Loss of zebrafish atp6v1e1b, encoding a subunit of vacuolar ATPase, recapitulates human ARCL type 2C syndrome and identifies multiple pathobiological signatures

Abstract

The inability to maintain a strictly regulated endo(lyso)somal acidic pH through the proton-pumping action of the vacuolar-ATPases (v-ATPases) has been associated with various human diseases including heritable connective tissue disorders. Autosomal recessive (AR) cutis laxa (CL) type 2C syndrome is associated with genetic defects in the ATP6V1E1 gene and is characterized by skin wrinkles or loose redundant skin folds with pleiotropic systemic manifestations. The underlying pathological mechanisms leading to the clinical presentations remain largely unknown. Here, we show that loss of atp6v1e1b in zebrafish leads to early mortality, associated with craniofacial dysmorphisms, vascular anomalies, cardiac dysfunction, N-glycosylation defects, hypotonia, and epidermal structural defects. These features are reminiscent of the phenotypic manifestations in ARCL type 2C patients. Our data demonstrates that loss of atp6v1e1b alters endo(lyso)somal protein levels, and interferes with non-canonical v-ATPase pathways in vivo. In order to gain further insights into the processes affected by loss of atp6v1e1b, we performed an untargeted analysis of the transcriptome, metabolome, and lipidome in early atp6v1e1b-deficient larvae. We report multiple affected pathways including but not limited to oxidative phosphorylation, sphingolipid, fatty acid, and energy metabolism together with profound defects on mitochondrial respiration. Taken together, our results identify complex pathobiological effects due to loss of atp6v1e1b in vivo.

The transcription factor IRF-5 is essential for the metabolic rewiring of CD8 T cells during chronic infection

“…Interestingly, we found several genes that were expressed to a lower degree in Irf5 -/- compared to WT p14 cells. For example, Gpat2 , which encodes for glycerol-3-phosphate acyltransferase 2, an enzyme involved in triacylglycerol (TAG) synthesis (48); Awat2 , which encodes for acyl-CoA wax alcohol acyltransferase 2, an enzyme that catalyzes the synthesis of wax esters (49); Atp6v1b1 , which encodes for a vacuolar ATPase that has been shown to regulate oxidative phosphorylation, and sphingolipid, fatty acid, and energy metabolism, and to severely affect mitochondrial respiration in zebra fish (50); Ucp3 , which encodes for the uncoupling protein 3, a mitochondrial fatty acid anion exporter that protects mitochondria from lipid-induced damage (51); and Gpdl1 , which encodes for glycosylphosphatidylinositol-specific phospholipase D, an enzyme involved in triglyceride metabolism (Figure 5D). We confirmed the expression of some of the genes in WT and Irf5 -/- p14 cells purified ex-vivo from the spleen of female mice at d21 p.i.…”

Early mechanisms of aortic failure in a zebrafish model for thoracic aortic dissection and rupture

“…MITF is a known downstream target of BMP signaling that is inhibited by SMAD6 [59][60][61] . MITF also acts upstream of lysosome biogenesis and it was previously shown that lysosomal dysfunction in zebrafish due to loss of function of atp6v1e1b associates with dilatation of the ventral aorta and narrow aortic branches 28 .…”

“…MITF is a known downstream target of BMP signaling that is inhibited by SMAD6 59–61 . MITF also acts upstream of lysosome biogenesis and it was previously shown that lysosomal dysfunction in zebrafish due to loss of function of atp6v1e1b associates with dilatation of the ventral aorta and narrow aortic branches 28 . Finally, MITF is known to be important for cell metabolism and cell cycle regulation, promoting mitochondrial biogenesis and having complex effects on proliferation depending on the level of MITF activity 62 .…”

“…Zebrafish were outcrossed to the transgenic Tg(fli1:EGFP) line in order to visualize endothelial cells as previously described 28 . Head length was used as normalization factor.…”