The endoplasmic reticulum membrane protein complex localizes to the mitochondrial - endoplasmic reticulum interface and its subunits modulate phospholipid biosynthesis in Trypanosoma brucei

The endoplasmic reticulum membrane complex (EMC) is a versatile complex that plays a key role in membrane protein biogenesis in the ER. Deletion of the complex has wide-ranging consequences including ER stress, disturbance in lipid transport and organelle tethering, among others. Here we report the function and organization of the evolutionarily conserved EMC (TbEMC) in the highly diverged eukaryote, Trypanosoma brucei. Using (co-) immunoprecipitation experiments in combination with mass spectrometry and whole cell proteomic analyses of parasites after depletion of select TbEMC subunits, we demonstrate that the TbEMC is composed of 9 subunits that are present in a high molecular mass complex localizing to the mitochondrial-endoplasmic reticulum interface. Knocking out or knocking down of single TbEMC subunits led to growth defects of T. brucei procyclic forms in culture. Interestingly, we found that depletion of individual TbEMC subunits lead to disruption of de novo synthesis of phosphatidylcholine (PC) or phosphatidylethanolamine (PE), the two most abundant phospholipid classes in T. brucei. Downregulation of TbEMC1 or TbEMC3 inhibited formation of PC while depletion of TbEMC8 inhibited PE synthesis, pointing to a role of the TbEMC in phospholipid synthesis. In addition, we found that in TbEMC7 knock-out parasites, TbEMC3 is released from the complex, implying that TbEMC7 is essential for the formation or the maintenance of the TbEMC. Author summary The endoPlasmic reticulum membrane complex (EMC) was first reported more than a decade back Since then, emerging research has indicated that the EMC is involved in insertion of endoplasmic reticulum membrane proteins. Its disruption has diverse effects on cellular functions including protein and lipid synthesis and may contribute to neuro-developmental disorders and cancer. The EMC is highly conserved and has been extensively studied in humans and yeast. Here we use biochemical, microscopy and proteomic analyses to study the EMC (TbEMC) in the unicellular parasite, Trypanosma brucei. We report that the TbEMC forms a complex at the mitochondrial-endoplasmic reticulum interface and regulates the synthesis of the two most abundant membrane Phospholipid classes. In addition, our study provides new insights into the localization, complex formation, function and interaction partners of the TbEMC and its subunits. Deciphering the cellular role of the EMC is key to understanding its involvement in disease biology and designing preventive strategies.