Disruption of BMP9 signalling contributes to altered acid ceramidase expression and processing in preeclampsia

Sphingolipids are critical bioactive mediators of cellular events. Ceramide (CER) is central to sphingolipid metabolism and its levels are kept in balance via the action of key regulatory enzymes that function by modulating its synthesis and breakdown in a variety of patho-physiological conditions. In particular, lysosomal acid ceramidase (ASAH1) is responsible for hydrolyzing CER into sphingosine (SPH). We have recently reported that altered sphingolipid metabolism in preeclampsia and IUGR, is in part due to impaired TGFß signaling. Bone morphogenetic protein 9 is a member of the TGFß superfamily that functions as a high affinity ligand for the Activin Receptor-like Kinase 1(ALK1) thereby activating R-SMAD1 signalling. Herein, we investigated the role of BMP9 in regulating ASAH1 expression in physiological and pathological conditions. Human choriocarcinoma JEG3 cells were treated with BMP9 (5–10 ng/ml) or control vehicle and examined ASAH1 protein expression by Western Blotting. Exposure of JEG3 cells to BMP9 resulted in increased ASAH1 protein expression. Inhibition of ALK1 signalling using the ALK1 inhibitor Dorsomorphin reversed the BMP9 stimulatory effect and led to an accumulation of inactive ASAH1 precursors. Immunoprecipitation of ASAH1 followed by immunoblotting with concanavalin A (identifies N-glycans) revealed decreased ASAH1 glycosylation following dorsomorphin treatment indicating an ALK1 signalling dependent regulation of ASAH1 maturation. Immunofluorescence analysis showed that dorsomorphin treatment resulted in co-localization of ASAH1with the endoplasmic reticulum marker (ER) calreticulin, in line with reduced glycosylation and trafficking of ASAH1 from the ER to the lysosomes. Of clinical relevance, BMP9, ALK1 and pophorylated SMAD1 protein expression levels were markedly decreased in preeclamptic placentae relative to age-matched controls. Our data implicates a novel role for BMP9 signaling via ALK1 in regulating ASAH1 expression, processing and trafficking in the human placenta. Disruption of this signalling pathway may in part contribute to altered sphingolipid metabolism found in preeclampsia. (Supported by CIHR)