Article
Biochemistry & Molecular Biology
Qingcai Meng, Daniel Stoyko, Celine Marlin Andrews, Parthena Konstantinidou, Pavol Genzor, O. Timothy, Alexandra R. Elchert, Leif Benner, Sushil Sobti, Esther Y. Katz, Astrid D. Haase
Summary: The combination of genome-editing and epitope tagging provides a powerful strategy to study proteins while preserving their physiological expression patterns. A simple and fast strategy to generate stable, endogenously tagged alleles is presented, allowing for the emulation of wild type protein expression and functionality.
NUCLEIC ACIDS RESEARCH
(2022)
Article
Biochemical Research Methods
Stefan J. Tekel, Nicholas Brookhouser, Kylie Standage-Beier, Xiao Wang, David A. Brafman
Summary: The introduction of transient reporters of editing enrichment (TREE) has enabled highly efficient single-base editing of human cells using a transient episomal fluorescent reporter, allowing for rapid generation of clonal editing efficiencies exceeding 80% in biallelic or multiplexed edited isogenic human pluripotent stem cell lines within approximately 3-4 weeks.
Article
Biochemistry & Molecular Biology
Ming Lyu, Yongsen Sun, Nana Yan, Qiang Chen, Xin Wang, Zehui Wei, Zhiying Zhang, Kun Xu
Summary: CRISPR/Cas9 is a powerful tool for gene editing, but it is challenging to screen modified cells. This study developed two novel traffic light screening reporters, PMG-SSA and PMG-HDR, to measure nuclease activity and select genetically modified cells. The reporters can be self-repaired, resulting in a functional selection cassette that can be used for screening. The results showed improved enrichment efficiencies for gene knockout and knock-in cells. These surrogate reporters advance CRISPR/Cas9-mediated editing in mammalian cells.
INTERNATIONAL JOURNAL OF BIOLOGICAL MACROMOLECULES
(2023)
Article
Biochemistry & Molecular Biology
Chengxi Sun, Chen Li, Wen Liu, Helgi B. Schioth
Summary: Humans and other organisms are exposed to thousands of chemicals, many of which are present in low concentrations and may have adverse effects on the circadian rhythm. Traditional toxicological methods are not effective for assessing complex mixtures of contaminants. This study establishes a cell line that can be used for high-throughput screening of chemicals that disrupt the circadian rhythm.
Article
Biochemistry & Molecular Biology
Hengxing Lu, Jun Liu, Tao Feng, Zihang Guo, Yunjun Yin, Fei Gao, Gengsheng Cao, Xuguang Du, Sen Wu
Summary: The authors developed an insertional mutagenesis strategy (HIT-trapping) and successfully created mutant alleles for 21 disease-related genes in porcine fibroblasts, achieving a significant improvement over previous methods.
Article
Biotechnology & Applied Microbiology
Margarita Stritzler, Cecilia Pascuan, Emilia Bottero, Cristina Gomez, Romina Frare, Andrea Puebla, Hiromi Tajima, Nicolas Ayub, Eduardo Blumwald, Gabriela Soto
Summary: This study demonstrates that next-generation sequencing can significantly reduce the time required for the identification of edited alleles in polyploid crops with complex genetics such as alfalfa. Through the analysis of fifteen edited events containing different doses of edited alleles, it was found that the use of NGS technology greatly improves the efficiency of determining the genotype of individual plants, which is attributed to the high output of sequences produced by this alternative genotyping approach.
PLANT CELL TISSUE AND ORGAN CULTURE
(2022)
Review
Biochemistry & Molecular Biology
Mark Soave, Leigh A. Stoddart, Carl W. White, Laura E. Kilpatrick, Joelle Goulding, Stephen J. Briddon, Stephen J. Hill
Summary: GPCRs, the largest family of membrane receptors, are major drug targets approved by the FDA. Recent advances in genome editing and fluorescent ligand design have provided new possibilities for studying GPCRs in their native environment. This review discusses the technical advances used to study the localization and ligand binding characteristics of genome-edited and endogenously expressed GPCRs.
Article
Biochemistry & Molecular Biology
Yixin Zhang, Yanlan Mo, Liyuan Han, Zhenyuan Sun, Wenzhong Xu
Summary: This study compared the transcriptomes of S. plumbizincicola and the non-hyperaccumulating ecotype of S. alfredii, revealing differentially expressed genes involved in heavy metal transport and detoxification in S. plumbizincicola. The highly expressed transcription factor genes SpARR11 and SpMYB84 were identified as potential regulators of Cd hyperaccumulation. Knockout mutations of these genes resulted in decreased Cd accumulation, suggesting their roles in the regulation of Cd hyperaccumulation in S. plumbizincicola. Transcriptome analysis combined with CRISPR/Cas9 technology provides valuable insights into the transcriptional regulation mechanism of hyperaccumulation in S. plumbizincicola.
INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES
(2023)
Article
Dermatology
Huishan Shi, Jos P. H. Smits, Ellen H. van den Bogaard, Matthew G. Brewer
Summary: The CRISPR/Cas9 technology is a powerful tool used to alter genetic landscapes in various hosts, particularly in developing novel mouse lines modeling disease states. It has been successfully used in both in vitro and in vivo systems within fields like epithelial biology and dermatology. The delivery of single-guide RNA/Cas9 editing complex to host cells is crucial for its success, and different delivery techniques have been explored and evaluated for their benefits and drawbacks in genetically modifying epidermal cells. Recent advances in dermatology have utilized these methods to better understand epidermal biology, identify therapeutic targets, and develop new treatment approaches for disease states.
JOURNAL OF INVESTIGATIVE DERMATOLOGY
(2021)
Article
Biology
Ganna Reint, Zhuokun Li, Kornel Labun, Salla Keskitalo, Inkeri Soppa, Katariina Mamia, Eero Tolo, Monika Szymanska, Leonardo A. Meza-Zepeda, Susanne Lorenz, Artur Cieslar-Pobuda, Xian Hu, Diana L. Bordin, Judith Staerk, Eivind Valen, Bernhard Schmierer, Markku Varjosalo, Jussi Taipale, Emma Haapaniemi
Summary: Precision CRISPR gene editing can be improved by fusing DNA repair proteins with Cas9, with different fusion proteins showing varying effects based on cell type and genomic site. Optimization is necessary to account for the diverse factors contributing to locus-specific genome editing outcomes.
Article
Agronomy
Jianwei Liu, Shufen Wang, Hao Wang, Bote Luo, Yiyong Cai, Xiaodong Li, Yanfeng Zhang, Xiaofeng Wang
Summary: The creation of two types of tomato nuclear male-sterile lines with different screening markers in a CRISPR/Cas9 system has made hybrid seed production in tomatoes more efficient. This approach allows for easy selection of the male-sterile lines at the seedling stage and ensures high seed purity during hybrid seed production.
MOLECULAR BREEDING
(2021)
Article
Multidisciplinary Sciences
Kendall J. Condon, Jose M. Orozco, Charles H. Adelmann, Jessica B. Spinelli, Pim W. van der Helm, Justin M. Roberts, Tenzin Kunchok, David M. Sabatini
Summary: Our study identified a catalog of genes that impact the mTORC1 pathway and clarified the multifaceted ways in which mTORC1 senses mitochondria dysfunction.AMPK and HRI are two of the key kinases that signal mitochondria distress to mTORC1, and their loss renders mTORC1 signaling largely resistant to mitochondria dysfunction induced by various inhibitors.
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
(2021)
Article
Immunology
Jack T. Whelan, Ragunath Singaravelu, Fuan Wang, Adrian Pelin, Levi A. Tamming, Giuseppe Pugliese, Nikolas T. Martin, Mathieu J. F. Crupi, Julia Petryk, Bradley Austin, Xiaohong He, Ricardo Marius, Jessie Duong, Carter Jones, Emily E. F. Fekete, Nouf Alluqmani, Andrew Chen, Stephen Boulton, Michael S. Huh, Matt Y. Tang, Zaid Taha, Elena Scut, Jean-Simon Diallo, Taha Azad, Brian D. Lichty, Carolina S. Ilkow, John C. Bell
Summary: The use of CRISPR/Cas9 assisted-recombinant vaccinia virus engineering system allows for the rapid generation of poxvirus vectors with multiple transgenes, enabling the creation of novel immunotherapies.
FRONTIERS IN IMMUNOLOGY
(2023)
Article
Biochemistry & Molecular Biology
Yihao Yang, Ziyan Shen, Youguang Li, Chenda Xu, Han Xia, Hao Zhuang, Shengyuan Sun, Min Guo, Changjie Yan
Summary: This article presents a method to improve rice cooking and eating quality by using CRISPR/Cas9 technology to knock out genes that encode grain storage proteins.
JOURNAL OF INTEGRATIVE PLANT BIOLOGY
(2022)
Article
Virology
Myung-Ju Lee, Jisu Lee, Su-Kyung Kang, Dagmar Wirth, Seung-Min Yoo, Changhoon Park, Myung-Shin Lee
Summary: This study investigated the role of cytokines in KSHV-infected endothelial cells by manipulating target genes using the CRISPR/Cas9 system. CXCL1 was found to be highly upregulated following KSHV infection and its knockout resulted in increased cell death and suppressed phosphorylation of STAT3. These findings suggest that CXCL1 production is essential for the survival of KSHV-infected endothelial cells.
JOURNAL OF MEDICAL VIROLOGY
(2023)
Article
Multidisciplinary Sciences
Guillermo Valenzuela Nieto, Ronald Jara, Daniel Watterson, Naphak Modhiran, Alberto A. Amarilla, Johanna Himelreichs, Alexander A. Khromykh, Constanza Salinas-Rebolledo, Teresa Pinto, Yorka Cheuquemilla, Yago Margolles, Natalia Lopez Gonzalez del Rey, Zaray Miranda-Chacon, Alexei Cuevas, Anne Berking, Camila Deride, Sebastian Gonzalez-Moraga, Hector Mancilla, Daniel Maturana, Andreas Langer, Juan Pablo Toledo, Ananda Muller, Benjamin Uberti, Paola Krall, Pamela Ehrenfeld, Javier Blesa, Pedro Chana-Cuevas, German Rehren, David Schwefel, Luis Angel Fernandez, Alejandro Rojas-Fernandez
Summary: Despite global efforts to develop SARS-CoV-2 treatments, the urgent need for effective diagnosis, treatment, and prophylactic measures remains critical. The development of alpaca Nanobody W25 shows promise as a potential antiviral agent, efficiently neutralizing SARS-CoV-2 strains with high affinity.
SCIENTIFIC REPORTS
(2021)
Article
Geriatrics & Gerontology
Natalia Lopez-Gonzalez del Rey, Tiziano Balzano, Lucia Martin-Rodriguez, Constanza Salinas-Rebolledo, Ines Trigo-Damas, Alejandro Rojas-Fernandez, Lydia Alvarez-Erviti, Javier Blesa
Summary: Proteinaceous inclusions known as LBs are used as a pathological hallmark for PD, with early deposits in the OB and amygdala containing insoluble alpha-synuclein and other ubiquitinated proteins. Injecting lactacystin into the OB and amygdala of mice did not result in significant differences in neuronal numbers, alpha-synuclein expression, or dopaminergic integrity, indicating that long-term injections in extra nigrostriatal regions may not mimic the spreading aspects of PD.
FRONTIERS IN AGING NEUROSCIENCE
(2021)
Article
Cell Biology
Gabriela Vargas, Omar Cortes, Eloisa Arias-Munoz, Sergio Hernandez, Cristobal Cerda-Troncoso, Laura Hernandez, Alexis E. Gonzalez, Michael H. Tatham, Hianara A. Bustamante, Claudio Retamal, Jorge Cancino, Manuel Varas-Godoy, Ronald T. Hay, Alejandro Rojas-Fernandez, Viviana A. Cavieres, Patricia V. Burgos
Summary: Macroautophagy and the ubiquitin proteasome system are interconnected in maintaining cellular homeostasis. HERPUD1, a protein regulated by proteasomal degradation, stabilizes ER and lysosomal network and negatively regulates autophagy, promoting cell survival under stress conditions.
FRONTIERS IN CELL AND DEVELOPMENTAL BIOLOGY
(2022)
Article
Biochemistry & Molecular Biology
Eduardo A. Albornoz, Alberto A. Amarilla, Naphak Modhiran, Sandra Parker, Xaria X. Li, Danushka K. Wijesundara, Julio Aguado, Adriana Pliego Zamora, Christopher L. D. McMillan, Benjamin Liang, Nias Y. G. Peng, Julian D. J. Sng, Fatema Tuj Saima, Jenny N. Fung, John D. Lee, Devina Paramitha, Rhys Parry, Michael S. Avumegah, Ariel Isaacs, Martin W. Lo, Zaray Miranda-Chacon, Daniella Bradshaw, Constanza Salinas-Rebolledo, Niwanthi W. Rajapakse, Ernst J. Wolvetang, Trent P. Munro, Alejandro Rojas-Fernandez, Paul R. Young, Katryn J. Stacey, Alexander A. Khromykh, Keith J. Chappell, Daniel Watterson, Trent M. Woodruff
Summary: COVID-19 infection may lead to neurological symptoms similar to Parkinson's disease. The study found that SARS-CoV-2 and spike protein can directly activate the NLRP3 inflammasome in microglial cells, which may contribute to the development of neuroinflammation.
MOLECULAR PSYCHIATRY
(2023)
Review
Hematology
Elena Martinez-Balsalobre, Jean-Hugues Guervilly, Jenny van Asbeck van der Wijst, Ana Belen Perez-Oliva, Christophe Lachaud
Summary: Fanconi anemia is a rare inherited disorder that affects the bone marrow and leads to decreased production of blood cells. Defective repair of DNA interstrand crosslinks is one of the causes, with mutations in over 20 genes linked to the disease. Advances in science and molecular biology have shed light on the relationship between FA gene mutations and the severity of clinical manifestations. This article highlights the current and promising therapeutic options for this rare disease.
Article
Cell Biology
Ellis G. Jaffray, Michael H. Tatham, Barbara Mojsa, Magda Liczmanska, Alejandro Rojas-Fernandez, Yili Yin, Graeme Ball, Ronald T. Hay
Summary: Jaffray et al. demonstrated that the degradation of PML and PML-RARA induced by arsenic relies on the VCP/p97 segregase protein. Arsenic trioxide treatment leads to degradation of PML-RARA and PML, curing the disease. The UFD1-NPLOC4-p97 segregase complex is essential for extracting poly-ubiquitinated, poly-SUMOylated PML from PML bodies prior to proteasomal degradation.
JOURNAL OF CELL BIOLOGY
(2023)
Article
Medicine, Research & Experimental
Lola Rodriguez-Ruiz, Juan M. Lozano-Gil, Elena Naranjo-Sanchez, Elena Martinez-Balsalobre, Alicia Martinez-Lopez, Christophe Lachaud, Miguel Blanquer, Toan K. Phung, Diana Garcia-Moreno, Maria L. Cayuela, Sylwia D. Tyrkalska, Ana B. Perez-Oliva, Victoriano Mulero
Summary: Chronic inflammatory diseases are associated with changes in hematopoiesis, including increased neutrophil counts and anemia. The NLRP1 inflammasome regulates hematopoiesis through the cleavage of GATA1, resulting in neutrophil reduction and increased erythrocyte counts. Inhibition of NLRP1 by LRRFIP1 and FLII, along with activation of the ZAKa/P38 kinase axis, promotes erythroid differentiation and offers potential therapeutic targets for hematopoietic alterations associated with chronic inflammatory and rare diseases.
EMBO MOLECULAR MEDICINE
(2023)
Article
Virology
Wing-Hang Ip, Michael H. Tatham, Steewen Krohne, Julia Gruhne, Michael Melling, Tina Meyer, Britta Gornott, Luca D. Bertzbach, Ronald T. Hay, Estefania Rodriguez, Thomas Dobner
Summary: This study used quantitative SUMO proteomics to analyze the cellular consequences of E1B-55K-mediated host cell modulation and adenovirus infection. The results showed that changes in the SUMOylated proteome could regulate DNA damage response, cell cycle control, chromatin assembly, and gene transcription. Additionally, a SUMO-dependent, ubiquitin-mediated degradation mechanism for some SUMO substrates was identified, suggesting that E1B-55K might use multiple mechanisms to alter restrictive cellular pathways.
JOURNAL OF VIROLOGY
(2023)
Article
Biochemistry & Molecular Biology
Emil P. T. Hertz, Ignacio Alonso-de Vega, Thomas Kruse, Yiqing Wang, Ivo A. Hendriks, Anna H. Bizard, Ania Eugui-Anta, Ronald T. Hay, Michael L. Nielsen, Jakob Nilsson, Ian D. Hickson, Niels Mailand
Summary: Hertz et al. use CRISPR screening to identify genetic vulnerabilities to inhibition of SUMOylation in human cells. They show that SUMO exerts its essential role in cell proliferation via NIP45- and BTRR-PICH-mediated DNA catenane resolution pathways. NIP45 mediates a TOP2-independent DNA catenane resolution process through its SUMO-like domains, promoting SUMOylation of specific factors including the SLX4 multi-nuclease complex, which contributes to catenane conversion into DSBs. Their findings establish the importance of SUMOylation in enabling resolution of toxic DNA catenanes via non-epistatic NIP45- and BTRR-PICH-dependent pathways to prevent mitotic failure.
NATURE STRUCTURAL & MOLECULAR BIOLOGY
(2023)
Article
Biochemistry & Molecular Biology
Sara Berrada, Elena Martinez-Balsalobre, Lise Larcher, Violette Azzoni, Nadia Vasquez, Melanie Da Costa, Sebastien Abel, Gilles Audoly, Lara Lee, Camille Montersino, Remy Castellano, Sebastien Combes, Camille Gelot, Raphael Ceccaldi, Jean-Hugues Guervilly, Jean Soulier, Christophe Lachaud
Summary: Fanconi anemia (FA) is a genetic disorder that affects DNA interstrand crosslink (ICL) repair. Researchers developed a new tool, click-melphalan, to investigate ICL repair and found that FANCD2 knockout cells are deficient in repairing click-melphalan-induced lesions.
NUCLEIC ACIDS RESEARCH
(2023)
Article
Cell Biology
Magda Liczmanska, Michael H. Tatham, Barbara Mojsa, Ania Eugui-Anta, Alejandro Rojas-Fernandez, Adel F. M. Ibrahim, Ronald T. Hay
Summary: The small ubiquitin-like modifier (SUMO) protease SENP6 disassembles SUMO chains from cellular substrate proteins. Proteins of the lamin family show increased SUMO modification after SENP6 depletion, accompanied by nuclear structural changes similar to laminopathies. Proximity-induced SUMO modification (PISM) directly targets lamin A/C for SUMO conjugation, recapitulating altered nuclear structure after SENP6 depletion. SENP6 activity protects the nucleus against hyperSUMOylation-induced laminopathy-like alterations.
Article
Biochemistry & Molecular Biology
Luke M. Simpson, Luke J. Fulcher, Gajanan Sathe, Abigail Brewer, Jin-Feng Zhao, Daniel R. Squair, Jennifer Crooks, Melanie Wightman, Nicola T. Wood, Robert Gourlay, Joby Varghese, Renata F. Soares, Gopal P. Sapkota
Summary: Reversible protein phosphorylation is crucial for controlling protein function and intracellular signaling. This study presents the development and application of the AdPhosphatase system, which allows targeted dephosphorylation of specific phospho-substrates. By using antigen-stabilized anti-GFP nanobody conjugated with Protein Phosphatase 1 or 2A catalytic subunits, the researchers successfully achieved specific dephosphorylation of two phospho-proteins, FAM83D and ULK1, with exquisite specificity. AdPhosphatase provides a new modality for potential drug discovery approaches by altering the phospho-status and function of target proteins.
CELL CHEMICAL BIOLOGY
(2023)
Article
Biochemistry & Molecular Biology
Luke M. Simpson, Lorraine Glennie, Abigail Brewer, Jin-Feng Zhao, Jennifer Crooks, Natalia Shpiro, Gopal P. Sapkota
Summary: This study investigates whether the subcellular context of a protein of interest (POI) affects the efficacy of PROTAC-mediated protein degradation. The results suggest that the subcellular localization of the POI can influence the effectiveness of PROTAC-induced POI degradation.
CELL CHEMICAL BIOLOGY
(2022)
Meeting Abstract
Hematology
Mathilde Poplineau, Leonard Herault, Adrien Mazuel, Nadine Platet, Shuhei Koide, Wakako Kuribayashi, Nadine Carbuccia, Lia N'Guyen, Julien Vernerey, Motohiko Oshima, Daniel Birnbaum, Christophe Lachaud, Atsushi Iwama, Estelle Duprez
EXPERIMENTAL HEMATOLOGY
(2021)