Reduced Stability and pH-Dependent Activity of a Common Obesity-Linked PCSK1 Polymorphism, N221D

• Published in: Endocrinology (Philadelphia) 2019-11, Vol.160 (11), p.2630-2645
• Main Authors: Jarvela, Timothy S, Shakya, Manita, Bachor, Tomas, White, Anne, Low, Malcolm J, Lindberg, Iris
• Format: Article
• Language: English
• Subjects: Adrenocorticotropic hormone; alpha-MSH - metabolism; Animals; Body weight; Cloning; CRISPR; Endocrinology; Endoplasmic reticulum; Endoplasmic Reticulum Stress; Enzymatic activity; Enzyme Stability; Enzymes; Female; Females; Gene mutations; Gene polymorphism; Genetic aspects; Genotypes; Glucose Intolerance; Golgi apparatus; Health aspects; Heat stress; Heat tolerance; High fat diet; Humans; Hydrogen-Ion Concentration; Hypothalamus; Hypothalamus - metabolism; Male; Males; Mice; Mutants; Mutation; Neuropeptide Y - metabolism; Nucleotides; Obesity; Obesity - genetics; Peptides; pH effects; Pituitary; Pituitary Gland - metabolism; Polymorphism; Polymorphism, Single Nucleotide; Pro-Opiomelanocortin - metabolism; Proopiomelanocortin; Proprotein Convertase 1 - genetics; Proprotein Convertase 1 - metabolism; Proprotein convertases; Risk; Risk factors; Sex Characteristics; Sexual dimorphism; Single nucleotide polymorphisms; Single-nucleotide polymorphism; Tunicamycin
• ISSN: 1945-7170; 0013-7227; 1945-7170
• DOI: 10.1210/en.2019-00418

Common mutations in the human prohormone convertase (PC)1/3 gene (PCKSI) are linked to increased risk of obesity. Previous work has shown that the rs6232 single-nucleotide polymorphism (N221D) results in slightly decreased activity, although whether this decrease underlies obesity risk is not clear. We observed significantly decreased activity of the N221D PC1/3 enzyme at the pH of the trans-Golgi network; at this pH, the mutant enzyme was less stable than wild-type enzyme. Recombinant N221D PC1/3 also showed enhanced susceptibility to heat stress. Enhanced susceptibility to tunicamycin-induced endoplasmic reticulum stress was observed in AtT-20/PC2 cell clones in which murine PC1/3 was replaced by human N221D PC1/3, as compared with wild-type human PC1/3. However, N221D PC1/3-expressing AtT-20/PC2 clones processed proopiomelanocortin to α-MSH similarly to wild-type PC1/3. We also generated a CRISPR-edited mouse line expressing the N221D mutation in the PCKSI gene. When homozygous N221D mice were fed either a standard or a high-fat diet, we found no increase in body weight compared with their wild-type sibling controls. Sexual dimorphism was observed in pituitary ACTH for both genotypes, with females exhibiting lower levels of pituitary ACTH. In contrast, hypothalamic α-MSH content for both genotypes was higher in females compared with males. Hypothalamic corticotropin-like intermediate peptide content was higher in wild-type females compared with wild-type, but not N221D, males. Taken together, these data suggest that the increased obesity risk linked to the N221D allele in humans may be due in part to PC1/3-induced loss of resilience to stressors rather than strictly to decreased enzymatic activity on peptide precursors.