Molecular Characterization, Expression Analysis, and CRISPR/Cas9 Mediated Gene Disruption of Myogenic Regulatory Factor 4 (MRF4) in Nile Tilapia
Myogenic regulator factors (MRFs) are essential for skeletal muscle development in vertebrates, including fish. This study aimed to characterize the role of <i>myogenic regulatory factor 4</i> (<i>MRF4</i>) in muscle development in Nile tilapia by cloning <i>NT-MRF4<...
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| Main Authors: | , , , , |
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| Format: | Article |
| Language: | English |
| Published: |
MDPI AG
2024-12-01
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| Series: | Current Issues in Molecular Biology |
| Subjects: | |
| Online Access: | https://www.mdpi.com/1467-3045/46/12/820 |
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| Summary: | Myogenic regulator factors (MRFs) are essential for skeletal muscle development in vertebrates, including fish. This study aimed to characterize the role of <i>myogenic regulatory factor 4</i> (<i>MRF4</i>) in muscle development in Nile tilapia by cloning <i>NT-MRF4</i> from muscle tissues. To explore the function of <i>NT-MRF4</i>, CRISPR/Cas9 gene editing was employed. The <i>NT-MRF4</i> cDNA was 1146 bp long and had encoded 225 amino acids, featuring a myogenic basic domain, a helix-loop-helix domain, and a nuclear localization signal. <i>NT</i>-MRF4 mRNA was exclusively expressed in adult muscle tissues, with expression also observed during embryonic and larval stages. Food-deprived Nile tilapia exhibited significantly lower <i>NT-MRF4</i> mRNA levels than the controls while re-feeding markedly increased expression. The CRISPR/Cas9 gene editing of <i>NT-MRF4</i> successfully generated two types of gene disruption, leading to a frame-shift mutation in the NT-MRF4 protein. Expression analysis of <i>MRF</i> and <i>MEF2</i> genes in gene-edited (GE) Nile tilapia revealed that <i>MyoG</i> expressions nearly doubled compared to wild-type (WT) fish, suggesting that <i>MyoG</i> compensates for the loss of MRF4 function. Additionally, <i>MEF2b</i>, <i>MEF2d</i>, and <i>MEF2a</i> expressions significantly increased in GE Nile tilapia, supporting continued muscle development. Overall, these findings suggest that <i>NT-MRF4</i> regulates muscle development, while <i>MyoG</i> may compensate for its inactivation to sustain normal muscle growth. |
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| ISSN: | 1467-3037 1467-3045 |