GPR37 is processed in the N‐terminal ectodomain by ADAM10 and furin

GPR37 is an orphan G protein-coupled receptor (GPCR) implicated in several neurological diseases and important physiological pathways in the brain. We previously reported that its long N-terminal ectodomain undergoes constitutive metalloprotease-mediated cleavage and shedding, which have been rarely described for class A GPCRs. Here, we demonstrate that the protease that cleaves GPR37 at Glu167↓Gln168 is a disintegrin and metalloprotease 10 (ADAM10). This was achieved by employing selective inhibition, RNAi-mediated downregulation, and genetic depletion of ADAM10 in cultured cells as well as in vitro cleavage of the purified receptor with recombinant ADAM10. In addition, the cleavage was restored in ADAM10 knockout cells by overexpression of the wild type but not the inactive mutant ADAM10. Finally, postnatal conditional depletion of ADAM10 in mouse neuronal cells was found to reduce cleavage of the endogenous receptor in the brain cortex and hippocampus, confirming the physiological relevance of ADAM10 as a GPR37 sheddase. Additionally, we discovered that the receptor is subject to another cleavage step in cultured cells. Using site-directed mutagenesis, the site (Arg54↓Asp55) was localized to a highly conserved region at the distal end of the ectodomain that contains a recognition site for the proprotein convertase furin. The cleavage by furin was confirmed by using furin-deficient human colon carcinoma LoVo cells and proprotein convertase inhibitors. GPR37 is thus the first multispanning membrane protein that has been validated as an ADAM10 substrate and the first GPCR that is processed by both furin and ADAM10. The unconventional N-terminal processing may represent an important regulatory element for GPR37

GPR37 is processed in the N-terminal ectodomain by ADAM10 and furin

Abstract GPR37 is an orphan G protein-coupled receptor (GPCR) implicated in several neurological diseases and important physiological pathways in the brain. We previously reported that its long N-terminal ectodomain undergoes constitutive metalloprotease-mediated cleavage and shedding, which have been rarely described for class A GPCRs. Here, we demonstrate that the protease that cleaves GPR37 at Glu167↓Gln168 is a disintegrin and metalloprotease 10 (ADAM10). This was achieved by employing selective inhibition, RNAi-mediated downregulation, and genetic depletion of ADAM10 in cultured cells as well as in vitro cleavage of the purified receptor with recombinant ADAM10. In addition, the cleavage was restored in ADAM10 knockout cells by overexpression of the wild type but not the inactive mutant ADAM10. Finally, postnatal conditional depletion of ADAM10 in mouse neuronal cells was found to reduce cleavage of the endogenous receptor in the brain cortex and hippocampus, confirming the physiological relevance of ADAM10 as a GPR37 sheddase. Additionally, we discovered that the receptor is subject to another cleavage step in cultured cells. Using site-directed mutagenesis, the site (Arg54↓Asp55) was localized to a highly conserved region at the distal end of the ectodomain that contains a recognition site for the proprotein convertase furin. The cleavage by furin was confirmed by using furin-deficient human colon carcinoma LoVo cells and proprotein convertase inhibitors. GPR37 is thus the first multispanning membrane protein that has been validated as an ADAM10 substrate and the first GPCR that is processed by both furin and ADAM10. The unconventional N-terminal processing may represent an important regulatory element for GPR37

Ecto-GPR37:a potential biomarker for Parkinson’s disease

Abstract Objective: α-Synuclein has been studied as a potential biomarker for Parkinson’s disease (PD) with no concluding results. Accordingly, there is an urgent need to find out reliable specific biomarkers for PD. GPR37 is an orphan G protein-coupled receptor that toxically accumulates in autosomal recessive juvenile parkinsonism. Here, we investigated whether GPR37 is upregulated in sporadic PD, and thus a suitable potential biomarker for PD. Methods: GPR37 protein density and mRNA expression in postmortem substantia nigra (SN) from PD patients were analysed by immunoblot and RT-qPCR, respectively. The presence of peptides from the N-terminus-cleaved domain of GPR37 (i.e. ecto-GPR37) in human cerebrospinal fluid (CSF) was determined by liquid chromatography-mass spectrometric analysis. An engineered in-house nanoluciferase-based immunoassay was used to quantify ecto-GPR37 in CSF samples from neurological control (NC) subjects, PD patients and Alzheimer’s disease (AD) patients. Results: GPR37 protein density and mRNA expression were significantly augmented in sporadic PD. Increased amounts of ecto-GPR37 peptides in the CSF samples from PD patients were identified by mass spectrometry and quantified by the in-house ELISA method. However, the CSF total α-synuclein level in PD patients did not differ from that in NC subjects. Similarly, the cortical GPR37 mRNA expression and CSF ecto-GPR37 levels in AD patients were also unaltered. Conclusions: GPR37 expression is increased in SN of sporadic PD patients. The ecto-GPR37 peptides are significantly increased in the CSF of PD patients, but not in AD patients. These results open perspectives and encourage further clinical studies to confirm the validity and utility of ecto-GPR37 as a potential PD biomarker

Estudio descriptivo de los trabajadores de servicios sanitarios de una dirección de Atención primaria confinados por COVID19

Background: The Covid-19 pandemic is testing the resistance of health systems, the preservation of health professionals is a priority in processes of this type. The professionals’ exposure to suspicious contacts often requires their confinement. The objective was to know the epidemiological characteristics of the primary care professionals who required confinement. Methods: The research was carried out in the North Metropolitan Primary Care Area of Barcelona, from February 17 to May 3, 2020. 1,418 professionals who required confinement due to the epidemic by Covid-19 participated. The reasons for confinement, symptomatology, the confinement time and the results of PCR tests results were recorded. Univariate descriptive analysis was performed. Results: 78.8% of the professionals were women and the mean age was 45.2 years. 67.8% were doctors and nurses, in the remaining 32.2% there were different healthcare and non-care professionals. 64.1% of the sample presented symptoms compatible with Covid-19. Participants described multiple symptoms during confinement. 1,050 diagnostic RT- PCR tests were performed, being positive in 323 cases, of which 33 were in asymptomatic people. Conclusions: The impact of the epidemic by Covid-19 is anticipated in health personnel compared to the general population. The distribution of symptoms in healthcare professionals is similar to that of other studies in the general population. Of the total number of professionals requiring isolation, 22.7% confirmed the diagnosis.Fundamentos: La pandemia de la Covid-19 está poniendo a prueba la resistencia de los sistemas sanitarios. La preservación de los profesionales sanitarios es prioritaria siempre y especialmente ante situaciones de este tipo. La exposición de los profesionales frente a contactos sospechosos obliga en muchas ocasiones a su confinamiento. El objetivo fue conocer las características epidemiológicas de los profesionales de atención primaria que han precisado confinamiento. Métodos: La investigación se llevó a cabo en el área de atención primaria Metropolitana Nord de Barcelona, desde el 17 de febrero al 3 de mayo de 2020. Participaron 1.418 profesionales que requirieron confinamiento debido a la epidemia por la Covid-19. Se registraron los motivos de confinamiento, sintomatología, tiempo de confinamiento y resultados de las pruebas PCR. Se realizó análisis descriptivo univariante. Resultados: El 78,8% de los profesionales eran mujeres y la edad media de fue 45,2 años. El 67,8% fueron facultativos y enfermeras, en el 32,2% restante había diferentes profesionales asistenciales y no asistenciales. El 64,1% de la muestra presentó sintomatología compatible con Covid-19. Los participantes describieron múltiples síntomas durante el confinamiento. Se realizaron 1.050 pruebas diagnósticas RT-PCR resultando positivas en 323 casos, de los que 33 fueron en personas asintomáticas. Conclusiones: El impacto de la epidemia por Covid-19 se adelanta en el personal sanitario respecto a la población general. La distribución de síntomas en profesionales sanitarios es similar a la de otros estudios en población general. Del total de profesionales que precisan confinamiento en el 22,7% se confirma el diagnóstico