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Pathogenicity evaluation and splicing correction of COL4A5 splice mutation in X-linked Alport syndrome / 中华肾脏病杂志
Chinese Journal of Nephrology ; (12): 872-880, 2021.
Article in Chinese | WPRIM | ID: wpr-911907
ABSTRACT

Objective:

To analyze the splicing mutation site of COL4A5 gene in a family with X-linked dominant Alport syndrome and explore the possibility of exon specific U1 small nuclear RNA (snRNA) gene therapy.

Methods:

The clinical data of the proband and family members of Alport syndrome were collected, and the gene mutations in the whole exon of a series of nephropathy genes in the proband were detected by high-throughput sequencing. The splice site changes and pathogenicity caused by COL4A5 c.546+5G>A mutation were analyzed by online software. Minigene experiment was used to verify and analyze the effect of COL4A5 gene mutation site c.546+5G>A in the proband of Alport syndrome family, and transient transfection and introduction of modified U1 snRNA to correct splicing mutation.

Results:

The results of gene sequencing showed that there was a hemizygous variation of COL4A5 gene in the proband and his half brother, and the variation site was c.546+5G>A. The results of online software for analyzing the pathogenicity of splice variation showed that the original donor splicing site could not be detected after mutation, suggesting that there was a great possibility of affecting splicing. The abnormal splicing mode of COL4A5 gene with c.546+5G>A mutation—deletion of exon 9 was verified by hybridized small gene detection. The abnormal splicing mutation could be partially corrected by the modified U1 snRNA. The correction ratios of ExSpeU1 (MT), ExSpeU1(E9+1), ExSpeU1(E9+9) and ExSpeU1(E9+11) to exon 9 deletion caused by c.546+5G>A were 0, 43.81%, 52.09% and 48.12%, respectively.

Conclusions:

The pathogenicity of the new splicing mutation of COL4A5 is verified, and the modified U1 snRNA can partially correct the abnormal splicing.

Full text: Available Index: WPRIM (Western Pacific) Language: Chinese Journal: Chinese Journal of Nephrology Year: 2021 Type: Article

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Full text: Available Index: WPRIM (Western Pacific) Language: Chinese Journal: Chinese Journal of Nephrology Year: 2021 Type: Article