Haploinsufficiency of the Insulin Receptor in the Presence of a Splice-Site Mutation inPpp2r2aResults in a Novel Digenic Mouse Model of Type 2 Diabetes

Abstract

Insulin resistance in mice typically does not manifest as diabetes due to multiple compensatory mechanisms. Here, we present a novel digenic model of type 2 diabetes in mice heterozygous for a null allele of the insulin receptor and an N-ethyl-N-nitrosourea-induced alternative splice mutation in the regulatory protein phosphatase 2A (PP2A) subunit PPP2R2A. Inheritance of either allele independently results in insulin resistance but not overt diabetes. Doubly heterozygous mice exhibit progressive hyperglycemia, hyperinsulinemia, and impaired glucose tolerance from 12 weeks of age without significant increase in body weight. Alternative splicing of Ppp2r2a decreased PPP2R2A protein levels. This reduction in PPP2R2A containing PP2A phosphatase holoenzyme was associated with decreased serine/threonine protein kinase AKT protein levels. Ultimately, reduced insulin-stimulated phosphorylated AKT levels were observed, a result that was confirmed in Hepa1-6, C2C12, and differentiated 3T3-L1 cells knocked down using Ppp2r2a small interfering RNAs. Altered AKT signaling and expression of gluconeogenic genes in the fed state contributed to an insulin resistance and hyperglycemia phenotype. This model demonstrates how genetic changes with individually small phenotypic effects interact to cause diabetes and how differences in expression of hypomorphic alleles of PPP2R2A and potentially other regulatory proteins have deleterious effects and may therefore be relevant in determining diabetes risk.Type 2 diabetes is a complex disease where cellular resistance to insulin combined with a failure in b-cell compensation results in the development of the disease. Underlying this process are multiple genetic and environmental factors that interact to determine susceptibility risk. However, there are relatively few examples of patients with diabetes whose disease can be demonstrated to be due to the interaction of mutations in two or more genes. One of these is due to heterozygous mutations in two unlinked genes, peroxisome proliferator-activated receptor g (PPARG) and protein phosphatase 1, regulatory (inhibitor) subunit 3A (PPP1R3A), expressed in adipocytes and skeletal muscle, respectively, resulting in severe insulin resistance and lipodystrophy (1). A second example is haploinsufficiency for the insulin receptor (IR) in combination with chimerin 2 (CHN2), a GTPase-activating protein, that results in insulin resistance and deficiency in intrauterine growth (2). In this latter example, the CHN2 mutation implicates a novel gene in insulin signaling and its regulation of metabolism and growth (2). Although there are other examples of doubly heterozygous individuals with diabetes, e.g., in the maturity-onset diabetes of the young HNF1A and HNF4A genes, it is unclear how these impact the severity of disease (3). In a mouse model, a digenic insulin resistance phenotype has been described whereby 40% of mice heterozygous for both IR and insulin receptor substrate 1 (IRS-1) null alleles develop overt diabetes at 4-6 months of age, demonstra...

The Oncogenic Functions of MASTL Kinase

“…In contrast to ENSA, loss of PP2A-B55 has been widely implicated in regulating diverse biological pathways, including neurodegeneration ( Taleski and Sontag, 2018 ), metabolism ( Reid et al, 2013 ), diabetes ( Goldsworthy et al, 2016 ), DNA repair ( Kalev et al, 2012 ; Wang et al, 2015 ), the cell cycle ( Burgess et al, 2017 ), and of course tumor suppression ( Ruvolo, 2016 ). Importantly, knockdown of PP2A-B55 closely phenocopies MASTL overexpression, with loss of PP2A-B55 in MCF10A cells inducing excessive proliferation resulting in the formation of large lobular acini in 3D culture ( Watt et al, 2017 ).…”

Impaired insulin signaling in the B10.D2-Hc0H2dH2-T18c/oSnJ mouse model of complement factor 5 deficiency

“…We showed that C5 def mice exhibited severe glucose intolerance and systemic IR in both lean and obese states that was inflammation independent. IR in the C5 def B10.D2-Hc 0 H2 d H2-T18 c / oSnJ mouse strain was associated with a genetic mutation in the insulin receptor (Insr) gene and improper processing of pro-INSR in multiple metabolically active tissues, as has been reported in other models of INSR haplodeficiency (20). Interestingly, adenoviral overexpression of C5 improved insulin sensitivity in both C5 control (C5 cont ) and C5 def mice, lending support for a beneficial role of C5 in metabolism.…”

A cell cycle kinase-phosphatase module restrains PI3K-Akt activity in an mTORC1-dependent manner

“…Of note, Ensa knockout mice have improved glucose tolerance and are more insulin sensitive 57 . In the same direction, haploinsuficiency of the B55α subunit of PP2A causes insulin resistance likely due to the defective insulin-induced AKT stimulation in insulin-responsive tissues 58 . These observations are in line with a model in which MASTL phosphorylates ENSA to inhibit PP2A/B55 and AKT signaling (Fig.…”