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

Abstract

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.

The endoplasmic reticulum of trypanosomatids: An unrevealed road for chemotherapy

“…(Volkmar and Christianson, 2020). In a study by Iyer et al (2022), biochemical, proteomic, and imaging approaches were used to study the EMC (TbEMC) in T. brucei. This study revealed that TbEMC is formed by 9 subunits with 20-40% of similarity with yeast and human homologs.…”

“…( Volkmar and Christianson, 2020 ). In a study by Iyer et al. (2022) , biochemical, proteomic, and imaging approaches were used to study the EMC (TbEMC) in T. brucei .…”

“…Indeed, the knocking out or knocking down of single TbEMC subunits led to growth defects on T. brucei procyclic forms and the disruption of de novo synthesis of phosphatidylcholine and or phosphatidylethanolamine, the two most abundant phospholipid classes in T. brucei , pointing to a role of the TbEMC in phospholipid synthesis. In addition, in TbEMC7 knockout parasites, TbEMC3 is released from the complex, implying that TbEMC7 is essential for the maintenance of the TbEMC integrity ( Iyer et al., 2022 ).…”

ER membrane complex (EMC): Structure, functions, and roles in diseases

“…Loss or reduction of EMC members results in defective cellular processes and renders them susceptible to various viruses 16–19 . Moreover, elimination or reduced levels of a single EMC subunit leads to growth defects in Trypanosoma brucei procyclic forms due to disruption of ab initio synthesis of phosphatidylcholine (PC) and phosphatidylethanolamine (PE) 20 …”

“…[16][17][18][19] Moreover, elimination or reduced levels of a single EMC subunit leads to growth defects in Trypanosoma brucei procyclic forms due to disruption of ab initio synthesis of phosphatidylcholine (PC) and phosphatidylethanolamine (PE). 20 Recent studies underscore the vital role of EMC in maintaining ER function, impacting processes such as protein and lipid synthesis, organelle communications, ER stress, viral infection, and various diseases. 7,8,[21][22][23][24][25][26] Given the newfound significance of EMC in biochemical processes and diseases, investigations into their structure, functions, and pathogenesis-related studies have become an area of considerable interest.…”