
Josef JiricnyETH Zurich | ETH Zürich · Department of Biology
Josef Jiricny
PhD
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299
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Introduction
Publications
Publications (299)
Prime editing (PE) is a powerful genome engineering approach that enables the introduction of base substitutions, insertions and deletions into any given genomic locus. However, the efficiency of PE varies widely and depends not only on the genomic region targeted, but also on the genetic background of the edited cell. Here, to determine which cell...
Prime editing is a powerful genome engineering approach that enables the introduction of base substitutions, insertions and deletions, into any given genomic locus. But prime editing, at even the same locus, can exhibit wildly different efficiencies in various cell backgrounds. It is unclear what determines these variations in efficiencies in a giv...
Germline mutations in the mismatch repair (MMR) genes MSH2, MSH6, MLH1 and PMS2 are linked to cancer of the colon and other organs, characterised by microsatellite instability and a large increase in mutation frequency. Unexpectedly, mutations in EXO1, encoding the only exonuclease genetically implicated in MMR, are not linked to familial cancer an...
FAN1, a DNA structure-specific nuclease, interacts with MLH1, but the repair pathways in which this complex acts are unknown. FAN1 processes DNA interstrand crosslinks (ICLs) and FAN1 variants are modifiers of the neurodegenerative Huntington’s disease (HD), presumably by regulating HD-causing CAG repeat expansions. Here, we identify specific amino...
Mutational signatures are imprints of pathophysiological processes arising through tumorigenesis. We generated isogenic CRISPR-Cas9 knockouts (Δ) of 43 genes in human induced pluripotent stem cells, cultured them in the absence of added DNA damage, and performed whole-genome sequencing of 173 subclones. ΔOGG1, ΔUNG, ΔEXO1, ΔRNF168, ΔMLH1, ΔMSH2, ΔM...
Mutational signatures are imprints of pathophysiological processes arising through tumorigenesis. We generated isogenic CRISPR–Cas9 knockouts (∆) of 43 genes in human induced pluripotent stem cells, cultured them in the absence of added DNA damage and performed whole-genome sequencing of 173 subclones. ∆OGG1, ∆UNG, ∆EXO1, ∆RNF168, ∆MLH1, ∆MSH2, ∆MS...
Inhibition of the TOR pathway (TORC2, or Ypk1/2), or the depolymerization of actin filaments results in catastrophic fragmentation of the yeast genome upon exposure to low doses of the radiomimetic drug Zeocin. We find that the accumulation of double-strand breaks (DSB) is not due to altered DSB repair, but by the uncoordinated activity of base exc...
p>Cancer whole-genome sequencing has revealed characteristic mutational signatures associated with defective DNA repair that underpin human genetic diseases. To define the direct mutagenic effects of DNA repair deficiency at the genome-wide level, we investigate mutational signatures generated by CRISPR-Cas9-based knockouts of 42 genes involved in...
Mutational signatures are imprints of pathophysiological processes arising through tumorigenesis. Here, we generate isogenic CRISPR-Cas9 knockouts (Δ) of 43 genes in human induced pluripotent stem cells, culture them in the absence of added DNA damage, and perform whole-genome sequencing of 173 daughter subclones. ΔOGG1, ΔUNG, ΔEXO1, ΔRNF168, ΔMLH1...
The mechanisms that underpin how insertions or deletions (indels) become fixed in DNA have primarily been ascribed to replication-related and/or double-strand break (DSB)-related processes. Here, we introduce a method to evaluate indels, orientating them relative to gene transcription. In so doing, we reveal a number of surprising findings: First,...
Replication factor C (RFC), a heteropentamer of RFC1-5, loads PCNA onto DNA during replication and repair. Once DNA synthesis has ceased, PCNA must be unloaded. Recent findings assign the uloader role primarily to an RFC-like (RLC) complex, in which the largest RFC subunit, RFC1, has been replaced with ATAD5 (ELG1 in Saccharomyces cerevisiae). ATAD...
The enhancer/promoter of the vitellogenin II gene (VTG) has been extensively studied as a model system of vertebrate transcriptional control. While deletion mutagenesis and in vivo footprinting identified the transcription factor (TF) binding sites governing its tissue specificity, DNase hypersensitivity and DNA methylation studies revealed the epi...
The enhancer/promoter of the vitellogenin II ( VT G) gene has been extensively studied as a model system of vertebrate transcriptional control. While deletion mutagenesis and in vivo footprinting identified the transcription factor (TF) binding sites governing its tissue specificity, DNase hypersensitivity- and DNA methylation studies revealed the...
The enhancer/promoter of the vitellogenin II (VTG) gene has been extensively studied as a model system of vertebrate transcriptional control. While deletion mutagenesis and in vivo footprinting identified the transcription factor (TF) binding sites governing its tissue specificity, DNase hypersensitivity- and DNA methylation studies revealed the ep...
Poly(ADP-ribose) polymerases (PARPs) facilitate the repair of DNA single-strand breaks (SSBs). When PARPs are inhibited, unrepaired SSBs colliding with replication forks give rise to cytotoxic double-strand breaks. These are normally rescued by homologous recombination (HR), but, in cells with suboptimal HR, PARP inhibition leads to genomic instabi...
Introductory paragraph
The mechanisms that underpin how insertions or deletions (indels) become fixed in DNA have primarily been ascribed to replication-related and/or double-strand break (DSB)-related processes. We introduce a novel way to evaluate indels, orientating them relative to gene transcription. In so doing, we reveal a number of surprisi...
The financial support for this Article was not fully acknowledged. The Acknowledgements should have included the following: This study was in part supported by the Swiss National Foundation Grant No.: 31003A-156023 to Alessandro Sartori.
Interstrand cross-link (ICL) hypersensitivity is a characteristic trait of Fanconi anemia (FA). Although FANCD2-associated nuclease 1 (FAN1) contributes to ICL repair, FAN1 mutations predispose to karyomegalic interstitial nephritis (KIN) and cancer rather than to FA. Thus, the biological role of FAN1 remains unclear. Because fork stalling in FAN1-...
Interstrand cross-link (ICL) hypersensitivity is a characteristic trait of Fanconi anemia (FA). Although FANCD2-associated nuclease 1 (FAN1) contributes to ICL repair, FAN1 mutations predispose to karyomegalic interstitial nephritis (KIN) and cancer rather than to FA. Thus, the biological role of FAN1 remains unclear. Because fork stalling in FAN1-...
DNA mismatch repair (MMR) is an evolutionarily-conserved process responsible for the repair of replication errors. In Escherichia coli, MMR is initiated by MutS and MutL, which activate MutH to incise transiently-hemimethylated GATC sites. MMR efficiency depends
on the distribution of these GATC sites. To understand which molecular events determine...
During class switch recombination (CSR), antigen-stimulated B-cells rearrange their immunoglobulin constant heavy chain (CH) loci to generate antibodies with different effector functions. CSR is initiated by activation-induced deaminase (AID), which
converts cytosines in switch (S) regions, repetitive sequences flanking the CH loci, to uracils. Alt...
Cisplatin and its derivatives, nitrogen mustards (NMs) and mitomycin C (MMC) are widely used in cancer chemotherapy. Their efficacy is linked primarily to their ability to generate DNA interstrand cross-links (ICLs), which effectively block the progression of transcription and replication machineries. Release of this block, referred to as unhooking...
RUVBL1 (RuvB-like1) and RUVBL2 (RuvB-like 2) are integral components of multisubunit protein complexes involved in processes ranging from cellular metabolism, transcription and chromatin remodeling to DNA repair. Here, we show that although RUVBL1 and RUVBL2 are known to form heterodimeric complexes in which they stabilize each other, the subunits...
Replicative DNA polymerases are high-fidelity enzymes that misincorporate nucleotides into nascent DNA with a frequency lower than 1/105, and this precision is improved to about 1/107 by their proofreading activity. Because this fidelity is insufficient to replicate most genomes without error, nature evolved postreplicative mismatch repair (MMR), w...
The cytotoxicity of SN1-type alkylating agents such as N-methyl-N'-nitrosourea (MNU), N-methyl-N'-nitro-N-nitrosoguanidine (MNNG), or the cancer chemotherapeutics temozolomide, dacarbazine and streptozotocin has been ascribed to the persistence of O(6)-methylguanine ((me)G) in genomic DNA. One hypothesis posits that (me)G toxicity is caused by futi...
Next-generation sequencing has revolutionized the search for disease-causing genetic alterations. Unfortunately, the task of distinguishing the handful of causative mutations from rare variants remains daunting. We now describe an assay that permits the analysis of all types of mutations in any gene of choice through the generation of stable human...
OBJECTIVES: The outcome of patients with primary melanoma (PM) cannot be completely explained based on currently adopted clinical-histopathologic criteria. In this study, we evaluated the potential prognostic value of mismatch repair protein expression in PMs.
METHODS: We examined the immunohistochemical staining of mismatch repair proteins in 18 b...
Mutations in the mismatch repair (MMR) genes MSH2, MSH6, MLH1 and PMS2 are associated with Lynch Syndrome (LS), a familial predisposition to early-onset cancer of the colon and other organs. Because
not all LS families carry mutations in these four genes, the search for cancer-associated mutations was extended to genes
encoding other members of the...
Biological processes are controlled by transcription networks. Expression changes of transcription factor (TF) genes in precancerous lesions are therefore crucial events in tumorigenesis. Our aim was to obtain a comprehensive picture of these changes in colorectal adenomas.
Using a 3-pronged selection procedure, we analyzed transcriptomic data on 3...
We previously reported that the expression of KIAA1199 in human colorectal tumors (benign and malignant) is markedly higher than that in the normal colonic mucosa. In this study, we investigated the functions of the protein encoded by this gene, which are thus far unknown. Immunostaining studies were used to reveal its subcellular localization, and...
KIAA1199 is an N-linked glycoprotein and a putative target of Wnt signaling. A. Western blot comparing KIAA1199 mobility in whole cell extracts from LS174T cells that were untreated, denatured by boiling at 100°C followed by PNGaseF treatment, or treated with PNGaseF alone. Mobility in the latter two extracts was similarly increased (vs. that obser...
Partial colocalization of KIAA1199 and ITPR3 in the ER. In general, the anti-KIAA1199 antibodies performed poorly in immunofluorescence experiments. However, a clear perinuclear staining in SW480 KIAA1199-Cl.18 cells strongly pointed to a localization of this protein in the ER. A similar staining pattern was detected with antibodies against ITPR3,...
Expression of KIAA1199 mRNA and protein in colorectal tissues and cell lines.
A. Expression values for KIAA1199 mRNA in 32 samples of normal colorectal mucosa, 32 colorectal adenomas, 25 colorectal cancers, and 8 colorectal cancer cell lines (Affymetrix U133Plus2.0 gene expression data from a previous study of ours [1]) B. Western blots showing KIA...
mRNA and protein expression levels of EPHA2 and ITPR3 in SW480 cell clones with or without KIAA1199. In triplicate microarray experiments, both EPHA2 and ITPR3 mRNA levels were found to be upregulated upon expression of KIAA1199 (panel A), but in qRT-PCR and Western blotting experiments appreciable increases were seen only in the expression of EPHA...
(A) Ectopic
KIAA1199 expression downregulates CTNNB1 expression in HEK293 cells. Cells were trasfected with an empty vector, Myc-tagged wild-type (CTNNB1wt) or Myc-tagged constitutively active beta-catenin (CTNNB1T41A), with (lanes 4–6) or without (lanes 1–3) KIAA1199. Western blotting performed 48 h after transfection revealed significant decrease...
List of putative KIAA1199 interactors identified by MS after immunoprecipitation. Molecular weight, cellular localization, functions, and number of unique peptide hits obtained on MS for each putative interactor is reported.
(PDF)
Genes whose expression significantly changed (p value <0.025, fold change ≥1.2) upon doxycycline-induced expression of KIAA1199 in SW480 Clone 13.
(PDF)
Epigenetic silencing of protein-encoding genes is common in early-stage colorectal tumorigenesis. Less is known about the methylation-mediated silencing of genes encoding microRNAs (miRNAs), which are also important epigenetic modulators of gene expression. Using quantitative PCR, we identified 56 miRNAs that were expressed in normal colorectal muc...
To improve replication fidelity, mismatch repair (MMR) must detect non-Watson-Crick base pairs and direct their repair to the nascent DNA strand. Eukaryotic MMR in vitro requires pre-existing strand discontinuities for initiation; consequently, it has been postulated that MMR in vivo initiates at Okazaki fragment termini in the lagging strand and a...
The mismatch repair (MMR) system detects non-Watson-Crick base pairs and strand misalignments arising during DNA replication and mediates their removal by catalyzing excision of the mispair-containing tract of nascent DNA and its error-free resynthesis. In this way, MMR improves the fidelity of replication by several orders of magnitude. It also ad...
The mammalian antibody repertoire is shaped by somatic hypermutation (SHM) and class switch recombination (CSR) of the immunoglobulin
(Ig) loci of B lymphocytes. SHM and CSR are triggered by non-canonical, error-prone processing of G/U mismatches generated by
activation-induced deaminase (AID). In birds, AID does not trigger SHM, but it triggers Ig...
Mismatch repair (MMR) is a key antimutagenic process that increases the fidelity of DNA replication and recombination. Yet genetic experiments showed that MMR is required for antibody maturation, a process during which the immunoglobulin loci of antigen-stimulated B cells undergo extensive mutagenesis and rearrangements. In an attempt to elucidate...
DNA interstrand crosslinks (ICLs) formed by antitumor agents, such as cisplatin or mitomycin C, are highly cytotoxic DNA lesions.
Their repair is believed to be triggered primarily by the stalling of replication forks at ICLs in S-phase. There is, however,
increasing evidence that ICL repair can also occur independently of replication. Using a repo...
Cells with DNA repair defects have increased genomic instability and are more likely to acquire secondary mutations that bring about cellular transformation. We describe the frequency and spectrum of somatic mutations involving several tumor suppressor genes in the rectal carcinoma of a 13-year-old girl harboring biallelic, germline mutations in th...
Cells with DNA repair defects have increased genomic instability and are more likely to acquire secondary mutations that bring about cellular transformation. We describe the frequency and spectrum of somatic mutations involving several tumor suppressor genes in the rectal carcinoma of a 13-year-old girl harboring biallelic, germline mutations in th...
A considerable surge of interest in the mismatch repair (MMR) system has been brought about by the discovery of a link between Lynch syndrome, an inherited predisposition to cancer of the colon and other organs, and malfunction of this key DNA metabolic pathway. This review focuses on recent advances in our understanding of the molecular mechanisms...
The LEM domain (for lamina-associated polypeptide, emerin, MAN1 domain) defines a group of nuclear proteins that bind chromatin through interaction of the LEM motif with the conserved DNA crosslinking protein, barrier-to-autointegration factor (BAF). Here, we describe a LEM protein annotated in databases as 'Ankyrin repeat and LEM domain-containing...
The small nematode Caenorhabditis elegans displays a spectrum of DNA damage responses similar to humans. In order to identify new DNA damage response genes, we isolated in a forward genetic screen 14 new mutations conferring hypersensitivity to ionizing radiation. We present here our characterization of lem-3, one of the genes identified in this sc...
YFP::LEM-3 expression pattern. (A) The transgene opIs383 [Pnpp-1::YFP::lem-3::3′UTRlem-3] rescues the hypersensitivity of lem-3(op444) mutants. Data represent the average of three experiments ± S.D. The progeny of 10 worms were analysed for each experiment. (B) Representative DIC and fluorescence images of embryos expressing YFP::LEM-3; in green: Y...
Chromatin segregation in wild-type embryos after irradiation. Wild-type hermaphrodites carrying the transgene GFP::H2B were irradiated with 30 Gy. Embryos were dissected 2 h after irradiation treatment. Images were taken every 20 sec (as described in Materials and Methods).
(RAR)
Embryonic lethality of isolated mutants. Mutants were irradiated with the indicated doses and embryonic lethality was scored. Data shown represent the average number of dead embryos of five hermaphrodites ± S.D.
(EPS)
Complementation tests.
lem-3(op444) fails to complement (A) rad-1(mn155) and (B) lem-3(tm3468). F1 embryonic lethality after irradiation with 30 Gy was quantified. Data shown represent the average embryonic lethality of the progeny of 10 hermaphrodites.
(EPS)
Enrichment of Sf9-expressed MBP-LEM-3 fusion protein. The protein, containing an N-terminal MBP-tag, was enriched over an Amylose-column. M: marker; prior inf.: prior infection; post inf.: post infection; sup.: supernatant; WT: LEM-3 wild type; F→L mutant corresponding to lem-3(op444).
(EPS)
Chromatin segregation defects in lem-3(op444) embryos after irradiation.
lem-3(op444) hermaphrodites carrying the transgene GFP::H2B were irradiated with 30 Gy. Embryos were dissected 2 h after irradiation treatment. Images were taken every 20 sec (as described in Materials and Methods).
(RAR)
Single strand nicks and gaps in DNA have been reported to increase the efficiency of nucleosome loading mediated by chromatin assembly factor 1 (CAF-1). However, on mismatch-containing substrates, these strand discontinuities are utilized by the mismatch repair (MMR) system as loading sites for exonuclease 1, at which degradation of the error-conta...
DNA interstrand crosslinks (ICLs) formed by antitumor agents, such as cisplatin or mitomycin C, are highly cytotoxic DNA lesions. Their repair is believed to be triggered primarily by the stalling of replication forks at ICLs in S-phase. There is, however, increasing evidence that ICL repair can also occur independently of replication. Using a repo...
Deleterious germ-line variants involving the DNA mismatch repair (MMR) genes have been identified as the cause of the hereditary nonpolyposis colorectal cancer syndrome known as the Lynch syndrome, but in numerous familial clusters of colon cancer, the cause remains obscure. We analyzed data for 235 German-speaking Swiss families with nonpolyposis...
Improved colonoscopy is revealing precancerous lesions that were frequently missed in the past, and ∼30% of those detected today have nonpolypoid morphologies ranging from slightly raised to depressed. To characterize these lesions molecularly, we assessed transcription of 23,768 genes in 42 precancerous lesions (25 slightly elevated nonpolypoid an...
Interstrand cross-links (ICLs) block replication and transcription and thus are highly cytotoxic. In higher eukaryotes, ICLs processing involves the Fanconi anemia (FA) pathway and homologous recombination. Stalled replication forks activate the eight-subunit FA core complex, which ubiquitylates FANCD2-FANCI. Once it is posttranslationally modified...
Cigarette smoke causes lung tumorigenesis; however, the mechanisms underlying transformation are unknown. We investigated if tobacco compounds induce DNA promoter hypermethylation in BEAS-2B cells treated with low doses of cigarette smoke condensate (CSC) for one month. Transcriptional profiles and anchorage-independent growth were explored using A...
Thymine DNA glycosylase (TDG) is a member of the uracil DNA glycosylase (UDG) superfamily of DNA repair enzymes. Owing to its ability to excise thymine when mispaired with guanine, it was proposed to act against the mutability of 5-methylcytosine (5-mC) deamination in mammalian DNA. However, TDG was also found to interact with transcription factors...
Fanconi anemia (FA) is a rare genetic disease characterized by congenital defects, bone marrow failure, chromosomal instability, and cancer susceptibility. One hallmark of cells from FA patients is hypersensitivity to interstrand cross-linking agents, such as the chemotherapeutics cisplatin and mitomycin C (MMC). We have recently characterized a FA...