Haploinsufficiency of Lipin3 leads to hypertriglyceridemia and obesity by disrupting the expression and nucleocytoplasmic localization of Lipin1

doi:10.1007/s11684-023-1003-0

2024, Vol. 18

Issue (1) : 180-191 https://doi.org/10.1007/s11684-023-1003-0

Haploinsufficiency of Lipin3 leads to hypertriglyceridemia and obesity by disrupting the expression and nucleocytoplasmic localization of Lipin1

Fang Wang¹, Yuxing Liu^1,², Yi Dong², Meifang Zhao², Hao Huang², Jieyuan Jin², Liangliang Fan^1,²(

), Rong Xiang^1,²(

)

¹. Department of Endocrinology, The Third Xiangya Hospital of Central South University, Changsha 410013, China
². Department of Cellular Biology, School of Life Sciences, Key Laboratory of Pediatric Rare Diseases, Ministry of Education, Central South University, Changsha 410013, China

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Abstract

Lipin proteins including Lipin 1–3 act as transcriptional co-activators and phosphatidic acid phosphohydrolase enzymes, which play crucial roles in lipid metabolism. However, little is known about the function of Lipin3 in triglyceride (TG) metabolism. Here, we identified a novel mutation (NM_001301860: p.1835A>T/p.D612V) of Lipin3 in a large family with hypertriglyceridemia (HTG) and obesity through whole-exome sequencing and Sanger sequencing. Functional studies revealed that the novel variant altered the half-life and stability of the Lipin3 protein. Hence, we generated Lipin3 heterozygous knockout (Lipin3-heKO) mice and cultured primary hepatocytes to explore the pathophysiological roles of Lipin3 in TG metabolism. We found that Lipin3-heKO mice exhibited obvious obesity, HTG, and non-alcoholic fatty liver disorder. Mechanistic study demonstrated that the haploinsufficiency of Lipin3 in primary hepatocytes may induce the overexpression and abnormal distribution of Lipin1 in cytosol and nucleoplasm. The increased expression of Lipin1 in cytosol may contribute to TG anabolism, and the decreased Lipin1 in nucleoplasm can reduce PGC1α, further leading to mitochondrial dysfunction and reduced TG catabolism. Our study suggested that Lipin3 was a novel disease-causing gene inducing obesity and HTG. We also established a relationship between Lipin3 and mitochondrial dysfunction.

Keywords Lipin3 Lipin1 hypertriglyceridemia obesity mitochondrial dysfunction

Corresponding Author(s): Liangliang Fan,Rong Xiang

About author:

Li Liu and Yanqing Liu contributed equally to this work.

Just Accepted Date: 31 August 2023 Online First Date: 10 October 2023 Issue Date: 22 April 2024

Cite this article:

Fang Wang,Yuxing Liu,Yi Dong, et al. Haploinsufficiency of Lipin3 leads to hypertriglyceridemia and obesity by disrupting the expression and nucleocytoplasmic localization of Lipin1[J]. Front. Med., 2024, 18(1): 180-191.

URL:

https://academic.hep.com.cn/fmd/EN/10.1007/s11684-023-1003-0
https://academic.hep.com.cn/fmd/EN/Y2024/V18/I1/180

Fig.1 Whole-exome sequencing identified a novel mutation of Lipin3 in a family with obesity and hypertriglyceridemia. (A) Pedigree of the family. Black circles/squares are affected, white circles/squares are unaffected. Arrow indicates the proband. (B) Overlapping filter strategy. Asterisks denote the remaining variants for further analysis that are present in two affected members (II-8 and III-3) but not in the normal control (III-2). (C) Schematic of the filter strategies used in our study. (D) Sanger DNA sequencing chromatogram demonstrates the heterozygosity for a Lipin3 missense mutation (p.1835A>T/p.D612V).

Tab.1 Characteristics and serum lipid levels of the family members

Fig.2 The novel mutation (p.D612V) may disrupt the stability of Lipin3 protein and lead to triglyceride accumulation. (A) Conservation analysis of the D612 amino acid was conducted with ConSurf Server software. (B) Structure prediction of the mutant protein. The wild-type Lipin3 (Lipin3-WT) protein structure and the p.D612V mutant Lipin3 (Lipin3-p.D612V) protein structure were predicted using SWISS-MODEL online software. (C) WB detected the expression of total Lipin3 and transfected Lipin3 (Flag) in the WT and Lipin3-mutated groups. (D,E) WB detected the degradation rate of transfected Lipin3 (Flag) in mutated and WT groups.

Fig.3 Haploinsufficiency of Lipin3 leads to obesity and hypertriglyceridemia in mice. (A) Sanger sequencing and WB confirmed that Lipin3-KO mice were generated successfully. (B,C) Body shape and weight of WT mice (n = 5) and Lipin3-heKO mice (n = 5). Weight of epididymal fat tissue (D) and liver tissue (E) in WT mice (n = 5) and Lipin3-heKO mice (n = 5). (F) Peripheral blood TG levels of WT mice (n = 5) and Lipin3-heKO mice (n = 5). (G) HE staining analysis showing the size of fat cells in WT mice and Lipin3-heKO mice. (H) Oil red O staining showing the lipid accumulation in liver tissues of WT mice and Lipin3-heKO mice. (I) TG levels in WT and Lipin3-heKO mouse primary cultured hepatocytes.

Fig.4 Lipin3-heKO mice exhibited a severe overweight phenotype underlie high-diet feeding. (A,B) Body shape and weight of HFD-WT mice (n = 5) and HFD-Lipin3-heKO mice (n = 5). Weight of epididymal fat tissue (C) and liver tissue (D) in HFD-WT mice (n = 5) and HFD-Lipin3-heKO mice (n = 5). (E) Peripheral blood TG levels of HFD-WT mice (n = 5) and HFD-Lipin3-heKO mice (n = 5). (F) HE staining analysis showing the size of fat cells in HFD-WT mice and HFD-Lipin3-heKO mice. (G) HE staining and analysis of NAS score for the hepatic lipid accumulation in HFD-WT mice and HFD-Lipin3-heKO mice.

Fig.5 Haploinsufficiency of Lipin3 may disrupt the nucleocytoplasmic localization of Lipin1 and induce mitochondrial dysfunction. (A) WB analysis revealed the expression of Lipin3 in the primary hepatocytes of WT and Lipin3-heKO. (B) WB analysis detected the levels of Lipin3, GAPDH, and histone in the cytoplasm and nucleus of primary cultured hepatocytes from WT and Lipin3-heKO mice. (C) WB analysis revealed the expression of PGC1α, PPARα, and cleaved PPARα in the primary hepatocytes of WT and Lipin3-heKO. (D) WB detected the expressions of mitochondrial electronic respiratory chain-related proteins. (E) JC-1 staining in WT and Lipin3-heKO mouse primary-cultured hepatocytes. (F) TEM analysis shows the mitochondria structures in primary cultured hepatocytes of WT and Lipin3-heKO mice. (G) Expression of Mfn2, Opa1, and Fis1 in WT and Lipin3-heKO mouse primary-cultured hepatocytes. ATP (H) and ROS (I) levels in WT and Lipin3-heKO mouse primary cultured hepatocytes.

Fig.6 Potential mechanism of how haploinsufficiency of Lipin3 induced TG accumulation.

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