This page has only limited features, please log in for full access.

Dr. Thomas Caspari
Paracelsus Medical University

Basic Info


Research Keywords & Expertise

0 DNA Repair
0 Microbiology
0 Philosophy
0 antibioitc resistance
0 Cell cycle checkpoint

Fingerprints

DNA Repair

Honors and Awards

The user has no records in this section


Career Timeline

The user has no records in this section.


Short Biography

The user biography is not available.
Following
Followers
Co Authors
The list of users this user is following is empty.
Following: 0 users

Feed

Review
Published: 27 May 2021 in Viruses
Reads 0
Downloads 0

The high sequence identity of the first SARS-CoV-2 samples collected in December 2019 at Wuhan did not foretell the emergence of novel variants in the United Kingdom, North and South America, India, or South Africa that drive the current waves of the pandemic. The viral spike receptor possesses two surface areas of high mutagenic plasticity: the supersite in its N-terminal domain (NTD) that is recognised by all anti-NTD antibodies and its receptor binding domain (RBD) where 17 residues make contact with the human Ace2 protein (angiotensin I converting enzyme 2) and many neutralising antibodies bind. While NTD mutations appear at first glance very diverse, they converge on the structure of the supersite. The mutations within the RBD, on the other hand, hone in on only a small number of key sites (K417, L452, E484, N501) that are allosteric control points enabling spike to escape neutralising antibodies while maintaining or even gaining Ace2-binding activity. The D614G mutation is the hallmark of all variants, as it promotes viral spread by increasing the number of open spike protomers in the homo-trimeric receptor complex. This review discusses the recent spike mutations as well as their evolution.

ACS Style

Anna Winger; Thomas Caspari. The Spike of Concern—The Novel Variants of SARS-CoV-2. Viruses 2021, 13, 1002 .

AMA Style

Anna Winger, Thomas Caspari. The Spike of Concern—The Novel Variants of SARS-CoV-2. Viruses. 2021; 13 (6):1002.

Chicago/Turabian Style

Anna Winger; Thomas Caspari. 2021. "The Spike of Concern—The Novel Variants of SARS-CoV-2." Viruses 13, no. 6: 1002.

Communication
Published: 13 December 2020 in Microorganisms
Reads 0
Downloads 0

The antibiotic nitrofurantoin is a furan flanked by a nitro group and a hydantoin ring. It is used to treat lower urinary tract infections (UTIs) that have a lifetime incidence of 50−60% in adult women. UTIs are typically caused by uropathogenic Escherichia coli (UPEC), which are increasingly expressing extended-spectrum beta-lactamases (ESBL), rendering them multi-drug resistant. Nitrofurantoin is a first-line treatment for gram-negative ESBL-positive UTI patients, given that resistance to it is still rare (0% to 4.4%). Multiplex PCR of β-lactamase genes of the blaCTX-M groups 1, 2, 9 and 8/25 from ESBL-positive UTI patients treated at three referral hospitals in North Wales (UK) revealed the presence of a novel CTX-M-14-like gene harbouring the missense mutations T55A, A273P and R277C. While R277 is close to the active site, T55 and A273 are both located in external loops. Recombinant expression of CTX-M-14 and the mutated CTX-M-14 in the periplasm of E. coli revealed a significant increase in the Minimum Inhibitory Concentration (MIC) for nitrofurantoin from ≥6 μg/mL (CTX-M-14) to ≥512 μg/mL (mutated CTX-M-14). Consistent with this finding, the mutated CTX-M protein hydrolysed nitrofurantoin in a cell-free assay. Detection of a novel nitrofurantoin resistance gene indicates an emerging clinical problem in the treatment of gram-negative ESBL-positive UTI patients.

ACS Style

Yasir Edowik; Thomas Caspari; Hugh Merfyn Williams. The Amino Acid Changes T55A, A273P and R277C in the Beta-Lactamase CTX-M-14 Render E. coli Resistant to the Antibiotic Nitrofurantoin, a First-Line Treatment of Urinary Tract Infections. Microorganisms 2020, 8, 1983 .

AMA Style

Yasir Edowik, Thomas Caspari, Hugh Merfyn Williams. The Amino Acid Changes T55A, A273P and R277C in the Beta-Lactamase CTX-M-14 Render E. coli Resistant to the Antibiotic Nitrofurantoin, a First-Line Treatment of Urinary Tract Infections. Microorganisms. 2020; 8 (12):1983.

Chicago/Turabian Style

Yasir Edowik; Thomas Caspari; Hugh Merfyn Williams. 2020. "The Amino Acid Changes T55A, A273P and R277C in the Beta-Lactamase CTX-M-14 Render E. coli Resistant to the Antibiotic Nitrofurantoin, a First-Line Treatment of Urinary Tract Infections." Microorganisms 8, no. 12: 1983.

Journal article
Published: 23 February 2018 in Cells
Reads 0
Downloads 0

The S. pombe checkpoint kinase, Cds1, protects the integrity of stalled DNA replication forks after its phosphorylation at threonine-11 by Rad3 (ATR). Modified Cds1 associates through its N-terminal forkhead-associated domain (FHA)-domain with Mrc1 (Claspin) at stalled forks. We report here that nutrient starvation results in post-translational changes to Cds1 and the loss of Mrc1. A drop in glucose after a down-shift from 3% to 0.1–0.3%, or when cells enter the stationary phase, triggers a sharp decline in Mrc1 and the accumulation of insoluble Cds1. Before this transition, Cds1 is transiently activated and phosphorylated by Rad3 when glucose levels fall. Because this coincides with the phosphorylation of histone 2AX at S129 by Rad3, an event that occurs towards the end of every unperturbed S phase, we suggest that a glucose limitation promotes the exit from the S phase. Since nitrogen starvation also depletes Mrc1 while Cds1 is post-translationally modified, we suggest that nutrient limitation is the general signal that promotes exit from S phase before it inactivates the Mrc1–Cds1 signalling component. Why Cds1 accumulates in resting cells while its activator Mrc1 declines is, as yet, unclear but suggests a novel function of Cds1 in non-replicating cells.

ACS Style

Jessica Fletcher; Liam Griffiths; Thomas Caspari. Nutrient Limitation Inactivates Mrc1-to-Cds1 Checkpoint Signalling in Schizosaccharomyces pombe. Cells 2018, 7, 15 .

AMA Style

Jessica Fletcher, Liam Griffiths, Thomas Caspari. Nutrient Limitation Inactivates Mrc1-to-Cds1 Checkpoint Signalling in Schizosaccharomyces pombe. Cells. 2018; 7 (2):15.

Chicago/Turabian Style

Jessica Fletcher; Liam Griffiths; Thomas Caspari. 2018. "Nutrient Limitation Inactivates Mrc1-to-Cds1 Checkpoint Signalling in Schizosaccharomyces pombe." Cells 7, no. 2: 15.

Journal article
Published: 06 October 2017 in Scientific Reports
Reads 0
Downloads 0

Chlorination of drinking water protects humans from water-born pathogens, but it also produces low concentrations of dibromoacetonitrile (DBAN), a common disinfectant by-product found in many water supply systems. DBAN is not mutagenic but causes DNA breaks and elevates sister chromatid exchange in mammalian cells. The WHO issued guidelines for DBAN after it was linked with cancer of the liver and stomach in rodents. How this haloacetonitrile promotes malignant cell transformation is unknown. Using fission yeast as a model, we report here that DBAN delays G1-S transition. DBAN does not hinder ongoing DNA replication, but specifically blocks the serine 345 phosphorylation of the DNA damage checkpoint kinase Chk1 by Rad3 (ATR) at broken replication forks. DBAN is particularly damaging for cells with defects in the lagging-strand DNA polymerase delta. This sensitivity can be explained by the dependency of pol delta mutants on Chk1 activation for survival. We conclude that DBAN targets a process or protein that acts at the start of S phase and is required for Chk1 phosphorylation. Taken together, DBAN may precipitate cancer by perturbing S phase and by blocking the Chk1-dependent response to replication fork damage.

ACS Style

Thomas Caspari; James Dyer; Nathalie Fenner; Christian Dunn; Chris Freeman. The drinking water contaminant dibromoacetonitrile delays G1-S transition and suppresses Chk1 activation at broken replication forks. Scientific Reports 2017, 7, 12730 .

AMA Style

Thomas Caspari, James Dyer, Nathalie Fenner, Christian Dunn, Chris Freeman. The drinking water contaminant dibromoacetonitrile delays G1-S transition and suppresses Chk1 activation at broken replication forks. Scientific Reports. 2017; 7 (1):12730.

Chicago/Turabian Style

Thomas Caspari; James Dyer; Nathalie Fenner; Christian Dunn; Chris Freeman. 2017. "The drinking water contaminant dibromoacetonitrile delays G1-S transition and suppresses Chk1 activation at broken replication forks." Scientific Reports 7, no. 1: 12730.

Journal article
Published: 01 October 2017 in Toxicology in Vitro
Reads 0
Downloads 0

Cylindrospermopsin (CYN) is a naturally occurring alkaloid produced by a variety of cyanobacteria and known to induce oxidative stress-mediated toxicity in eukaryotic cells. Despite extensive research on the mechanism of CYN toxicity, an understanding of the structural features responsible for this toxicity and the mechanism by which it can enter the cell are still not clear. It was established that the presence of both the uracil and guanidine groups is essential in biological activity of CYN whilst not much is known in this regard on the role of tether that separates them and the attached hydroxyl group. Therefore, in the present study we have prepared three synthetic analogues possessing uracil and guanidine groups separated by a variable length tether (4-6 carbons) and containing a hydroxyl function in a position orientation to CYN, together with a tetracyclic analogue of CYN lacking the hydroxyl group at C-7. The toxicity of these compounds was then compared with CYN and guanidinoacetate (GAA; the primary substrate in CYN biosynthesis) in an in vitro model using human neutrophils isolated from healthy subjects. The lowest activity measured by means of reactive oxygen species generation, lipid peroxidation and cell death was observed for GAA and the tetracyclic analogue. The greatest toxicity was found in an analogue with a 6-carbon tether, but all three analogues and CYN caused rapid onset of redox imbalance. These results add to the general understanding of CYN toxicity and preliminary findings suggest that the -OH group at C-7 may be significant for the cellular transport of CYN and/or be involved in its toxic activity inside the cell, a hypothesis which requires further testing.

ACS Style

Christopher Cartmell; Daniel M. Evans; Jessica Elwood; Hisham S. Fituri; Patrick J. Murphy; Thomas Caspari; Barbara Poniedziałek; Piotr Rzymski. Synthetic analogues of cyanobacterial alkaloid cylindrospermopsin and their toxicological activity. Toxicology in Vitro 2017, 44, 172 -181.

AMA Style

Christopher Cartmell, Daniel M. Evans, Jessica Elwood, Hisham S. Fituri, Patrick J. Murphy, Thomas Caspari, Barbara Poniedziałek, Piotr Rzymski. Synthetic analogues of cyanobacterial alkaloid cylindrospermopsin and their toxicological activity. Toxicology in Vitro. 2017; 44 ():172-181.

Chicago/Turabian Style

Christopher Cartmell; Daniel M. Evans; Jessica Elwood; Hisham S. Fituri; Patrick J. Murphy; Thomas Caspari; Barbara Poniedziałek; Piotr Rzymski. 2017. "Synthetic analogues of cyanobacterial alkaloid cylindrospermopsin and their toxicological activity." Toxicology in Vitro 44, no. : 172-181.

Other
Published: 16 July 2017
Reads 0
Downloads 0

Why the DNA damage checkpoint kinase Chk1 protects the genome of lower and higher eukaryotic cells differentially is still unclear. Mammalian Chk1 regulates replication origins, safeguards DNA replication forks and promotes fork progression. Conversely, yeast Chk1 acts only in G1 and G2. We report here that the mutation of serine 173 (S173A) in the activation loop of fission yeast Chk1 abolishes the G1-M and S-M checkpoints without affecting the G2-M arrest. Although Chk1-S173A is fully phosphorylated at serine 345 by the DNA damage sensor Rad3 (ATR) when DNA replication forks break, cells fail to stop the cell cycle. Mutant cells are uniquely sensitive to the DNA alkylation agent methyl- methanesulfate (MMS). This MMS sensitivity is genetically linked with the lagging strand DNA polymerase delta. Chk1-S173A is also unable to block mitosis when the G1 transcription factor Cdc10 is impaired. Serine 173 is equivalent to lysine 166 in human Chk1, an amino acid important for substrate specificity. We conclude that the removal of serine 173 impairs the phosphorylation of a Chk1 target that is important to protect cells from DNA replication stress.Summary statementMutation of serine-173 in the activation loop of Chk1 kinase may promote cancer as it abolishes the response to genetic alterations that arise while chromosomes are being copied.

ACS Style

Naomi Coulton; Thomas Caspari. The activation loop residue serine 173 of S.pombe Chk1 kinase is critical for the response to DNA replication stress. 2017, 164244 .

AMA Style

Naomi Coulton, Thomas Caspari. The activation loop residue serine 173 of S.pombe Chk1 kinase is critical for the response to DNA replication stress. . 2017; ():164244.

Chicago/Turabian Style

Naomi Coulton; Thomas Caspari. 2017. "The activation loop residue serine 173 of S.pombe Chk1 kinase is critical for the response to DNA replication stress." , no. : 164244.

Journal article
Published: 01 January 2017 in Biology Open
Reads 0
Downloads 0

While mammalian Chk1 kinase regulates replication origins, safeguards fork integrity and promotes fork progression, yeast Chk1 acts only in G1 and G2. We report here that the mutation of serine 173 (S173A) in the kinase domain of fission yeast Chk1 abolishes the G1-M and S-M checkpoints with little impact on the G2-M arrest. This separation-of-function mutation strongly reduces the Rad3-dependent phosphorylation of Chk1 at serine 345 during logarithmic growth, but not when cells experience exogenous DNA damage. Loss of S173 lowers the restrictive temperature of a catalytic DNA polymerase epsilon mutant (cdc20.M10) and is epistatic with a mutation in DNA polymerase delta (cdc6.23) when DNA is alkylated by methyl-methanesulfate (MMS). The chk1-S173A allele is uniquely sensitive to high MMS concentrations where it displays a partial checkpoint defect. A complete checkpoint defect occurs only when DNA replication forks break in cells without the intra-S phase checkpoint kinase Cds1. Chk1-S173A is also unable to block mitosis when the G1 transcription factor Cdc10 (cdc10.V50) is impaired. We conclude that serine 173, which is equivalent to lysine 166 in the activation loop of human Chk1, is only critical in DNA polymerase mutants or when forks collapse in the absence of Cds1.

ACS Style

Naomi Coulton; Thomas Caspari. The kinase domain residue serine 173 of S.pombe Chk1 kinase is critical for the response to DNA replication stress. Biology Open 2017, 6, 1840 -1850.

AMA Style

Naomi Coulton, Thomas Caspari. The kinase domain residue serine 173 of S.pombe Chk1 kinase is critical for the response to DNA replication stress. Biology Open. 2017; 6 (12):1840-1850.

Chicago/Turabian Style

Naomi Coulton; Thomas Caspari. 2017. "The kinase domain residue serine 173 of S.pombe Chk1 kinase is critical for the response to DNA replication stress." Biology Open 6, no. 12: 1840-1850.

Research article
Published: 01 July 2015 in PLOS ONE
Reads 0
Downloads 0

The activity of Cdc2 (CDK1) kinase, which coordinates cell cycle progression and DNA break repair, is blocked upon its phosphorylation at tyrosine 15 (Y15) by Wee1 kinase in the presence of DNA damage. How Cdc2 can support DNA repair whilst being inactivated by the DNA damage checkpoint remains to be explained. Human CDK1 is phosphorylated by Myt1 kinase at threonine 14 (T14) close to its ATP binding site before being modified at threonine 161 (T167Sp) in its T-loop by the CDK-activating kinase (CAK). While modification of T161 promotes association with the cyclin partner, phosphorylation of T14 inhibits the CDK1-cyclin complex. This inhibition is further enforced by the modification of Y15 by Wee1 in the presence of DNA lesions. In S.pombe, the dominant inhibition of Cdc2 is provided by the phosphorylation of Y15 and only a small amount of Cdc2 is modified at T14 when cells are in S phase. Unlike human cells, both inhibitory modifications are executed by Wee1. Using the novel IEFPT technology, which combines isoelectric focusing (IEF) with Phos-tag SDS electrophoresis (PT), we report here that S.pombe Cdc2 kinase exists in seven forms. While five forms are phosphorylated, two species are not. Four phospho-forms associate with cyclin B (Cdc13) of which only two are modified at Y15 by Wee1. Interestingly, only one Y15-modified species carries also the T14 modification. The fifth phospho-form has a low affinity for cyclin B and is neither Y15 nor T14 modified. The two unphosphorylated forms may contribute directly to the DNA damage response as only they associate with the DNA damage checkpoint kinase Chk1. Interestingly, cyclin B is also present in the unphosphorylated pool. We also show that the G146D mutation in Cdc2.1w, which renders Cdc2 insensitive to Wee1 inhibition, is aberrantly modified in a Wee1-dependent manner. In conclusion, our work adds support to the idea that two distinct Cdc2 pools regulate cell cycle progression and the response to DNA damage.

ACS Style

Thomas Caspari; Victoria Hilditch. Two Distinct Cdc2 Pools Regulate Cell Cycle Progression and the DNA Damage Response in the Fission Yeast S.pombe. PLOS ONE 2015, 10, e0130748 .

AMA Style

Thomas Caspari, Victoria Hilditch. Two Distinct Cdc2 Pools Regulate Cell Cycle Progression and the DNA Damage Response in the Fission Yeast S.pombe. PLOS ONE. 2015; 10 (7):e0130748.

Chicago/Turabian Style

Thomas Caspari; Victoria Hilditch. 2015. "Two Distinct Cdc2 Pools Regulate Cell Cycle Progression and the DNA Damage Response in the Fission Yeast S.pombe." PLOS ONE 10, no. 7: e0130748.

Journal article
Published: 26 May 2014 in Nucleic Acids Research
Reads 0
Downloads 0

Although it is well established that Cdc2 kinase phosphorylates the DNA damage checkpoint protein Crb253BP1 in mitosis, the full impact of this modification is still unclear. The Tudor-BRCT domain protein Crb2 binds to modified histones at DNA lesions to mediate the activation of Chk1 by Rad3ATR kinase. We demonstrate here that fission yeast cells harbouring a hyperactive Cdc2CDK1 mutation (cdc2.1w) are specifically sensitive to the topoisomerase 1 inhibitor camptothecin (CPT) which breaks DNA replication forks. Unlike wild-type cells, which delay only briefly in CPT medium by activating Chk1 kinase, cdc2.1w cells bypass Chk1 to enter an extended cell-cycle arrest which depends on Cds1 kinase. Intriguingly, the ability to bypass Chk1 requires the mitotic Cdc2 phosphorylation site Crb2-T215. This implies that the presence of the mitotic phosphorylation at Crb2-T215 channels Rad3 activity towards Cds1 instead of Chk1 when forks break in S phase. We also provide evidence that hyperactive Cdc2.1w locks cells in a G1-like DNA repair mode which favours non-homologous end joining over interchromosomal recombination. Taken together, our data support a model such that elevated Cdc2 activity delays the transition of Crb2 from its G1 to its G2 mode by blocking Srs2 DNA helicase and Casein Kinase 1 (Hhp1).

ACS Style

Salah Adam Mahyous Saeyd; Katarzyna Ewert-Krzemieniewska; Boyin Liu; Thomas Caspari. Hyperactive Cdc2 kinase interferes with the response to broken replication forks by trappingS.pombeCrb2 in its mitotic T215 phosphorylated state. Nucleic Acids Research 2014, 42, 7734 -7747.

AMA Style

Salah Adam Mahyous Saeyd, Katarzyna Ewert-Krzemieniewska, Boyin Liu, Thomas Caspari. Hyperactive Cdc2 kinase interferes with the response to broken replication forks by trappingS.pombeCrb2 in its mitotic T215 phosphorylated state. Nucleic Acids Research. 2014; 42 (12):7734-7747.

Chicago/Turabian Style

Salah Adam Mahyous Saeyd; Katarzyna Ewert-Krzemieniewska; Boyin Liu; Thomas Caspari. 2014. "Hyperactive Cdc2 kinase interferes with the response to broken replication forks by trappingS.pombeCrb2 in its mitotic T215 phosphorylated state." Nucleic Acids Research 42, no. 12: 7734-7747.

Review article
Published: 01 March 2014 in Open Biology
Reads 0
Downloads 0

Peregrine Laziosi (1265–1345), an Italian priest, became the patron saint of cancer patients when the tumour in his left leg miraculously disappeared after he developed a fever. Elevated body temperature can cause tumours to regress and sensitizes cancer cells to agents that break DNA. Why hyperthermia blocks the repair of broken chromosomes by changing the way that the DNA damage checkpoint kinases ataxia telangiectasia mutated (ATM) and ataxia telangiectasia and Rad3-related (ATR) are activated is an unanswered question. This review discusses the current knowledge of how heat affects the ATR–Chk1 and ATM–Chk2 kinase networks, and provides a possible explanation of why homeothermal organisms such as humans still possess this ancient heat response.

ACS Style

Thomas Turner; Thomas Caspari. When heat casts a spell on the DNA damage checkpoints. Open Biology 2014, 4, 140008 .

AMA Style

Thomas Turner, Thomas Caspari. When heat casts a spell on the DNA damage checkpoints. Open Biology. 2014; 4 (3):140008.

Chicago/Turabian Style

Thomas Turner; Thomas Caspari. 2014. "When heat casts a spell on the DNA damage checkpoints." Open Biology 4, no. 3: 140008.

Research article
Published: 28 June 2012 in PLOS Genetics
Reads 0
Downloads 0

DNA damage checkpoint activation can be subdivided in two steps: initial activation and signal amplification. The events distinguishing these two phases and their genetic determinants remain obscure. TopBP1, a mediator protein containing multiple BRCT domains, binds to and activates the ATR/ATRIP complex through its ATR-Activation Domain (AAD). We show that Schizosaccharomyces pombe Rad4TopBP1 AAD–defective strains are DNA damage sensitive during G1/S-phase, but not during G2. Using lacO-LacI tethering, we developed a DNA damage–independent assay for checkpoint activation that is Rad4TopBP1 AAD–dependent. In this assay, checkpoint activation requires histone H2A phosphorylation, the interaction between TopBP1 and the 9-1-1 complex, and is mediated by the phospho-binding activity of Crb253BP1. Consistent with a model where Rad4TopBP1 AAD–dependent checkpoint activation is ssDNA/RPA–independent and functions to amplify otherwise weak checkpoint signals, we demonstrate that the Rad4TopBP1 AAD is important for Chk1 phosphorylation when resection is limited in G2 by ablation of the resecting nuclease, Exo1. We also show that the Rad4TopBP1 AAD acts additively with a Rad9 AAD in G1/S phase but not G2. We propose that AAD–dependent Rad3ATR checkpoint amplification is particularly important when DNA resection is limiting. In S. pombe, this manifests in G1/S phase and relies on protein–chromatin interactions. DNA structure–dependent checkpoint activation and the amplification of checkpoint signals are carefully modulated to allow the checkpoint kinases to delay mitosis and regulate DNA metabolism. While much work has gone into understanding how this checkpoint functions, the mechanism by which the checkpoint signal is amplified is less clear. We have characterised a conserved domain in the Schizosaccharomyces pombe TopBP1 homolog, Rad4TopBP1 (also known as Cut5) that is capable of activating the ATR homolog Rad3ATR. We demonstrate that this domain is not required for initial checkpoint activation, but functions to amplify the checkpoint signal, likely when the presence of single-stranded DNA is limiting. Our data suggest that the function of the Rad4TopBP1 ATR-Activation Domain (AAD) is mediated by interactions between checkpoint proteins and phosphorylated histone H2A, which is itself promoted by Rad3ATR. We propose that the resulting amplification of the checkpoint signal is particularly important in G1-S phase, when resection is limited.

ACS Style

Su-Jiun Lin; Christopher Wardlaw; Takashi Morishita; Izumi Miyabe; Charly Chahwan; Thomas Caspari; Ulrike Schmidt; Antony M. Carr; Valerie Garcia. The Rad4TopBP1 ATR-Activation Domain Functions in G1/S Phase in a Chromatin-Dependent Manner. PLOS Genetics 2012, 8, e1002801 .

AMA Style

Su-Jiun Lin, Christopher Wardlaw, Takashi Morishita, Izumi Miyabe, Charly Chahwan, Thomas Caspari, Ulrike Schmidt, Antony M. Carr, Valerie Garcia. The Rad4TopBP1 ATR-Activation Domain Functions in G1/S Phase in a Chromatin-Dependent Manner. PLOS Genetics. 2012; 8 (6):e1002801.

Chicago/Turabian Style

Su-Jiun Lin; Christopher Wardlaw; Takashi Morishita; Izumi Miyabe; Charly Chahwan; Thomas Caspari; Ulrike Schmidt; Antony M. Carr; Valerie Garcia. 2012. "The Rad4TopBP1 ATR-Activation Domain Functions in G1/S Phase in a Chromatin-Dependent Manner." PLOS Genetics 8, no. 6: e1002801.

Journal article
Published: 01 January 2012 in Journal of Cell Science
Reads 0
Downloads 0

Exposure of human cells to heat switches DNA damage signaling from genotoxic to temperature stress. This change reduces mitotic commitment at the expense of DNA break repair. The thermal alterations behind this switch remain elusive despite the successful use of heat to sensitize cancer cells to DNA breaks. Rad9 is a highly conserved subunit of the Rad9-Rad1-Hus1 (9-1-1) checkpoint-clamp that is loaded by Rad17 onto damaged chromatin. At the DNA, Rad9 activates the checkpoint kinases Rad3ATR and Chk1 to arrest cells in G2. Using Schizosaccharomyces pombe as a model eukaryote, we discovered a new variant of Rad9, Rad9-M50, expression of which is specifically induced by heat. High temperatures promote alternative translation from a cryptic initiation codon at methionine-50. This process is restricted to cycling cells and independent of the temperature-sensing MAP kinase pathway. While full-length Rad9 delays mitosis in the presence of DNA lesions, Rad9-M50 functions in a remodeled checkpoint pathway to reduce mitotic commitment at elevated temperatures. This remodeled pathway still relies on Rad1 and Hus1, but acts independently of Rad17. Heat-induction of Rad9-M50 ensures that Chk1 kinase remains in a hypo-phosphorylated state. Elevated temperatures specifically reverse the DNA damage-induced modification of Chk1 in a manner dependent on Rad9-M50. Taken together, heat reprograms the DNA damage checkpoint at the level of Chk1 by inducing a Rad9 variant that can act outside of the canonical 9-1-1 complex.

ACS Style

Simon Janes; Ulrike Schmidt; Karim Ashour Garrido; Nadja Ney; Susanna Concilio; Mohamed Zekri; Thomas Caspari. Heat induction of a novel Rad9 variant from a cryptic translation initiation site reduces mitotic commitment. Journal of Cell Science 2012, 125, 4487 -4497.

AMA Style

Simon Janes, Ulrike Schmidt, Karim Ashour Garrido, Nadja Ney, Susanna Concilio, Mohamed Zekri, Thomas Caspari. Heat induction of a novel Rad9 variant from a cryptic translation initiation site reduces mitotic commitment. Journal of Cell Science. 2012; 125 (19):4487-4497.

Chicago/Turabian Style

Simon Janes; Ulrike Schmidt; Karim Ashour Garrido; Nadja Ney; Susanna Concilio; Mohamed Zekri; Thomas Caspari. 2012. "Heat induction of a novel Rad9 variant from a cryptic translation initiation site reduces mitotic commitment." Journal of Cell Science 125, no. 19: 4487-4497.

Journal article
Published: 01 April 2004 in Eukaryotic Cell
Reads 0
Downloads 0

The liz1 + gene of the fission yeast Schizosaccharomyces pombe was previously identified by complementation of a mutation that causes abnormal mitosis when ribonucleotide reductase is inhibited. Liz1 has similarity to transport proteins from Saccharomyces cerevisiae , but the potential substrate and its connection to the cell division cycle remain elusive. We report here that liz1 + encodes a plasma membrane-localized active transport protein for the vitamin pantothenate, the precursor of coenzyme A (CoA). Liz1 is required for pantothenate uptake at low extracellular concentrations. A lack of pantothenate uptake results in three phenotypes: (i) slow growth, (ii) delayed septation, and (iii) aberrant mitosis in the presence of hydroxyurea (HU). All three phenotypes are suppressed by high extracellular concentrations of pantothenate, where pantothenate uptake occurs by passive diffusion. liz1 Δ mutants are viable because they can synthesize pantothenate from uracil as an endogenous source. The use of uracil for both pantothenate biosynthesis and deoxyribonucleotide generation provides an explanation for the aberrant mitosis in the presence of HU. HU blocks ribonucleotide reductase, and we propose that the accumulation of ribonucleotides reduces uracil biosynthesis by feedback inhibition of aspartate transcarbamoylase. Thus, the addition of HU to liz1 Δ mutants results in a shortage of pantothenate. Because liz1 Δ mutants show striking similarities to mutants with defects in fatty acid biosynthesis, we propose that the shortage of pantothenate compromises fatty acid synthesis, resulting in slow growth and mitotic defects.

ACS Style

Jürgen Stolz; Thomas Caspari; Antony Michael Carr; Norbert Sauer. Cell Division Defects of Schizosaccharomyces pombe liz1 − Mutants Are Caused by Defects in Pantothenate Uptake. Eukaryotic Cell 2004, 3, 406 -412.

AMA Style

Jürgen Stolz, Thomas Caspari, Antony Michael Carr, Norbert Sauer. Cell Division Defects of Schizosaccharomyces pombe liz1 − Mutants Are Caused by Defects in Pantothenate Uptake. Eukaryotic Cell. 2004; 3 (2):406-412.

Chicago/Turabian Style

Jürgen Stolz; Thomas Caspari; Antony Michael Carr; Norbert Sauer. 2004. "Cell Division Defects of Schizosaccharomyces pombe liz1 − Mutants Are Caused by Defects in Pantothenate Uptake." Eukaryotic Cell 3, no. 2: 406-412.

Book chapter
Published: 01 January 2004 in The Molecular Biology of Schizosaccharomyces pombe
Reads 0
Downloads 0

The concept of checkpoint controls was first applied to biological systems by Weinert and Hartwell (Weinert and Hartwell 1988), when they observed that a rad9 mutant of S. cerevisiae was sensitive to DNA damage because it could not arrest the cell cycle before entering mitosis to allow time for the damage to be repaired. Subsequently, the checkpoint concept has been applied to many other biological processes, for example the monitoring of chromosome segregation during mitosis (Chap. 11). Checkpoints that respond directly to changes in DNA structure have become known as DNA-integrity checkpoints and encompass a number of related biological phenomena. In this chapter, we will run through the different DNA-integrity checkpoint sub-pathways, concentrating mainly on the DNA damage checkpoint about which the most is known. We will discuss the different checkpoint-protein complexes and how they might function in signal generation as well as in DNA repair and DNA replication. While checkpoint pathways were defined in terms of the induced delay they cause in cell cycle progression (Chap. 3), it has subsequently become clear that checkpoint pathways and proteins also participate in the coordination of repair with DNA replication and may regulate the choice of repair pathways for certain types of DNA damage (Chap. 7).

ACS Style

Antony M. Carr; Thomas Caspari. Checkpoint Controls Halting the Cell Cycle. The Molecular Biology of Schizosaccharomyces pombe 2004, 41 -56.

AMA Style

Antony M. Carr, Thomas Caspari. Checkpoint Controls Halting the Cell Cycle. The Molecular Biology of Schizosaccharomyces pombe. 2004; ():41-56.

Chicago/Turabian Style

Antony M. Carr; Thomas Caspari. 2004. "Checkpoint Controls Halting the Cell Cycle." The Molecular Biology of Schizosaccharomyces pombe , no. : 41-56.

Journal article
Published: 01 September 2003 in Journal of Cell Science
Reads 0
Downloads 0

The fission yeast BRCT domain protein Rad4/Cut5 is required for genome integrity checkpoint responses and DNA replication. Here we address the position at which Rad4/Cut5 acts within the checkpoint response pathways. Rad4 is shown to act upstream of the effector kinases Chk1 and Cds1, as both Chk1 phosphorylation and Cds1 kinase activity require functional Rad4. Phosphorylation of Rad9, Rad26 and Hus1 in response to either DNA damage or inhibition of DNA replication are independent of Rad4/Cut5 checkpoint function. Further we show that a novel, epitope-tagged allele of rad4+/cut5+ acts as a dominant suppressor of the checkpoint deficiencies of rad3-, rad26- and rad17- mutants. Suppression results in the restoration of mitotic arrest and is dependent upon the remaining checkpoint Rad proteins and the two effector kinases. High-level expression of the rad4+/cut5+ allele in rad17 mutant cells restores the nuclear localization of Rad9, but this does not fully account for the observed suppression. We conclude from these data that Rad4/Cut5 acts with Rad3, Rad26 and Rad17 to effect the checkpoint response, and a model for its function is discussed.

ACS Style

Sheila Harris; Caroline Kemplen; Thomas Caspari; Christopher Chan; Howard D. Lindsay; Marius Poitelea; Antony M. Carr; Clive Price. Delineating the position ofrad4+/cut5+ within the DNA-structure checkpoint pathways inSchizosaccharomyces pombe. Journal of Cell Science 2003, 116, 3519 -3529.

AMA Style

Sheila Harris, Caroline Kemplen, Thomas Caspari, Christopher Chan, Howard D. Lindsay, Marius Poitelea, Antony M. Carr, Clive Price. Delineating the position ofrad4+/cut5+ within the DNA-structure checkpoint pathways inSchizosaccharomyces pombe. Journal of Cell Science. 2003; 116 (17):3519-3529.

Chicago/Turabian Style

Sheila Harris; Caroline Kemplen; Thomas Caspari; Christopher Chan; Howard D. Lindsay; Marius Poitelea; Antony M. Carr; Clive Price. 2003. "Delineating the position ofrad4+/cut5+ within the DNA-structure checkpoint pathways inSchizosaccharomyces pombe." Journal of Cell Science 116, no. 17: 3519-3529.

Journal article
Published: 01 August 2003 in Molecular and Cellular Biology
Reads 0
Downloads 0

The Mre11-Rad50-Nbs1(Xrs2) complex and the Ku70-Ku80 heterodimer are thought to compete with each other for binding to DNA ends. To investigate the mechanism underlying this competition, we analyzed both DNA damage sensitivity and telomere overhangs in Schizosaccharomyces pombe rad50-d , rad50-d pku70-d , rad50-d exo1-d , and pku70-d rad50-d exo1-d cells. We found that rad50 exo1 double mutants are more methyl methanesulfonate (MMS) sensitive than the respective single mutants. The MMS sensitivity of rad50-d cells was suppressed by concomitant deletion of pku70 + . However, the MMS sensitivity of the rad50 exo1 double mutant was not suppressed by the deletion of pku70 + . The G-rich overhang at telomere ends in taz1-d cells disappeared upon deletion of rad50 + , but the overhang reappeared following concomitant deletion of pku70 + . Our data suggest that the Rad50 complex can process DSB ends and telomere ends in the presence of the Ku heterodimer. However, the Ku heterodimer inhibits processing of DSB ends and telomere ends by alternative nucleases in the absence of the Rad50-Rad32 protein complex. While we have identified Exo1 as the alternative nuclease targeting DNA break sites, the identity of the nuclease acting on the telomere ends remains elusive.

ACS Style

Kazunori Tomita; Akira Matsuura; Thomas Caspari; Antony M. Carr; Yufuko Akamatsu; Hiroshi Iwasaki; Ken'ichi Mizuno; Kunihiro Ohta; Masahiro Uritani; Takashi Ushimaru; Koichi Yoshinaga; Masaru Ueno. Competition between the Rad50 Complex and the Ku Heterodimer Reveals a Role for Exo1 in Processing Double-Strand Breaks but Not Telomeres. Molecular and Cellular Biology 2003, 23, 5186 -5197.

AMA Style

Kazunori Tomita, Akira Matsuura, Thomas Caspari, Antony M. Carr, Yufuko Akamatsu, Hiroshi Iwasaki, Ken'ichi Mizuno, Kunihiro Ohta, Masahiro Uritani, Takashi Ushimaru, Koichi Yoshinaga, Masaru Ueno. Competition between the Rad50 Complex and the Ku Heterodimer Reveals a Role for Exo1 in Processing Double-Strand Breaks but Not Telomeres. Molecular and Cellular Biology. 2003; 23 (15):5186-5197.

Chicago/Turabian Style

Kazunori Tomita; Akira Matsuura; Thomas Caspari; Antony M. Carr; Yufuko Akamatsu; Hiroshi Iwasaki; Ken'ichi Mizuno; Kunihiro Ohta; Masahiro Uritani; Takashi Ushimaru; Koichi Yoshinaga; Masaru Ueno. 2003. "Competition between the Rad50 Complex and the Ku Heterodimer Reveals a Role for Exo1 in Processing Double-Strand Breaks but Not Telomeres." Molecular and Cellular Biology 23, no. 15: 5186-5197.

Journal article
Published: 14 April 2003 in Genes & Development
Reads 0
Downloads 0

The signalosome is implicated in regulating cullin-dependent ubiquitin ligases. We find that two signalosome subunits, Csn1 and Csn2, are required to regulate ribonucleotide reductase (RNR) through the degradation of a small protein, Spd1, that acts to anchor the small RNR subunit in the nucleus. Spd1 destruction correlates with the nuclear export of the small RNR subunit, which, in turn, correlates with a requirement for RNR in replication and repair. Spd1 degradation is promoted by two separate CSN-dependent mechanisms. During unperturbed S phase, Spd1 degradation is independent of checkpoint proteins. In irradiated G2 cells, Spd1 degradation requires the DNA damage checkpoint. The signalosome copurifies with Pcu4 (cullin 4). Pcu4, Csn1, and Csn2 promote the degradation of Spd1, identifying a new function for the signalosome as a regulator of Pcu4-containing E3 ubiquitin ligase.

ACS Style

Cong Liu; Kelly A. Powell; Kirsten Mundt; Lejung Wu; Antony Michael Carr; Thomas Caspari. Cop9/signalosome subunits and Pcu4 regulate ribonucleotide reductase by both checkpoint-dependent and -independent mechanisms. Genes & Development 2003, 17, 1130 -1140.

AMA Style

Cong Liu, Kelly A. Powell, Kirsten Mundt, Lejung Wu, Antony Michael Carr, Thomas Caspari. Cop9/signalosome subunits and Pcu4 regulate ribonucleotide reductase by both checkpoint-dependent and -independent mechanisms. Genes & Development. 2003; 17 (9):1130-1140.

Chicago/Turabian Style

Cong Liu; Kelly A. Powell; Kirsten Mundt; Lejung Wu; Antony Michael Carr; Thomas Caspari. 2003. "Cop9/signalosome subunits and Pcu4 regulate ribonucleotide reductase by both checkpoint-dependent and -independent mechanisms." Genes & Development 17, no. 9: 1130-1140.

Journal article
Published: 15 May 2002 in Genes & Development
Reads 0
Downloads 0

The availability of a sister chromatid, and thus the cell cycle phase in which DNA double-strand breaks (DSBs) occur, influences the choice between homologous recombination (HR) or nonhomologous end joining (NHEJ). The sequential activation and destruction of CDK–cyclin activities controls progression through the cell cycle. Here we provide evidence that the major Schizosaccharomyces pombe CDK, Cdc2–cyclin B, influences recombinational repair of radiation-induced DSBs during the G2 phase at two distinct stages. At an early stage in HR, a defect in Cdc2 kinase activity, which is caused by a single amino acid change in cyclin B, affects the formation of Rhp51 (Rad51sp) foci in response to ionizing radiation in a process that is redundant with the function of Rad50. At a late stage in HR, low Cdc2–cyclin B activity prevents the proper regulation of topoisomerase III (Top3) function, disrupting a recombination step that occurs after the assembly of Rhp51 foci. This effect of Cdc2–cyclin B kinase on Top3 function is mediated by the BRCT-domain-containing checkpoint protein Crb2, thus linking checkpoint proteins directly with recombinational repair in G2. Our data suggest a model in which CDK activity links processing of recombination intermediates to cell cycle progression via checkpoint proteins.

ACS Style

Thomas Caspari; Johanne M. Murray; Antony Michael Carr. Cdc2-cyclin B kinase activity links Crb2 and Rqh1-topoisomerase III. Genes & Development 2002, 16, 1195 -1208.

AMA Style

Thomas Caspari, Johanne M. Murray, Antony Michael Carr. Cdc2-cyclin B kinase activity links Crb2 and Rqh1-topoisomerase III. Genes & Development. 2002; 16 (10):1195-1208.

Chicago/Turabian Style

Thomas Caspari; Johanne M. Murray; Antony Michael Carr. 2002. "Cdc2-cyclin B kinase activity links Crb2 and Rqh1-topoisomerase III." Genes & Development 16, no. 10: 1195-1208.

Dispatch
Published: 01 February 2002 in Current Biology
Reads 0
Downloads 0

How checkpoint pathways recognise double-strand breaks has long been a mystery. Recent studies have found that two distinct checkpoint protein complexes associate independently with chromatin at the sites of DNA damage. Why do two distinct mechanisms recognise strand lesions, and what does this tell us about the checkpoint pathways?

ACS Style

Thomas Caspari; Antony Michael Carr. Checkpoints: How to Flag Up Double-Strand Breaks. Current Biology 2002, 12, R105 -R107.

AMA Style

Thomas Caspari, Antony Michael Carr. Checkpoints: How to Flag Up Double-Strand Breaks. Current Biology. 2002; 12 (3):R105-R107.

Chicago/Turabian Style

Thomas Caspari; Antony Michael Carr. 2002. "Checkpoints: How to Flag Up Double-Strand Breaks." Current Biology 12, no. 3: R105-R107.

Journal article
Published: 01 April 2000 in Current Biology
Reads 0
Downloads 0
ACS Style

Thomas Caspari. Checkpoints: How to activate p53. Current Biology 2000, 10, R315 -R317.

AMA Style

Thomas Caspari. Checkpoints: How to activate p53. Current Biology. 2000; 10 (8):R315-R317.

Chicago/Turabian Style

Thomas Caspari. 2000. "Checkpoints: How to activate p53." Current Biology 10, no. 8: R315-R317.