Reduced stability and pH-dependent activity of a common obesity-linked PCSK1 polymorphism, N221D

Common mutations in the human prohormone convertase 1/3 (PC1/3) gene (PCSK1) are linked to increased risk of obesity. Previous work has shown that the rs6232 SNP (N221D) results in slightly decreased activity, though 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 ER stress was observed in AtT-20/PC2 cell clones in which murine PC1/3 was replaced by human N221D PC1/3, as compared to 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 Pcsk1 gene. When homozygous N221D mice were fed either a standard or a high fat diet, we found no increase in body weight compared to 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 to males. Hypothalamic CLIP content was higher in wild-type females compared to wild-type, but not N221D, males. 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.