The circulating enzyme peptidase M20 domain containing 1 (PM20D1) as a metabolic sensor : molecular basis and clinical implications

Abstract

PM20D1 is a secreted enzyme catalysing condensation of medium- or long-chain fatty acids and amino acids into N-acyl amino acids (NAAA) and the reverse hydrolytic reaction. Although overexpression of PM20D1 increases energy expenditure and counteracts high fat diet (HFD)-induced obesity in mice, the clinical relevance of the finding remains elusive. Moreover, the controversial conclusions between gain- and loss-of-function study in animals indicate that the role of PM20D1 in regulating energy metabolism still needs further confirmation. Therefore, this study aimed to: 1) explore the associations of the circulating PM20D1 and major NAAA levels with obesity-related metabolic complications in humans; 2) compare the pharmacological roles of PM20D1 under different environmental conditions in animals; 3) illustrate the underlying mechanisms whereby PM20D1 as a metabolic sensor based on its potential Zinc-binding property. Serum concentrations of PM20D1, oleic acid (C18:1) and major NAAA (C18:1-Leu and C18:1-Phe) in 256 subjects, including 78 lean and 178 overweight/obese individuals with or without type 2 diabetes (T2D), were measured with immunoassays and liquid chromatography-mass spectrometry, respectively. The impact of sulfonylurea and rosiglitazone on circulating PM20D1 and its metabolites levels was examined in 62 patients with T2D. Furthermore, recombinant adenovirus associated virus (AAV) was generated and intravenously injected into mice fed on either STC or HFD for overexpression of PM20D1 or luciferase (as control). Followed by the comparison of metabolic phenotypes, circulating C18:1, C18:1-Leu, C18:1-Phe and Zinc levels under different environmental temperatures (22°C, 30°C, and 6°C). Lastly, Zinc-binding property of PM20D1 was validated using Zinc chelating resin, and the role of Zinc on enzymatic activity were evaluated through the measurement of C18:1-Phe synthesized/hydrolyzed by PM20D1. In clinical study, serum PM20D1 level was significantly elevated in both obese mice and overweight/obese individuals, and was closely associated with circulating levels of C18:1-Leu and C18:1-Phe. Serum PM20D1, C18:1-Leu and C18:1-Phe concentrations correlated positively with several parameters of adiposity as well as fasting and 2-h postprandial glucose, HbA1c, fasting insulin, and HOMA-IR independent of BMI and age. Moreover, a significant elevation in circulating levels of PM20D1, C18:1-Leu and C18:1-Phe corresponding with increases in the number of components of the metabolic syndrome was observed. Treatment with sulfonylurea significantly decreased circulating PM20D1, C18:1-Leu and C18:1-Phe in patients with T2D. In animal studies, no significant difference in metabolic phenotypes was observed between PM20D1 overexpressed and control mice when housed under 22 °C or 30 °C. By contrast, HFD-fed mice with PM20D1 overexpression exhibited significant decrease in fat mass and improvement in glucose tolerance when being exposed to 6 °C. Mechanistically, mice serum level of Zinc increased during cold exposure, which preferentially suppressed the synthase activity of PM20D1 by binding with 192E site, but had little influence on its hydrolase activity, resulting to the elevated circulating level of C18:1, then led to the improvement of obesity-related metabolic disorders. The clinical results, taken together with the animal and in vitro studies, support the important role of PM20D1 involved in obesity-related metabolic complications, and suggest that PM20D1 may acts as a metabolic sensor to “sense” the changes in environmental temperature to adjust its metabolic/enzymatic activities.