scholarly journals The Shu Complex Prevents Mutagenesis and Cytotoxicity of Single-Strand Specific Alkylation Lesions

2021 ◽  
Author(s):  
Braulio Bonilla ◽  
Alexander I Brown ◽  
Sarah R Hengel ◽  
Kyle S Rapchak ◽  
Debra Mitchell ◽  
...  
Keyword(s):  
Homologous Recombination ◽  
Ectopic Expression ◽  
Dna Lesions ◽  
Methyl Methanesulfonate ◽  
Base Pairing ◽  
Growth Defects ◽  
Dna Base ◽  
Genome Wide ◽  
A Genome ◽  
Methyl Cytosine

Three-methyl cytosine (3meC) are toxic DNA lesions, blocking base pairing. Bacteria and humans, express members of the AlkB enzymes family, which directly remove 3meC. However, other organisms, including budding yeast, lack this class of enzymes. It remains an unanswered evolutionary question as to how yeast repairs 3meC, particularly in single-stranded DNA. The yeast Shu complex, a conserved homologous recombination factor, aids in preventing replication-associated mutagenesis from DNA base damaging agents such as methyl methanesulfonate (MMS). We found that MMS-treated Shu complex-deficient cells, exhibit a genome-wide increase in A:T and G:C substitutions mutations. The G:C substitutions displayed transcriptional and replicational asymmetries consistent with mutations resulting from 3meC. Ectopic expression of a human AlkB homolog in Shu-deficient yeast rescues MMS-induced growth defects and increased mutagenesis. Finally, the Shu complex exhibits increased affinity for 3meC-containing DNA. Thus, our work identifies a novel mechanism for coping with alkylation adducts.

scholarly journals The chromatin remodeler ALC1 underlies resistance to PARP inhibitor treatment

2020 ◽  
Vol 6 (51) ◽  
pp. eabb8626
Author(s):  
Szilvia Juhász ◽  
Rebecca Smith ◽  
Tamás Schauer ◽  
Dóra Spekhardt ◽  
Hasan Mamar ◽  
...  
Keyword(s):  
Homologous Recombination ◽  
Parp Inhibitor ◽  
Parp Inhibitors ◽  
Dna Lesions ◽  
End Joining ◽  
Multiple Tumor ◽  
Genome Wide ◽  
A Genome ◽  
Inhibitor Resistance ◽  
Tumor Types

Poly(ADP-ribose) polymerase (PARP) inhibitors are used in the treatment of BRCA-deficient cancers, with treatments currently extending toward other homologous recombination defective tumors. In a genome-wide CRISPR knockout screen with olaparib, we identify ALC1 (Amplified in Liver Cancer 1)—a cancer-relevant poly(ADP-ribose)-regulated chromatin remodeling enzyme—as a key modulator of sensitivity to PARP inhibitor. We found that ALC1 can remove inactive PARP1 indirectly through binding to PARylated chromatin. Consequently, ALC1 deficiency enhances trapping of inhibited PARP1, which then impairs the binding of both nonhomologous end-joining and homologous recombination repair factors to DNA lesions. We also establish that ALC1 overexpression, a common feature in multiple tumor types, reduces the sensitivity of BRCA-deficient cells to PARP inhibitors. Together, we conclude that ALC1-dependent PARP1 mobilization is a key step underlying PARP inhibitor resistance.

scholarly journals Metabolic precision labeling enables selective probing of O-linkedN-acetylgalactosamine glycosylation

2020 ◽  
Vol 117 (41) ◽  
pp. 25293-25301
Author(s):  
Marjoke F. Debets ◽  
Omur Y. Tastan ◽  
Simon P. Wisnovsky ◽  
Stacy A. Malaker ◽  
Nikolaos Angelis ◽  
...  
Keyword(s):  
Cell Surface ◽  
Secretory Pathway ◽  
Ectopic Expression ◽  
Protein Glycosylation ◽  
Major Type ◽  
Metabolic Labeling ◽  
Uridine Diphosphate ◽  
Superresolution Microscopy ◽  
Genome Wide ◽  
A Genome

Protein glycosylation events that happen early in the secretory pathway are often dysregulated during tumorigenesis. These events can be probed, in principle, by monosaccharides with bioorthogonal tags that would ideally be specific for distinct glycan subtypes. However, metabolic interconversion into other monosaccharides drastically reduces such specificity in the living cell. Here, we use a structure-based design process to develop the monosaccharide probeN-(S)-azidopropionylgalactosamine (GalNAzMe) that is specific for cancer-relevant Ser/Thr(O)–linkedN-acetylgalactosamine (GalNAc) glycosylation. By virtue of a branchedN-acylamide side chain, GalNAzMe is not interconverted by epimerization to the correspondingN-acetylglucosamine analog by the epimeraseN-acetylgalactosamine–4-epimerase (GALE) like conventional GalNAc–based probes. GalNAzMe enters O-GalNAc glycosylation but does not enter other major cell surface glycan types including Asn(N)-linked glycans. We transfect cells with the engineered pyrophosphorylase mut-AGX1 to biosynthesize the nucleotide-sugar donor uridine diphosphate (UDP)-GalNAzMe from a sugar-1-phosphate precursor. Tagged with a bioorthogonal azide group, GalNAzMe serves as an O-glycan–specific reporter in superresolution microscopy, chemical glycoproteomics, a genome-wide CRISPR-knockout (CRISPR-KO) screen, and imaging of intestinal organoids. Additional ectopic expression of an engineered glycosyltransferase, “bump-and-hole” (BH)–GalNAc-T2, boosts labeling in a programmable fashion by increasing incorporation of GalNAzMe into the cell surface glycoproteome. Alleviating the need for GALE-KO cells in metabolic labeling experiments, GalNAzMe is a precision tool that allows a detailed view into the biology of a major type of cancer-relevant protein glycosylation.

scholarly journals Pan-cancer landscape of homologous recombination deficiency

2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Luan Nguyen ◽  
John W. M. Martens ◽  
Arne Van Hoeck ◽  
Edwin Cuppen
Keyword(s):  
Prostate Cancer ◽  
Homologous Recombination ◽  
Cancer Genomics ◽  
Strand Break ◽  
Diagnostic Value ◽  
Cancer Type ◽  
Homologous Recombination Deficiency ◽  
Genome Wide ◽  
A Genome ◽  
Pan Cancer

Abstract Homologous recombination deficiency (HRD) results in impaired double strand break repair and is a frequent driver of tumorigenesis. Here, we develop a genome-wide mutational scar-based pan-cancer Classifier of HOmologous Recombination Deficiency (CHORD) that can discriminate BRCA1- and BRCA2-subtypes. Analysis of a metastatic (n = 3,504) and primary (n = 1,854) pan-cancer cohort reveals that HRD is most frequent in ovarian and breast cancer, followed by pancreatic and prostate cancer. We identify biallelic inactivation of BRCA1, BRCA2, RAD51C or PALB2 as the most common genetic cause of HRD, with RAD51C and PALB2 inactivation resulting in BRCA2-type HRD. We find that while the specific genetic cause of HRD is cancer type specific, biallelic inactivation is predominantly associated with loss-of-heterozygosity (LOH), with increased contribution of deep deletions in prostate cancer. Our results demonstrate the value of pan-cancer genomics-based HRD testing and its potential diagnostic value for patient stratification towards treatment with e.g. poly ADP-ribose polymerase inhibitors (PARPi).

scholarly journals Systematic Analysis of Gibberellin Pathway Components in Medicago truncatula Reveals the Potential Application of Gibberellin in Biomass Improvement

2020 ◽  
Vol 21 (19) ◽  
pp. 7180
Author(s):  
Hongfeng Wang ◽  
Hongjiao Jiang ◽  
Yiteng Xu ◽  
Yan Wang ◽  
Lin Zhu ◽  
...  
Keyword(s):  
Medicago Truncatula ◽  
Expression Profiles ◽  
Expression Patterns ◽  
Ectopic Expression ◽  
Feedback Regulation ◽  
Specific Expression ◽  
Systematic Analysis ◽  
Model Legume ◽  
Genome Wide ◽  
A Genome

Gibberellins (GAs), a class of phytohormones, act as an essential natural regulator of plant growth and development. Many studies have shown that GA is related to rhizobial infection and nodule organogenesis in legume species. However, thus far, GA metabolism and signaling components are largely unknown in the model legume Medicago truncatula. In this study, a genome-wide analysis of GA metabolism and signaling genes was carried out. In total 29 components, including 8 MtGA20ox genes, 2 MtGA3ox genes, 13 MtGA2ox genes, 3 MtGID1 genes, and 3 MtDELLA genes were identified in M. truncatula genome. Expression profiles revealed that most members of MtGAox, MtGID1, and MtDELLA showed tissue-specific expression patterns. In addition, the GA biosynthesis and deactivation genes displayed a feedback regulation on GA treatment, respectively. Yeast two-hybrid assays showed that all the three MtGID1s interacted with MtDELLA1 and MtDELLA2, suggesting that the MtGID1s are functional GA receptors. More importantly, M. truncatula exhibited increased plant height and biomass by ectopic expression of the MtGA20ox1, suggesting that enhanced GA response has the potential for forage improvement.

scholarly journals AP4 is a mediator of epithelial–mesenchymal transition and metastasis in colorectal cancer

2013 ◽  
Vol 210 (7) ◽  
pp. 1331-1350 ◽  
Author(s):  
Rene Jackstadt ◽  
Simone Röh ◽  
Jens Neumann ◽  
Peter Jung ◽  
Reinhard Hoffmann ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Target Genes ◽  
Epithelial Mesenchymal Transition ◽  
Ectopic Expression ◽  
Migration And Invasion ◽  
Mesenchymal Transition ◽  
Helix Loop Helix ◽  
Genome Wide ◽  
A Genome ◽  
Tumor Types

The basic helix-loop-helix transcription factor AP4/TFAP4/AP-4 is encoded by a c-MYC target gene and displays up-regulation concomitantly with c-MYC in colorectal cancer (CRC) and numerous other tumor types. Here a genome-wide characterization of AP4 DNA binding and mRNA expression was performed using a combination of microarray, genome-wide chromatin immunoprecipitation, next-generation sequencing, and bioinformatic analyses. Thereby, hundreds of induced and repressed AP4 target genes were identified. Besides many genes involved in the control of proliferation, the AP4 target genes included markers of stemness (LGR5 and CD44) and epithelial–mesenchymal transition (EMT) such as SNAIL, E-cadherin/CDH1, OCLN, VIM, FN1, and the Claudins 1, 4, and 7. Accordingly, activation of AP4 induced EMT and enhanced migration and invasion of CRC cells. Conversely, down-regulation of AP4 resulted in mesenchymal–epithelial transition and inhibited migration and invasion. In addition, AP4 induction was required for EMT, migration, and invasion caused by ectopic expression of c-MYC. Inhibition of AP4 in CRC cells resulted in decreased lung metastasis in mice. Elevated AP4 expression in primary CRC significantly correlated with liver metastasis and poor patient survival. These findings imply AP4 as a new regulator of EMT that contributes to metastatic processes in CRC and presumably other carcinomas.

scholarly journals A genome-wide homologous recombination screen identifies the RNA-binding protein RBMX as a component of the DNA-damage response

2012 ◽  
Vol 14 (3) ◽  
pp. 318-328 ◽  
Author(s):  
Britt Adamson ◽  
Agata Smogorzewska ◽  
Frederic D. Sigoillot ◽  
Randall W. King ◽  
Stephen J. Elledge
Keyword(s):  
Dna Damage ◽  
Homologous Recombination ◽  
Dna Damage Response ◽  
Rna Binding ◽  
Binding Protein ◽  
Rna Binding Protein ◽  
Genome Wide ◽  
A Genome ◽  
Damage Response

scholarly journals SATB2 Overexpression Promotes the Proliferation, Migration and Invasion of Oral Squamous Cell Carcinoma by Up-Regulating NOX4

2020 ◽  
Author(s):  
Jie Gao ◽  
Ying Meng ◽  
Xin Ge ◽  
Chen-xing Hou ◽  
Qing-hai Zhu ◽  
...  
Keyword(s):  
Squamous Cell Carcinoma ◽  
Oral Squamous Cell Carcinoma ◽  
Cell Carcinoma ◽  
Squamous Cell ◽  
Ectopic Expression ◽  
Migration And Invasion ◽  
Clinical Samples ◽  
Genome Wide ◽  
A Genome ◽  
Sirna Knockdown

Abstract Background: While atypical expression of special AT-rich sequence-binding protein 2 (SATB2) has been approved associated with tumor progression, metastasis and unfavourable prognosis in various carcinomas. However, in oral squamous cell carcinoma (OSCC), both the expressive state and associated functions of SATB2’s are still undefined.Methods: Real-time PCR, western blotting, and immunohistochemistry were used to examine SATB2 expression. In vitro experiments including Flow Cytometry, CCK8 assay, migration assay, wound-healing assay were used to investigate the effects of SATB2 on HN4 cell proliferation, migration and invasion ability. Additionally, an orthotopic implantation assay was performed in nude mice to confirm the effects of SATB2 in vivo. Furthermore, a genome wide siRNA knockdown experiment was performed to explore the potential downstream regulatory mechanism of SATB2 in OSCC.Results: We found that , in clinical samples from a retrospective cohort of 58 OSCC patients, high expression of SATB2 is associated with poor prognosis of OSCC patients. In this study, we investigated SATB2 is highly expressed in OSCC tissues and cell lines ,which can promotes OSCC cells’ proliferation, migration, invasion and tumor growth. Following a genome wide siRNA knockdown experiment, we identified NOX4, a bona fide downstream target of SATB2, which can partially suppress OSCC proliferation. Furthermore, NOX4 knockdown inhibits tumorigenicity, which can be rescued partially by ectopic expression of SATB2.Conclusion: Our findings not only indicate overexpression of SATB2 triggers the proliferative, migratory and invasive mechanisms which are important in the malignant phenotype of OSCC, but also identify NOX4 as the downstream gene for SATB2. These findings indicate that SATB2 may play a key role in OSCC tumorigenicity and may be a future target for the development of new therapeutic regimens.

scholarly journals TRIM26 is a critical host factor for HCV replication and contributes to host tropism

2021 ◽  
Vol 7 (2) ◽  
pp. eabd9732
Author(s):  
Yisha Liang ◽  
Guigen Zhang ◽  
Qiheng Li ◽  
Lin Han ◽  
Xiaoyou Hu ◽  
...  
Keyword(s):  
Hepatoma Cell ◽  
Ectopic Expression ◽  
Host Factor ◽  
Hepatoma Cell Line ◽  
Genome Replication ◽  
Host Tropism ◽  
Host Interactions ◽  
Genome Wide ◽  
Factor Deficiency ◽  
A Genome

Hepatitis C virus (HCV) remains a major human pathogen that requires better understanding of virus-host interactions. In this study, we performed a genome-wide CRISPR-Cas9 screening and identified TRIM26, an E3 ligase, as a critical HCV host factor. Deficiency of TRIM26 specifically impairs HCV genome replication. Mechanistic studies showed that TRIM26 interacts with HCV-encoded NS5B protein and mediates its K27-linked ubiquitination at residue K51, and thus promotes the NS5B-NS5A interaction. Moreover, mouse TRIM26 does not support HCV replication because of its unique six–amino acid insert that prevents its interaction with NS5B. Ectopic expression of human TRIM26 in a mouse hepatoma cell line that has been reconstituted with other essential HCV host factors promotes HCV infection. In conclusion, we identified TRIM26 as a host factor for HCV replication and a new determinant of host tropism. These results shed light on HCV-host interactions and may facilitate the development of an HCV animal model.

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