Dietary rescue of lipotoxicity-induced mitochondrial damage in Peroxin19 mutants.

Mutations in peroxin (PEX) genes lead to loss of peroxisomes, resulting in the formation of peroxisomal biogenesis disorders (PBDs) and early lethality. Studying PBDs and their animal models has greatly contributed to our current knowledge about peroxisomal functions. Very-long-chain fatty acid (VLCFA) accumulation has long been suggested as a major disease-mediating factor, although the exact pathological consequences are unclear. Here, we show that a Drosophila Pex19 mutant is lethal due to a deficit in medium-chain fatty acids (MCFAs). Increased lipolysis mediated by Lipase 3 (Lip3) leads to accumulation of free fatty acids and lipotoxicity. Administration of MCFAs prevents lipolysis and decreases the free fatty acid load. This drastically increases the survival rate of Pex19 mutants without reducing VLCFA accumulation. We identified a mediator of MCFA-induced lipolysis repression, the ceramide synthase Schlank, which reacts to MCFA supplementation by increasing its repressive action on lip3. This shifts our understanding of the key defects in peroxisome-deficient cells away from elevated VLCFA levels toward elevated lipolysis and shows that loss of this important organelle can be compensated by a dietary adjustment.

Unbalanced lipolysis results in lipotoxicity and mitochondrial damage in peroxisome-deficient Pex19 mutants.

Inherited peroxisomal biogenesis disorders (PBDs) are characterized by the absence of functional peroxisomes. They are caused by mutations of peroxisomal biogenesis factors encoded by Pex genes, and result in childhood lethality. Owing to the many metabolic functions fulfilled by peroxisomes, PBD pathology is complex and incompletely understood. Besides accumulation of peroxisomal educts (like very-long-chain fatty acids [VLCFAs] or branched-chain fatty acids) and lack of products (like bile acids or plasmalogens), many peroxisomal defects lead to detrimental mitochondrial abnormalities for unknown reasons. We generated Pex19 Drosophila mutants, which recapitulate the hallmarks of PBDs, like absence of peroxisomes, reduced viability, neurodegeneration, mitochondrial abnormalities, and accumulation of VLCFAs. We present a model of hepatocyte nuclear factor 4 (Hnf4)-induced lipotoxicity and accumulation of free fatty acids as the cause for mitochondrial damage in consequence of peroxisome loss in Pex19 mutants. Hyperactive Hnf4 signaling leads to up-regulation of lipase 3 and enzymes for mitochondrial ß-oxidation. This results in enhanced lipolysis, elevated concentrations of free fatty acids, maximal ß-oxidation, and mitochondrial abnormalities. Increased acid lipase expression and accumulation of free fatty acids are also present in a Pex19-deficient patient skin fibroblast line, suggesting the conservation of key aspects of our findings.

Obstructor-A is required for epithelial extracellular matrix dynamics, exoskeleton function, and tubulogenesis.

The epidermis and internal tubular organs, such as gut and lungs, are exposed to a hostile environment. They form an extracellular matrix to provide epithelial integrity and to prevent contact with pathogens and toxins. In arthropods, the cuticle protects, shapes, and enables the functioning of organs. During development, cuticle matrix is shielded from premature degradation; however, underlying molecular mechanisms are poorly understood. Previously, we identified the conserved obstructor multigene-family, which encodes chitin-binding proteins. Here we show that Obstructor-A is required for extracellular matrix dynamics in cuticle forming organs. Loss of obstructor-A causes severe defects during cuticle molting, wound protection, tube expansion and larval growth control. We found that Obstructor-A interacts and forms a core complex with the polysaccharide chitin, the cuticle modifier Knickkopf and the chitin deacetylase Serpentine. Knickkopf protects chitin from chitinase-dependent degradation and deacetylase enzymes ensure extracellular matrix maturation. We provide evidence that Obstructor-A is required to control the presence of Knickkopf and Serpentine in the extracellular matrix. We propose a model suggesting that Obstructor-A coordinates the core complex for extracellular matrix protection from premature degradation. This mechanism enables exoskeletal molting, tube expansion, and epithelial integrity. The evolutionary conservation suggests a common role of Obstructor-A and homologs in coordinating extracellular matrix protection in epithelial tissues of chitinous invertebrates.

Expression and localization of clathrin heavy chain in Drosophila melanogaster.

Clathrin-coated vesicles mediate cellular endocytosis of nutrients and molecules that are involved in a variety of biological processes. Basic components of the vesicle coat are clathrin heavy chain (Chc) and clathrin light chain molecules. In Drosophila melanogaster the chc gene function has been analyzed in a number of previous studies mainly using genetic approaches. However, the chc mRNA and protein expression patterns have not been studied systematically. We have generated an antibody that specifically recognizes Chc and we have analyzed chc RNA and protein expression patterns throughout embryonic and larval stages. We found that chc mRNA and protein are highly expressed from early stages of embryogenesis onwards, consistent with genetic studies predicting a maternal contribution of the gene function. During subsequent stages mRNA and protein are co-expressed in all embryonic cells; however we found an up-regulation in specific tissues including the gut, the salivary glands, tracheal system and the epidermis. In addition the central nervous system and the nephrocyte-like garland cells show strong Chc expression at late embryogenesis. In larvae Chc is highly expressed in garland cells, imaginal discs, fat body, salivary glands and the ring gland. Subcellularly, we found Chc protein in a vesicle-like pattern within the cytoplasm and at the plasma membrane. Co-labeling studies show that Chc is partially in contact with the trans-Golgi network and co-localizes with markers for early endocytosis. Together, the antibody may serve as a new tool to study the function of Chc in clathrin-dependent cellular processes, such as endocytosis.

The Wurst protein: a novel endocytosis regulator involved in airway clearance and respiratory tube size control.

The mammalian lung and the Drosophila airways are composed of an intricate network of epithelial tubes that transports fluids or gases and converts during late embryogenesis from liquid- to air-filling. Conserved growth factor pathways have been characterized in model organisms such as Drosophila or the mouse that control patterning and branching of tubular networks. In contrast, knowledge of the coordination of respiratory tube size and physiology is still limited. Latest studies have shown that endocytosis plays a major role in size determination and liquid clearance of the respiratory tubes and a new key regulator of these processes was identified, the Drosophila Wurst protein. wurst encodes a J-domain transmembrane protein which is essential for Clathrin-mediated endocytosis. It is evolutionary conserved and single Wurst orthologs are found in mammals (termed DNAJC22). In this commentary, we discuss the role of Wurst/DNAJC22 and address whether these proteins may be general regulators of Clathrin-mediated endocytosis.

Wurst is essential for airway clearance and respiratory-tube size control.

The Drosophila melanogaster tracheal system and the mammalian lung are branching networks of tubular epithelia that convert during late embryogenesis from liquid- to air-filling. Little is known about how respiratory-tube size and physiology are coordinated. Here, we show that the Drosophila wurst gene encodes a unique J-domain transmembrane protein highly conserved in metazoa. In wurst mutants, respiratory-tube length is increased and lumen clearance is abolished, preventing gas filling of the airways. Wurst is essential for clathrin-mediated endocytosis, which is required for size determination and lumen clearance of the airways. wurst recruits heat shock cognate protein 70-4 and clathrin to the apical membrane of epithelial cells. The sequence conservation of the single Wurst orthologues in mice and humans offer new opportunities for genetic studies of clinically relevant lung syndromes caused by the failure of liquid clearance and respiratory-tube size control.