Increased AT(1) Receptor Expression and Its Mediated Vasoconstriction Lead to Hypertension in Snx1(−/−) Mice

Angiotensin II type 1 receptor (AT(1)R) is a vital therapeutic target for hypertension. Sorting nexin 1 (SNX1) participates in the sorting and trafficking of renal dopamine D(5) receptor, while angiotensin and dopamine are counter-regulatory factors in regulating blood pressure. Thus, we supposed whether SNX1 has an effect on the AT(1)R sorting and trafficking. Snx1(−/−) mice, generated using the CRISPR/Cas9 system, showed a dramatic elevation in blood pressure compared to wild-type littermates. The angiotensin II-mediated contractile reactivity of mesenteric arteries and AT(1)R expression in aortas also rised in Snx1(−/−) mice. Moreover, immunofluorescence and immunoprecipitation analysis, respectively, revealed that SNX1 and AT(1)R were colocalized and interacted in aortas from wild-type mice. In vitro studies disclosed that AT(1)R protein level and its downstream calcium signaling were upregulated in A10 cells treated with SNX1 siRNA. This may result from decreased AT(1)R protein degradation because AT(1)R mRNA level had no change, but the expression of AT(1)R protein was less degraded when SNX1 knockdown, as reflected by the cycloheximide chase assay. Furthermore, the proteasomal rather than lysosomal inhibition increased AT(1)R protein content, accompanied by decayed binding of ubiquitin and AT(1)R after SNX1 knockdown. Confocal microscopy uncovered that AT(1)R colocalized with PSMD6, a proteasome marker, and the colocalization was reduced after SNX1 knockdown. These findings suggest that SNX1 sorts AT(1)R to proteasomal degradation and SNX1 impairment increases arterial AT(1)R expression, leading to higher vasoconstriction and increased blood pressure.