An efficient miRNA knockout approach using CRISPR-Cas9 in Xenopus

Godden, Alice M.; Antonaci, Marco; Ward, Nicole J.; Lee, Michael van der; Abu-Daya, Anita; Guille, Matthew; Wheeler, Grant N.

doi:10.1016/j.ydbio.2021.12.015

Cited by 8 publications

(6 citation statements)

References 68 publications

(65 reference statements)

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“…Another way to overcome this issue is to use more than one Cas protein to target different parts of the interested gene. For instance, Godden et al [ 87 ] showed that when two sgRNAs are used together, the targeted miRNA gene loses all of its functions in embryos. Additionally, the discovery of new and more Cas nucleases with broader PAM recognition sequences gives more opportunities to design gRNAs in different parts of miRNA sequences [ 181 ].…”

Section: Challenges Of Targeting Mirnas By Crispr/cas and Its Strateg...mentioning

confidence: 99%

“…Interestingly, an increasing number of studies are selecting lentivirus as their vector of choice. However, using a viral vector in vivo has a number of drawbacks, including insertional restriction, immune response, and size capacity (Table 6 [ 78 , 87 , 88 , 119 , 215 – 222 ]). Moreover, the risks of off-targeting and further mutation rise with prolonged expansion following insertion [ 223 ].…”

Section: Challenges Of Targeting Mirnas By Crispr/cas and Its Strateg...mentioning

confidence: 99%

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Targeting miRNA by CRISPR/Cas in cancer: advantages and challenges

et al. 2023

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Clustered regulatory interspaced short palindromic repeats (CRISPR) has changed biomedical research and provided entirely new models to analyze every aspect of biomedical sciences during the last decade. In the study of cancer, the CRISPR/CRISPR-associated protein (Cas) system opens new avenues into issues that were once unknown in our knowledge of the noncoding genome, tumor heterogeneity, and precision medicines. CRISPR/Cas-based gene-editing technology now allows for the precise and permanent targeting of mutations and provides an opportunity to target small non-coding RNAs such as microRNAs (miRNAs). However, the development of effective and safe cancer gene editing therapy is highly dependent on proper design to be innocuous to normal cells and prevent introducing other abnormalities. This study aims to highlight the cutting-edge approaches in cancer-gene editing therapy based on the CRISPR/Cas technology to target miRNAs in cancer therapy. Furthermore, we highlight the potential challenges in CRISPR/Cas-mediated miRNA gene editing and offer advanced strategies to overcome them.

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Section: Challenges Of Targeting Mirnas By Crispr/cas and Its Strateg...mentioning

confidence: 99%

Section: Challenges Of Targeting Mirnas By Crispr/cas and Its Strateg...mentioning

confidence: 99%

Targeting miRNA by CRISPR/Cas in cancer: advantages and challenges

et al. 2023

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“…Introduction of these sgRNAs plus Cas9 into the embryo leads to deletion of the whole miRNA pri-RNA sequence. Using this method, we have knocked out miR-219 and miR-196a, showing clear NC phenotypes, including craniofacial and pigment abnormalities [ 39 ]. The observed phenotype for miR-219 could be due to the direct down-regulation of Pdgfra and Sox6 which have been shown to be targets of miR-219 in oligodendrocyte differentiation and myelination [ 40 ].…”

Section: The Role Of Mirnas In Nc Developmentmentioning

confidence: 99%

MicroRNAs in neural crest development and neurocristopathies

Antonaci

Wheeler

2022

Biochemical Society Transactions

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The neural crest (NC) is a vertebrate-specific migratory population of multipotent stem cells that originate during late gastrulation in the region between the neural and non-neural ectoderm. This population of cells give rise to a range of derivatives, such as melanocytes, neurons, chondrocytes, chromaffin cells, and osteoblasts. Because of this, failure of NC development can cause a variety of pathologies, often syndromic, that are globally called neurocristopathies. Many genes are known to be involved in NC development, but not all of them have been identified. In recent years, attention has moved from protein-coding genes to non-coding genes, such as microRNAs (miRNA). There is increasing evidence that these non-coding RNAs are playing roles during embryogenesis by regulating the expression of protein-coding genes. In this review, we give an introduction to miRNAs in general and then focus on some miRNAs that may be involved in NC development and neurocristopathies. This new direction of research will give geneticists, clinicians, and molecular biologists more tools to help patients affected by neurocristopathies, as well as broadening our understanding of NC biology.

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“…Over 70% of human genes have at least one zebrafish orthologue [ 36 ], with the Xenopus genome including orthologs of ~80% of human disease genes [ 37 ]. More recently, methods to increase the genome editing efficiency of the Clustered Regularly Interspaced Short Palindromic Repeat– (CRISPR–) Cas9 system in zebrafish [ 38 , 39 , 40 , 41 , 42 , 43 ] and Xenopus [ 44 , 45 , 46 , 47 , 48 ] have led to new human disease models.…”

Section: Aquatic Freshwater Vertebrate Animal Model Advantagesmentioning

confidence: 99%

“…However, it is less understood whether resistance is due to mutations that cause changes in the pharmacokinetics and pharmacodynamics of the therapeutic molecules or inherent differences in the pathophysiology of the seizure. The extensive genetic toolkit available in aquatic freshwater vertebrates permits modeling of knockdown [ 101 , 102 , 103 , 104 , 105 , 106 , 107 ], knockout [ 39 , 44 , 46 , 48 , 108 , 109 , 110 , 111 , 112 , 113 , 114 , 115 , 116 ], ectopic, and overexpression [ 38 , 117 , 118 , 119 , 120 , 121 ] of seizure-associated genes in combination with metabolic enzymes, transporters, and other proteins required for absorption, distribution, degradation, and excretion of therapeutics. A unique aspect of Xenopus is that their large eggs and embryos can be genetically modified by injecting the embryo on only one side at the two-cell stage, providing an internal control [ 120 ].…”

Section: Aquatic Freshwater Vertebrate Animal Model Advantagesmentioning

confidence: 99%

Aquatic Freshwater Vertebrate Models of Epilepsy Pathology: Past Discoveries and Future Directions for Therapeutic Discovery

Williams¹,

Mruk²

2022

IJMS

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Epilepsy is an international public health concern that greatly affects patients’ health and lifestyle. About 30% of patients do not respond to available therapies, making new research models important for further drug discovery. Aquatic vertebrates present a promising avenue for improved seizure drug screening and discovery. Zebrafish (Danio rerio) and African clawed frogs (Xenopus laevis and tropicalis) are increasing in popularity for seizure research due to their cost-effective housing and rearing, similar genome to humans, ease of genetic manipulation, and simplicity of drug dosing. These organisms have demonstrated utility in a variety of seizure-induction models including chemical and genetic methods. Past studies with these methods have produced promising data and generated questions for further applications of these models to promote discovery of drug-resistant seizure pathology and lead to effective treatments for these patients.

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An efficient miRNA knockout approach using CRISPR-Cas9 in Xenopus

Cited by 8 publications

References 68 publications

Targeting miRNA by CRISPR/Cas in cancer: advantages and challenges

Targeting miRNA by CRISPR/Cas in cancer: advantages and challenges

MicroRNAs in neural crest development and neurocristopathies

Aquatic Freshwater Vertebrate Models of Epilepsy Pathology: Past Discoveries and Future Directions for Therapeutic Discovery

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