Abstract Body (Do not enter title and authors here): Introduction: Natriuretic peptide receptor 1 (NPR1) is a membrane-bound guanylate cyclase activated by atrial and brain natriuretic peptides (ANP and BNP), playing a central role in blood pressure (BP) regulation via vasorelaxation and modulation of intravascular volume. Published data suggest that NPR1 loss-of-function variants, L1034F and E967K, localized to the guanylyl cyclase domain, exhibit a partial decrease in NPR1 activation in response to ANP and BNP. In this study, we report a newly identified variant, I751N, associated with elevated blood pressure; however, the impact of this variant on NPR1 function remains unclear.
Research Questions: Does the NPR1 variant I751N affect the function of NPR1? If it does, what are the underlying mechanisms?

Methods: We performed genetic association analysis for BP using whole-exome sequencing data from 1,252,240 individuals. The functional characterization of NPR1 variants was examined using flow cytometry, cyclic guanosine monophosphate (cGMP) assays, western blotting, and internalization assay in HEK293 cells overexpressing either the wild-type (WT) NPR1 or the variants E967K and I751N.

Results: The NPR1 I751N variant was significantly associated with higher systolic BP (p = 2.9e-11; effect size = +5.3 mmHg per allele; alternative allele frequency = 0.02%). Flow cytometry confirmed that NPR1 cell surface expression was comparable across WT and both variant-expressing HEK293 stable cell lines. cGMP assays demonstrated partial loss of signaling with E967K and near-complete loss with I751N. The I751N protein had a lower molecular weight (~75–90 kDa) than WT and E967K (~119 kDa), suggesting truncation or degradation due to the introduction of a polar residue (N, Asparagine) in a predicted hydrophobic region. Furthermore, biosensor-labeled NPR1 antibody experiments revealed accelerated ligand internalization in NPR1-I751N cells compared to NPR1-WT cells.

Conclusion: The NPR1-I751N variant encodes a truncated receptor that rapidly internalizes ligands without triggering downstream signaling, suggesting that this variant may function as a decoy receptor, akin to NPR3. These findings reveal new mechanisms by which the NPR1-I751N variant impacts BP in individuals carrying this mutation.