Journal of Guangxi Normal University(Natural Science Edition) ›› 2023, Vol. 41 ›› Issue (3): 163-170.doi: 10.16088/j.issn.1001-6600.2022040602

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Construction of AC16 Human Cardiomyocyte Cell Line with Mutant LMNA Gene by CRISPR/Cas9

ZOU Lei1,2, XING Bing1,2, YANG Liu1,2*   

  1. 1. College of Life Science, Guangxi Normal University, Guilin Guangxi 541006, China;
    2. Key Laboratory of Ecology of Rare and Endangered Species and Environmental Protection (Guangxi Normal University), Ministry of Education, Guilin Guangxi 541006, China
  • Received:2022-04-06 Revised:2022-07-27 Online:2023-05-25 Published:2023-06-01

Abstract: Mutations in LMNA (LaminA/C) gene trigger abnormal expression of nucleofibrillar proteins and their linked or interacting proteins, causing a series of physiological responses in the body and leading to the development of nucleofibrillar protein disease, however, the specific pathogenic mechanism of the disease is still unclear. In this paper, a cell line carrying LMNA mutation Q517X is successfully constructed by using CRISPR/Cas9 technology to select pathogenic mutation loci in accordance with the sgRNA screening principle, and examined the RNA expression levels and protein expression levels of LaminB1, PCNA, P53, PKCα and other genes linked to or interacting with LaminA/C in the mutant cell line and the nuclear membrane of the mutant cells. It was found that Q517X mutant cells were not mutated, and the mRNA expression of LaminB1, P53 and PCNA genes were significantly reduced by about 70% in Q517X mutant cells, and the expression of LaminA/C, PKCα, LaminB1 and P53 proteins were significantly reduced by 75%, 60% and 80% respectively in Q517X mutant cells. The results showed that the Q517X mutation altered the expression of LMNA-related genes in human cardiomyocytes and thus affected the normal cytoskeletal structure and function of the nucleus.

Key words: LMNA, LaminA, Laminopathies, CRISPR/Cas9, gene editing

CLC Number:  Q78;R542.2
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