A case report with functional characterization of a HNF1B mutation (p.Leu168Pro) causing MODY5

Kei Yoshida; Yuichi Mushimoto; Kanako Tanase-Nakao; Kazuhisa Akiba; Kanako Ishii; Tatsuhiko Urakami; Shigetaka Sugihara; Toru Kikuchi; Maki Fukami; Satoshi Narumi; Japanese Study Group of Insulin Therapy for Childhood and Adolescent Diabetes (JSGIT)

doi:10.1297/cpe.30.179

A case report with functional characterization of a HNF1B mutation (p.Leu168Pro) causing MODY5

Clin Pediatr Endocrinol. 2021;30(4):179-185. doi: 10.1297/cpe.30.179. Epub 2021 Oct 1.

Affiliations

¹ Department of Molecular Endocrinology, National Research Institute for Child Health and Development, Tokyo, Japan.
² Department of Pediatrics and Child Health, Nihon University School of Medicine, Tokyo, Japan.
³ Department of Pediatrics, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan.
⁴ Department of Pediatrics, Tokyo Women's Medical University Medical Center East, Tokyo, Japan.
⁵ Department of Pediatrics, Saitama Medical University, Saitama, Japan.

Abstract

We previously performed next-generation sequencing-based genetic screening in patients with autoantibody-negative type 1 diabetes, and identified the p.Leu168Pro mutation in HNF1B. Here,we report the clinical course of the patient and the results of functional characterization of this mutation. The proband had bilateral renal hypodysplasia and developed insulin-dependent diabetes during childhood. The pathogenicity of Leu168Pro-HNF1B was evaluated with three-dimensional structure modeling, Western blotting, immunofluorescence analysis and luciferase reporter assays using human embryonic kidney 293 cells. Three-dimensional structure modeling predicted that the Leu168 residue is buried in the DNA-binding Pit-Oct-Unc-specific (POU_S) domain and forms a hydrophobic core. Western blotting showed that the protein expression level of Leu168Pro-HNF1B was lower than that of wild-type (WT) HNF1B. Immunofluorescence staining showed that both WT- and Leu168Pro-HNF1B were normally localized in the nucleus. The cells transfected with WT-HNF1B exhibited 5-fold higher luciferase reporter activity than cells transfected with an empty vector. The luciferase activities were comparable between WT-HNF1B/Leu168Pro-HNF1B and WT-HNF1B/empty vector co-transfection. In conclusion, Leu168Pro is a protein-destabilizing HNF1B mutation, and the destabilization is likely due to the structural changes involving the hydrophobic core of POU_S. The disease-causing Leu168Pro HNF1B mutation is a loss-of-function mutation without a dominant-negative effect.

Keywords: HNF1B; MODY5; genetics; multicystic dysplastic kidney; mutation.