CRISPR Journal最新文献_第7页

CRISPRi-Mediated Treatment of Dominant Rhodopsin-Associated Retinitis Pigmentosa. CRISPRi-Mediated Treatment of Dominant Rhodopsin-Associated Retinitis Pigmentosa.

IF 3.7 4区生物学

CRISPR Journal Pub Date : 2023-12-01 DOI: 10.1089/crispr.2023.0039

Erin R Burnight, Luke A Wiley, Nathaniel K Mullin, Malavika K Adur, Mallory J Lang, Cathryn M Cranston, Chunhua Jiao, Stephen R Russell, Elliot H Sohn, Ian C Han, Jason W Ross, Edwin M Stone, Robert F Mullins, Budd A Tucker

{"title":"CRISPRi-Mediated Treatment of Dominant Rhodopsin-Associated Retinitis Pigmentosa.","authors":"Erin R Burnight, Luke A Wiley, Nathaniel K Mullin, Malavika K Adur, Mallory J Lang, Cathryn M Cranston, Chunhua Jiao, Stephen R Russell, Elliot H Sohn, Ian C Han, Jason W Ross, Edwin M Stone, Robert F Mullins, Budd A Tucker","doi":"10.1089/crispr.2023.0039","DOIUrl":"10.1089/crispr.2023.0039","url":null,"abstract":"Rhodopsin (RHO) mutations such as Pro23His are the leading cause of dominantly inherited retinitis pigmentosa in North America. As with other dominant retinal dystrophies, these mutations lead to production of a toxic protein product, and treatment will require knockdown of the mutant allele. The purpose of this study was to develop a CRISPR-Cas9-mediated transcriptional repression strategy using catalytically inactive Staphylococcus aureus Cas9 (dCas9) fused to the Krüppel-associated box (KRAB) transcriptional repressor domain. Using a reporter construct carrying green fluorescent protein (GFP) cloned downstream of the RHO promoter fragment (nucleotides -1403 to +73), we demonstrate a ∼74-84% reduction in RHO promoter activity in RHOpCRISPRi-treated versus plasmid-only controls. After subretinal transduction of human retinal explants and transgenic Pro23His mutant pigs, significant knockdown of rhodopsin protein was achieved. Suppression of mutant transgene in vivo was associated with a reduction in endoplasmic reticulum (ER) stress and apoptosis markers and preservation of photoreceptor cell layer thickness.","PeriodicalId":54232,"journal":{"name":"CRISPR Journal","volume":"6 6","pages":"502-513"},"PeriodicalIF":3.7,"publicationDate":"2023-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11304754/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"138807791","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

A Fluorescent Reporter Mouse for In Vivo Assessment of Genome Editing with Diverse Cas Nucleases and Prime Editors. 一种荧光报告小鼠，用于在体内评估使用多种 Cas 核酸酶和主编辑器进行基因组编辑的效果。

IF 3.7 4区生物学

CRISPR Journal Pub Date : 2023-12-01 DOI: 10.1089/crispr.2023.0048

Zexiang Chen, Suet-Yan Kwan, Aamir Mir, Max Hazeltine, Minwook Shin, Shun-Qing Liang, Io Long Chan, Karen Kelly, Krishna S Ghanta, Nicholas Gaston, Yueying Cao, Jun Xie, Guangping Gao, Wen Xue, Erik J Sontheimer, Jonathan K Watts

引用次数: 0

First Drug Approval Rises the CRISPR Tide. 首次药物批准掀起 CRISPR 浪潮。

IF 3.7 4区生物学

CRISPR Journal Pub Date : 2023-12-01 Epub Date: 2023-12-05 DOI: 10.1089/crispr.2023.29168.editorial

Rodolphe Barrangou

引用次数: 0

Correction to: APOBEC Reporter Systems for Evaluating diNucleotide Editing Levels by Rieffer et al. The CRISPR Journal, 2023;6(5):430-446; DOI: 10.1089/crispr.2023.0027. Rieffer等人对《评估双核核苷酸编辑水平的APOBEC报告系统》的更正。CRISPR期刊，2023；6（5）：430-446；DOI:10.1089/crispr.2023.0027。

IF 3.7 4区生物学

CRISPR Journal Pub Date : 2023-12-01 Epub Date: 2023-11-08 DOI: 10.1089/crispr.2023.0027.correx

引用次数: 0

Modified Cas9-Guided Oxford Nanopore Technology Sequencing Uncovers Single and Multiple Transgene Insertion Sites in a Zebrafish Melanoma Model. 改良 Cas9 引导的牛津纳米孔技术测序发现斑马鱼黑色素瘤模型中的单个和多个转基因插入位点。

IF 3.7 4区生物学

CRISPR Journal Pub Date : 2023-12-01 DOI: 10.1089/crispr.2023.0062

Raffaella De Paolo, Uday Munagala, Francesco Cucco, Samanta Sarti, Letizia Pitto, Filippo Martignano, Silvestro G Conticello, Laura Poliseno

引用次数: 0

A PCR-Induced Mutagenesis-Restriction Fragment Length Polymorphism Method for the Detection of CRISPR-Induced Indels. 用于检测 CRISPR 诱导的 Indels 的 PCR 诱导突变-限制性片段长度多态性方法。

IF 3.7 4区生物学

CRISPR Journal Pub Date : 2023-12-01 Epub Date: 2023-12-05 DOI: 10.1089/crispr.2023.0047

Lydia Angelopoulou, Electra Stylianopoulou, Konstantinos Tegopoulos, Ioanna Farmakioti, Maria Grigoriou, George Skavdis

引用次数: 0

Differential Divalent Metal Binding by SpyCas9's RuvC Active Site Contributes to Nonspecific DNA Cleavage. SpyCas9的RuvC活性位点与二价金属的不同结合导致了非特异性DNA裂解。

IF 3.7 4区生物学

CRISPR Journal Pub Date : 2023-12-01 DOI: 10.1089/crispr.2023.0022

Sydney N Newsom, Duen-Shian Wang, Saadi Rostami, Isabelle Schuster, Hari Priya Parameshwaran, Yadin G Joseph, Peter Z Qin, Jin Liu, Rakhi Rajan

{"title":"Differential Divalent Metal Binding by SpyCas9's RuvC Active Site Contributes to Nonspecific DNA Cleavage.","authors":"Sydney N Newsom, Duen-Shian Wang, Saadi Rostami, Isabelle Schuster, Hari Priya Parameshwaran, Yadin G Joseph, Peter Z Qin, Jin Liu, Rakhi Rajan","doi":"10.1089/crispr.2023.0022","DOIUrl":"10.1089/crispr.2023.0022","url":null,"abstract":"To protect against mobile genetic elements (MGEs), some bacteria and archaea have clustered regularly interspaced short palindromic repeats-CRISPR associated (CRISPR-Cas) adaptive immune systems. CRISPR RNAs (crRNAs) bound to Cas nucleases hybridize to MGEs based on sequence complementarity to guide the nucleases to cleave the MGEs. This programmable DNA cleavage has been harnessed for gene editing. Safety concerns include off-target and guide RNA (gRNA)-free DNA cleavages, both of which are observed in the Cas nuclease commonly used for gene editing, Streptococcus pyogenes Cas9 (SpyCas9). We developed a SpyCas9 variant (SpyCas9H982A) devoid of gRNA-free DNA cleavage activity that is more selective for on-target cleavage. The H982A substitution in the metal-dependent RuvC active site reduces Mn2+-dependent gRNA-free DNA cleavage by ∼167-fold. Mechanistic molecular dynamics analysis shows that Mn2+, but not Mg2+, produces a gRNA-free DNA cleavage competent state that is disrupted by the H982A substitution. Our study demonstrates the feasibility of modulating cation:protein interactions to engineer safer gene editing tools.","PeriodicalId":54232,"journal":{"name":"CRISPR Journal","volume":"6 6","pages":"527-542"},"PeriodicalIF":3.7,"publicationDate":"2023-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10753984/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"138807848","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Enhancing Precision and Efficiency of Cas9-Mediated Knockin Through Combinatorial Fusions of DNA Repair Proteins. 通过DNA修复蛋白的组合融合提高Cas9介导的Knockin的准确性和效率。

IF 3.7 4区生物学

CRISPR Journal Pub Date : 2023-10-01 Epub Date: 2023-09-15 DOI: 10.1089/crispr.2023.0036

Ryan R Richardson, Marilyn Steyert, Saovleak N Khim, Garrett W Crutcher, Cheryl Brandenburg, Colin D Robertson, Andrea J Romanowski, Jeffrey Inen, Bekir Altas, Alexandros Poulopoulos

{"title":"Enhancing Precision and Efficiency of Cas9-Mediated Knockin Through Combinatorial Fusions of DNA Repair Proteins.","authors":"Ryan R Richardson, Marilyn Steyert, Saovleak N Khim, Garrett W Crutcher, Cheryl Brandenburg, Colin D Robertson, Andrea J Romanowski, Jeffrey Inen, Bekir Altas, Alexandros Poulopoulos","doi":"10.1089/crispr.2023.0036","DOIUrl":"10.1089/crispr.2023.0036","url":null,"abstract":"Cas9 targets genomic loci with high specificity. For knockin with double-strand break repair, however, Cas9 often leads to unintended on-target knockout rather than intended edits. This imprecision is a barrier for direct in vivo editing where clonal selection is not feasible. In this study, we demonstrate a high-throughput workflow to comparatively assess on-target efficiency and precision of editing outcomes. Using this workflow, we screened combinations of donor DNA and Cas9 variants, as well as fusions to DNA repair proteins. This yielded novel high-performance double-strand break repair editing agents and combinatorial optimizations, yielding increases in knockin efficiency and precision. Cas9-RC, a novel fusion Cas9 flanked by eRad18 and CtIP[HE], increased knockin performance in vitro and in vivo in the developing mouse brain. Continued comparative assessment of editing efficiency and precision with this framework will further the development of high-performance editing agents for in vivo knockin and future genome therapeutics.","PeriodicalId":54232,"journal":{"name":"CRISPR Journal","volume":" ","pages":"447-461"},"PeriodicalIF":3.7,"publicationDate":"2023-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10611978/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10609434","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

APOBEC Reporter Systems for Evaluating diNucleotide Editing Levels. 评估双核核苷酸编辑水平的APOBEC报告系统。

IF 3.7 4区生物学

CRISPR Journal Pub Date : 2023-10-01 Epub Date: 2023-09-06 DOI: 10.1089/crispr.2023.0027

Amanda E Rieffer, Yanjun Chen, Daniel J Salamango, Sofia N Moraes, Reuben S Harris

{"title":"APOBEC Reporter Systems for Evaluating diNucleotide Editing Levels.","authors":"Amanda E Rieffer, Yanjun Chen, Daniel J Salamango, Sofia N Moraes, Reuben S Harris","doi":"10.1089/crispr.2023.0027","DOIUrl":"10.1089/crispr.2023.0027","url":null,"abstract":"Precision genome editing has become a reality with the discovery of base editors. Cytosine base editor (CBE) technologies are improving rapidly but are mostly optimized for TC dinucleotide targets. Here, we report the development and implementation of APOBEC Reporter Systems for Evaluating diNucleotide Editing Levels (ARSENEL) in living cells. The ARSENEL panel is comprised of four constructs that quantitatively report editing of each of the four dinucleotide motifs (AC/CC/GC/TC) through real-time accumulation of eGFP fluorescence. Editing rates of APOBEC3Bctd and AIDΔC CBEs reflect established mechanistic preferences with intrinsic biases to TC and GC, respectively. Twelve different (new and established) base editors are tested here using this system with a full-length APOBEC3B CBE showing the greatest on-target TC specificity and an APOBEC3A construct showing the highest editing efficiency. In addition, ARSENEL enables real-time assessment of natural and synthetic APOBEC inhibitors with the most potent to-date being the large subunit of the Epstein-Barr virus ribonucleotide reductase. These reporters have the potential to play important roles in research and development as precision genome engineering technologies progress toward achieving maximal specificity and efficiency.","PeriodicalId":54232,"journal":{"name":"CRISPR Journal","volume":" ","pages":"430-446"},"PeriodicalIF":3.7,"publicationDate":"2023-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10611974/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10168892","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Special Issue: CRISPR Trials. 特刊：CRISPR试验。

IF 3.7 4区生物学

CRISPR Journal Pub Date : 2023-10-01 DOI: 10.1089/crispr.2023.29166.cfp

Fyodor Urnov

引用次数: 0