CRISPR/Cas9 ADCY7 Knockout Stimulates the Insulin Secretion Pathway Leading to Excessive Insulin Secretion.
Front Endocrinol (Lausanne)
; 12: 657873, 2021.
Article
em En
| MEDLINE
| ID: mdl-34177802
ABSTRACT
Aim:
Despite the enormous efforts to understand Congenital hyperinsulinism (CHI), up to 50% of the patients are genetically unexplained. We aimed to functionally characterize a novel candidate gene in CHI. Patient A 4-month-old boy presented severe hyperinsulinemic hypoglycemia. A routine CHI genetic panel was negative.Methods:
A trio-based whole-exome sequencing (WES) was performed. Gene knockout in the RIN-m cell line was established by CRISPR/Cas9. Gene expression was performed using real-time PCR.Results:
Hyperinsulinemic hypoglycemia with diffuse beta-cell involvement was demonstrated in the patient, who was diazoxide-responsive. By WES, compound heterozygous variants were identified in the adenylyl cyclase 7, ADCY7 gene p.(Asp439Glu) and p.(Gly1045Arg). ADCY7 is calcium-sensitive, expressed in beta-cells and converts ATP to cAMP. The variants located in the cytoplasmic domains C1 and C2 in a highly conserved and functional amino acid region. RIN-m(-/-Adcy7) cells showed a significant increase in insulin secretion reaching 54% at low, and 49% at high glucose concentrations, compared to wild-type. In genetic expression analysis Adcy7 loss of function led to a 34.1-fold to 362.8-fold increase in mRNA levels of the insulin regulator genes Ins1 and Ins2 (p ≤ 0.0002), as well as increased glucose uptake and sensing indicated by higher mRNA levels of Scl2a2 and Gck via upregulation of Pdx1, and Foxa2 leading to the activation of the glucose stimulated-insulin secretion (GSIS) pathway.Conclusion:
This study identified a novel candidate gene, ADCY7, to cause CHI via activation of the GSIS pathway.Palavras-chave
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Adenilil Ciclases
/
Hiperinsulinismo Congênito
/
Células Secretoras de Insulina
/
Insulina
Tipo de estudo:
Prognostic_studies
Limite:
Animals
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Child, preschool
/
Humans
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Male
Idioma:
En
Ano de publicação:
2021
Tipo de documento:
Article