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Genetic Limitations to Athletic Performance
Published in Peter M. Tiidus, Rebecca E. K. MacPherson, Paul J. LeBlanc, Andrea R. Josse, The Routledge Handbook on Biochemistry of Exercise, 2020
A significant recent advancement in the field of gene therapy is the discovery and development of the CRISPR–Cas9 system for modifying DNA. The Cas9 protein assembles with a guide RNA, enabling DNA binding and cutting much more precisely than has previously been possible in humans. This opens up possibilities for gene therapy but also for gene doping. The CRISPR–Cas9 system can create either permanent or temporary changes to the genome causing insertion, deletion, or replacement of gene(s), single-base changes, or gene suppression or activation (36). New, potentially even better, genome editing tools are also emerging, such as prime editing, CRISPR–Cas3, and EvolvR (36). Whilst gene therapy is not currently mainstream, we move ever closer to that scenario.
Advances in Genome Editing
Published in Yashwant Pathak, Gene Delivery, 2022
CRISPR–Cas is a bacterial adaptive immune system that cleaves invading nucleic acids. CRISPRs were first discovered in E. coli in 1987 during an examination of genes involved in phosphate metabolism, and later in a variety of other bacterium species (Ishino et al., 2018). CRISPR-Cas systems are classified into two groups (class I and II), based on the structural variation of the Cas genes and the way they are organized. The classes are further subdivided into six types (type I–VI). Class I includes type I, III, and IV, and class II includes type II, V, and VI (Makarova and Koonin, 2015). Class 1 CRISPR–Cas systems have multiprotein effector complexes, whereas class II systems only have a single effector protein. Presently, six subtypes of the Type I system (Type I-A through Type I-F) have been found, each with a different number of Cas genes. All Type I systems, with the exception of cas1, cas2, and cas3, encode a Cascade-like complex. Cascade is the name given to the effector complex of type I systems (CRISPR-associated complex for antiviral defense) (Brouns et al., 2008). Cascade engages crRNA and locates the target, and the majority of variations are also in charge of crRNA processing. In rare circumstances, cascade can also help with spacer acquisition. Cas3 is a component of the Cascade complex in the Type I-A system, a protein with both helicase and DNase domains responsible for degrading the target (Huo et al., 2014; Gong et al., 2014). Cas1 and Cas2, the Cas9 hallmark protein, and occasionally a fourth protein are encoded by Type II CRISPR-Cas systems (Csn2 or Cas4) (Chylinski et al., 2014). Cas9 helps with adaptation, engages in crRNA processing, and cleaves the target DNA with the help of crRNA and tracrRNA (Karvelis et al., 2013). Type II systems are split into subtypes II-A, II-B, and II-C. In Type II-A and Type II-B, respectively, the csn2 and cas4 genes encoding adaptation proteins are present, whereas Type II-C lacks a fourth gene (Rath et al., 2015). The Type III CRISPR-Cas systems contain the hallmark protein Cas10 with unclear function (Dorsey et al., 2019).
Mechanisms underlying citrinin-induced toxicity via oxidative stress and apoptosis-mediated by mitochondrial-dependent pathway in SH-SY5Y cells
Published in Drug and Chemical Toxicology, 2023
Mahmoud Abudayyak, Ecem Fatma Karaman, Sibel Ozden
In the present study, Cas3, Cas9, and Bax were upregulated, while Bcl-2 was downregulated in CIT-treated SH-SY5Y cells, which also had a higher Bax/Bcl-2 ratio, compared to the solvent control group. It is known that the intrinsic pathway (or mitochondrial-dependent) is controlled by Bcl-2 family proteins (such as pro-apoptotic Bax, antiapoptotic Bcl-2, etc.) and the elevated Bax/Bcl-2 ratio serves as a sign of apoptosis (Cory and Adams 2002, Jeong et al.2009;; Bose et al.2016). The increased Bax/Bcl-2 ratio indicates mitochondrial dysfunction, resulting in the release of cytochrome-c. This sequential cascade subsequently induces several proteases, including Cas3, as the promoter of apoptosis, and Cas9 (Agarwal and Kaye, 2003, Abdel Wahab et al.2009). The activation of Cas3 and Cas9 leading to apoptosis has previously been shown in CIT-treated HL-60 and porcine kidney PK15 cells (Yu et al.2006, Klarić et al.2012). Meanwhile, Gayathri et al. (2015) reported that CIT induces apoptotic cell death through the mitochondria-mediated intrinsic pathway, not the extrinsic pathway. Our results suggest that CIT-induced neurotoxicity may be due to the induction of apoptosis via the mitochondrial-dependent pathway.
Potential of CRISPR/Cas system in the diagnosis of COVID-19 infection
Published in Expert Review of Molecular Diagnostics, 2021
V. Edwin Hillary, Savarimuthu Ignacimuthu, S. Antony Ceasar
Scientists started to explore modern molecular tools including genome-editing systems, especially the recently invented and noble prize winning clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated proteins (Cas) system for rapid diagnosis of SARS-CoV-2 [13,14]. CRISPR/Cas system has become a most popular genome-editing tool. This system uses a programmable protein that is attached to the targeted sequence with the help of guide RNA (gRNA) and creates double-stranded breaks (DSBs) [15]. There are many types of Cas proteins viz. Cas1, Cas2, Cas3, Cas6, Cas9, and Cas10 [16,17]. Among these, Cas9 protein is the efficient genome-editing system to target double-stranded DNA (dsDNA) or single-stranded DNA (ssDNA). Following Cas9 protein, the Cas12, and Cas13 proteins receive more attention in diagnosing viral diseases. Cas12 protein cleaves ssDNA and dsDNA, whereas Cas13 protein cleaves ssRNA; therefore, these proteins play an important role in the diagnosis of CoVs. In this review, we summarize the potential of CRISPR/Cas systems including Cas9, Cas12, and Cas13 proteins utilized for the diagnosis of SARS-CoV-2 infection.
Applications of the CRISPR-Cas system for infectious disease diagnostics
Published in Expert Review of Molecular Diagnostics, 2021
Peipei Li, Li Wang, Junning Yang, Li-Jun Di, Jingjing Li
Three types of class 1 CRISPR-Cas systems are type I, III and IV systems with different characteristic nucleases, including the enzymes Cas3, Cas10 and Cas8-like (csf1), respectively [65–67]. Type I systems are composed of Cas protein complexes and a crRNA molecule. The protospacer adjacent motif (PAM) is recognized by the Cascade complex, Cas6 or Cas5 mediates crRNA unwinding and binding with target DNA, the Cas7 backbone stabilizes the R-loop, then Cas3 cleaves the target DNA [68]. Type III systems comprise crRNA, Csm complexes (III-A systems), and Cmr complexes (III-B systems). Cas10 is the signature protein of type III systems, and Cas10 polymerase activity can generate cyclic oligo-(A) nucleotides (cOA), which bind with Csm6 and activate the RNase activity of Csm6, so they often cleave target RNA rather than DNA [69,70]. CRISPR-Cas type IV systems are found in plasmids, and the interference mechanisms are largely unknown [71].