microbiology, 2010

A screen of the commercially available collection of haploid deletion mutants of Saccharomyces ce... more A screen of the commercially available collection of haploid deletion mutants of Saccharomyces cerevisiae for spontaneous mutator mutants newly identified a deletion of SRL3. This gene had been previously isolated as a suppressor of lethality of checkpoint kinase deletions if overexpressed. We found DNA damage sensitivity and extended checkpoint arrests to be associated with this strain. However, when crossed to wild-type, a mutant gene conferring these phenotypes was found to segregate from the SRL3 deletion. The mutation was identified as a Cterminal truncation of Mms2, an E2 ubiquitin conjugating enzyme involved in error-free replicative bypass of lesions. This confirmed an earlier report that Mms2 may be required to restrain error-prone polymerase ζ activity and underscored that residues of the C-terminus are necessary for Mms2 function. Srl3, on the other hand, does not appear to influence DNA damage sensitivity or spontaneous mutability if deleted. However, the absence of these phenotypes does not contradict its likely role as a positive regulator of dNTP levels.

Molecular Genetics And Genomics, 1986

A double mutant being thermoconditionally defective in mutation induction as well as in repair of... more A double mutant being thermoconditionally defective in mutation induction as well as in repair of pre-lethal UV-induced DNA damage (rev2ts) and deficient in excision repair (rad3-2) was studied in temperature-shift experiments. The influence of inhibitors of DNA replication (hydroxyurea, aphidicolin) was determined. Additionally, an analysis of the dose-response pattern of mutation induction ("mutation kinetics") at several ochre alleles was carried out. It was concluded that the UV-inducible REV2 dependent mutagenic repair process is not induced in excision-deficient cells. In excision-deficient cells, REV2 dependent mutation fixation is slow and mostly post-replicative though not dependent on DNA replication. The REV2 mediated mutagenic process could be separated from the repair function.

Current Genetics, Nov 1, 1981

Selection of mutants of Saccharomyces cerevisiae sensitive to the DNA cross-linking agent nitroge... more Selection of mutants of Saccharomyces cerevisiae sensitive to the DNA cross-linking agent nitrogen mustard (HN2) at two temperatures (23 °C and 36 °C) yielded two isolates with thermoconditionally enhanced (ts) sensitivity to the mutagen. Both were due to single recessive nuclear genes. Mutant allele snm1-2 (ts) showed mainly ts-sensitivity to HN2, whereas mutant allele snm2-1 (ts) conferred ts-sensitivity to HN2, half mustard (HN1) and UV. In temperature-shift experiments it was determined that the functions of SNM1 and SNM2 are needed for recovery within 6 to 7 h. after mutagen exposure during incubation at 23 °C on YEPD when HN2 and UV are applied. After HN1 treatment the SNM2 coded function is required for recovery for about 14 hrs. This possibly indicates a handling of UV- and HN2-induced lesions different from that of HN1-induced lesions.

PubMed, 1984

In a thermoconditional mutant of mutagenic DNA repair (rev 2ts = rad 5-8) of Saccharomyces cerevi... more In a thermoconditional mutant of mutagenic DNA repair (rev 2ts = rad 5-8) of Saccharomyces cerevisiae recovery of survival and mutation frequencies can be monitored by incubating UV-irradiated cells in growth medium at a permissive temperature (23 degrees C) before plating and a shift to restrictive temperature (36 degrees C). Inhibition of protein synthesis with cycloheximide during incubation at permissive conditions blocks this REV 2 dependent recovery process in stationary phase rev 2ts cells, whereas it can be reduced but not totally abolished in exponentially growing cells. These results indicate a strict dependence on post-irradiation protein synthesis in stationary phase cells and argue for a considerable constitutive level and only limited inducibility in logarithmic phase cells. The UV inducibility of the REV 2 coded function in stationary phase cells could be confirmed by analysis of the dose-response pattern of the his 5-2 reversion: in stationary phase rev 2ts cells, the quadratic component of the biphasic linear-quadratic induction kinetics found at 23 degrees C, which is interpreted as the consequence of induction of mutagenic repair, is eliminated at 36 degrees C.

Crosslinking agents such as psoralens, nitrogen mustards or cisplatin are bifunctionally acting c... more Crosslinking agents such as psoralens, nitrogen mustards or cisplatin are bifunctionally acting chemicals that generate a fraction of their adducts as covalent linkages between complementary deoxyribonucleic acid strands. Since many of these agents are of importance in genetic toxicology and cancer therapy, repair of interstrand crosslinks has been studied extensively in bacteria and in lower and higher eukaryotes. The main repair pathway in Escherichia coli involves the sequential action of nucleotide excision repair (NER) and recombinational repair. In eukaryotes, several repair pathways play important roles not only in repair including NER, translesion synthesis and recombination, but also mismatch repair. Relative contributions of the various pathways depend on cell cycle position and agent used. Eukaryotic proteins that specifically enhance resistance to crosslinking agents have been identified (FANC family of proteins, SNM1). Key Concepts: Chemicals with two or more correctly spaced reactive groups can covalently link opposing DNA strands. Several repair or tolerance pathways such as nucleotide excision repair, recombination and translesion synthesis can work together to overcome such complex damage. Cell cycle stage may determine the choice of repair pathway combinations. A heritable human syndrome with multiple diverse phenotypes (Fanconi anaemia) has been associated with defects in crosslink repair. By integrating in vitro studies and analysis of Fanconi proteins, current models of replication-dependent crosslink repair assume the creation of double strand breaks at stalled replication forks that are repaired by homologous recombination. Keywords: DNA repair; crosslinks; excision; bifunctional alkylation; recombination; translesion synthesis

Journal of Biological Chemistry, Mar 1, 2003

Molecular Biology of the Cell, Jun 1, 2005

How mitochondrial DNA (mtDNA) copy number is determined and modulated according to cellular deman... more How mitochondrial DNA (mtDNA) copy number is determined and modulated according to cellular demands is largely unknown. Our previous investigations of the related DNA helicases Pif1p and Rrm3p uncovered a role for these factors and the conserved Mec1/Rad53 nuclear checkpoint pathway in mtDNA mutagenesis and stability in Saccharomyces cerevisiae. Here, we demonstrate another novel function of this pathway in the regulation of mtDNA copy number. Deletion of RRM3 or SML1, or overexpression of RNR1, which recapitulates Mec1/Rad53 pathway activation, resulted in an approximately twofold increase in mtDNA content relative to the corresponding wild-type yeast strains. In addition, deletion of RRM3 or SML1 fully rescued the ϳ50% depletion of mtDNA observed in a pif1 null strain. Furthermore, deletion of SML1 was shown to be epistatic to both a rad53 and an rrm3 null mutation, placing these three genes in the same genetic pathway of mtDNA copy number regulation. Finally, increased mtDNA copy number via the Mec1/Rad53 pathway could occur independently of Abf2p, an mtDNA-binding protein that, like its metazoan homologues, is implicated in mtDNA copy number control. Together, these results indicate that signaling through the Mec1/Rad53 pathway increases mtDNA copy number by altering deoxyribonucleoside triphosphate pools through the activity of ribonucleotide reductase. This comprises the first linkage of a conserved signaling pathway to the regulation of mitochondrial genome copy number and suggests that homologous pathways in humans may likewise regulate mtDNA content under physiological conditions.

CRC Press eBooks, Sep 19, 2005

Humana Press eBooks, Feb 17, 2004

Biochemical and Biophysical Research Communications, Feb 1, 2009

The collection of gene deletion mutants of Saccharomyces cerevisiae was used to screen for novel ... more The collection of gene deletion mutants of Saccharomyces cerevisiae was used to screen for novel genes required for UV-induced mutagenesis. We found the SBF transcription factor (Swi4/Swi6 protein complex) to be required for wild-type levels of UV mutability in forward and reverse mutation assay. Expression of translesion polymerase ζ component Rev7 was identified as a target of SBFdependent regulation.

Springer eBooks, 1985

This article deals primarily with the practical aspects of mutagen testing with yeast. Equipment ... more This article deals primarily with the practical aspects of mutagen testing with yeast. Equipment necessary for a laboratory where mutagen testing with yeast is performed, and the most commonly used media, are listed. Some general procedures are described and, finally, for those who have little experience with work of this kind, a precise protocol is given for an experiment with stationary phase cells of the strain D7 of Saccharomyces cerevisiae using the heteroallelic ade2 system as the genetic endpoint. Some experimental data were obtained by students following this protocol using the direct-acting mutagen ethyl methanesulfonate (EMS); these data are discussed and analyzed. More details on the various genetic endpoints available in numerous yeast strains and on the interpretation of dose-dependence data, as well as an extended list of yeast literature, can be found in an article by Eckardt and von Borstel in this volume. Further technical advice is provided in our references to Zimmermann (1975), von Borstel (1981), and Zimmermann et al. (1984).

Current Genetics, May 1, 1982

Three haploid yeast mutants (snm) sensitive or thermoconditionally sensitive to the DNA cross-lin... more Three haploid yeast mutants (snm) sensitive or thermoconditionally sensitive to the DNA cross-linking agent nitrogen mustard (HN2) were crossed with four rad strains representing mutations in the three pathways of DNA dark repair. The resulting haploid double and triple mutant strains were tested for their sensitivity to UV, HN2 and HN1. From the observed epistatic or synergistic interactions of the combinations of mutant alleles we could derive the relation of the SNM1 and SNM2 genes to the postulated repair pathways. Alleles snm1-1 and snml-2 (ts) were found epistatic to genes of the rad3 group, whereas snm2-1 (ts) was epistatic to rad6. The snm1 and snm2 mutant alleles interacted synergistically. From these data it is concluded that the SNM1 gene product plays a cross-link specific role in excision repair while the SNM2 gene product may be involved in a system of error-prone repair.

Heliyon, Jul 1, 2018

Handling of the medically important dermatophyte Trichophyton rubrum in the laboratory typically ... more Handling of the medically important dermatophyte Trichophyton rubrum in the laboratory typically requires the generation of spores d for storage, treatment and plating when needed. The described method allows technically simple but efficient generation and harvesting of microconidia by cutting holes in Sabouraud dextrose agar medium that is covered by a mature T. rubrum mycelium.

Mutation research, Sep 1, 1995

The Internet journal of microbiology, May 28, 2013

For flow cytometry applications, chromosomal DNA of budding yeast cells has been stained routinel... more For flow cytometry applications, chromosomal DNA of budding yeast cells has been stained routinely with propidium iodide but SYBR® Green tends to yield superior results. Use of the recently introduced Midori Green results in staining of equal quality at significantly lower costs.

Molecular Genetics And Genomics, 1996

A mismatch repair-based model can explain some features of u.v.

DNA Repair and Mutagenesis, Second Edition, 2006

Featuring more than 10,000 references and a text lavishly complemented by over 700 illustrations,... more Featuring more than 10,000 references and a text lavishly complemented by over 700 illustrations, DNA Repair and Mutagenesis, 2nd Edition, is a timely update to the original edition published in 1995. The addition of three new authors, including an expert in the field of structural biology, ensures a comprehensive review of the most current research in diverse subject areas. An ideal textbook for advanced undergraduate and graduate students, the book is also an essential resource for all scientists researching cellular responses to DNA damage. Completely reorganized, the new edition presents a significant overhaul of the existing chapters and introduces important new material, reflective of the major changes and developments that have occurred in the field over the last decade. As with the first edition, this new volume presents the field within a strong historical framework and all aspects of biological responses to DNA damage are detailed. The book consists of 30 chapters divided into five main parts, covering: sources and consequences of DNA damage; correcting altered bases in DNA: DNA repair; DNA damage tolerance and mutagenesis; regulatory responses to DNA damage in eukaryotes; and disease states associated with defective biological responses to DNA damage. Basic principles are appropriately stressed; however, the book focuses on the significant progress to date and future research directions. Hardcover, 1,118 pages, two-color throughout with full-color insert, illustrations, index.

Cancer Research, 2011

The antibiotic salinomycin represents a selective potassium ionophore. It has recently attracted ... more The antibiotic salinomycin represents a selective potassium ionophore. It has recently attracted considerable attention due to its stem-cell specific mode of action as elucidated in a breast cancer stem cell model [1]. Further studies revealed additional benefits, e.g. the triggering of apoptosis in resistant cancer cell lines and reversal of multidrug resistance [2,3]. In budding yeast, we discovered that this agent is active and, similar to nigericin, prominently causes mitochondrial damage as revealed by the emergence of respiration-defective cells. We also noted the frequent occurrence of salinomycin-resistant clones which may prove to be an obstacle in chemotherapy. We used the collection of systematic gene deletions in haploid yeast to characterize genes whose deletion confers sensitivity or resistance to salinomycin if a carbon source is provided (glycerol) that requires active respiration for growth. With this toxicogenomics approach in a model organism, we attempt to elucid...