GPRASP1 loss-of-function links to arteriovenous malformations by endothelial activating GPR4 signals

Abstract

Arteriovenous malformations (AVMs) are fast-flow vascular malformations and refer to important causes of intracerebral hemorrhage in young adults. Getting deep insight into the genetic pathogenesis of AVMs is necessary. Herein, we identified two vital missense variants of G protein-coupled receptor (GPCR) associated sorting protein 1 (GPRASP1) in AVM patients for the first time and congruously determined to be loss-of-function (LoF) variants in endothelial cells. GPRASP1 LoF caused endothelial dysfunction in vitro and in vivo. Endothelial Gprasp1 knockout mice suffered a high probability of cerebral hemorrhage, AVMs, and exhibited vascular anomalies in multiple organs. GPR4 was identified to be an effective GPCR binding with GPRASP1 to develop endothelial disorders. GPRASP1 deletion activated GPR4/cAMP/MAPK signaling to disturb endothelial functions, thus contributing to vascular anomalies. Mechanistically, GPRASP1 promoted GPR4 degradation. GPRASP1 enabled GPR4 K63-linked ubiquitination, enhancing the binding of GPR4 and RABGEF1 to activate RAB5 for conversions from endocytic vesicles to endosomes, and subsequently increasing the interactions of GPR4 and ESCRT members to package GPR4 into multivesicular bodies or late endosomes for lysosome degradation. Notably, the GPR4 antagonist NE 52-QQ57 and JNK inhibitor SP600125 effectively rescued the vascular phenotype caused by endothelial Gprasp1 deletion. Our findings provided novel insights into the roles of GPRASP1 in AVMs and hinted at new therapeutic strategies.

Expression of Endogenous Epitope-Tagged GPR4 in the Mouse Brain

“…GPR4 has been implicated in both peripheral and central physiological functions. It plays a role in angiogenesis ( Kim et al, 2005 ; Qiao et al, 2006 ; Yang et al, 2007 ; Ren et al, 2016 ; Miltz et al, 2017 ; Ouyang et al, 2021 ; Li et al, 2023 ), monocyte migration ( Huang et al, 2007 ; Chen et al, 2011 ; Dong et al, 2013 ), chronic inflammation ( Miltz et al, 2017 ; Sanderlin et al, 2017 , 2019 ; Krewson et al, 2020 ; Liu et al, 2020 ; Marie et al, 2023 ), ischemia/reperfusion injury ( Dong et al, 2017 ), maintenance of acid–base balance by the kidney ( Yang et al, 2007 ; Sun et al, 2010 , 2015 ; Cheval et al, 2021 ), and cancer cell migration/metastases ( Sin et al, 2004 ; Castellone et al, 2011 ; Wyder et al, 2011 ; Jing et al, 2016 ; Yu et al, 2019 ; Xue et al, 2020 ; Marie et al, 2023 ). In the central nervous system, GPR4 has been associated with central respiratory chemosensitivity, contributing both to an atypical CO 2 /H + -dependent vasoconstriction in brainstem regions associated with CO 2 -regulated breathing and to direct modulation by CO 2 /H + of putative respiratory chemosensory neurons in the retrotrapezoid nucleus ( Kumar et al, 2015 ; Hosford et al, 2018 ; Wenzel et al, 2020 ).…”

“…GPR4 has been implicated in both peripheral and central physiological functions. It plays a role in angiogenesis (Kim et al, 2005;Li et al, 2023;Miltz et al, 2017;Ouyang et al, 2021;Qiao et al, 2006;Ren et al, 2016;Yang et al, 2007), monocyte migration (Chen et al, 2011;L. Dong et al, 2013;Huang et al, 2007), chronic inflammation (Krewson et al, 2020;Liu et al, 2020;Marie et al, 2023;Miltz et al, 2017;Sanderlin et al, 2017Sanderlin et al, , 2019, ischemia/reperfusion injury (B.…”