CCT as a modulator of extracellular vesicles production and T cell metabolism through kinesin dynamics

• Autores: Amelia Rodriguez Gómez
• Directores de la Tesis: Francisco Sánchez Madrid (dir. tes.), Noa Beatriz Martín Cófreces (dir. tes.)
• Lectura: En la Universidad Autónoma de Madrid ( España ) en 2023
• Idioma: inglés
• Número de páginas: 259
• Títulos paralelos:
  • CCT como modulador de la producción de vesículas extracelulares y el metabolismo de linfocitos T mediante las dinámicas de kinesina
• Tribunal Calificador de la Tesis: Carlos Cabañas (presid.), María Yañez Mo (secret.), Héctor Peinado (voc.), Pedro Roda Navarro (voc.), Begoña Sot Sanz (voc.)
• Programa de doctorado: Programa de Doctorado en Biociencias Moleculares por la Universidad Autónoma de Madrid
• Materias:
  • Química
    • Bioquímica
      • Biología molecular
  • Ciencias de la vida
    • Biología celular
    • Inmunología
    • Biología molecular
• Enlaces
  • Tesis en acceso abierto en: Biblos-e Archivo
• Resumen

  • Cell proteostasis includes gene transcription, protein translation, folding of de novo proteins, post-translational modifications, secretion, degradation and recycling. One of the main mechanisms of protein homeostasis maintenance involves the action of the endolysosomal system (ELS). ELS includes lysosome-mediated degradation pathway, but also extracellular vesicles (EVs) biosynthesis and release as a clearance mechanism. EVs contain proteins, lipids, nucleic acids and metabolites in their composition. Another relevant modulators of cell proteostasis are chaperonins. The eukaryotic chaperonin complex CCT has been described to be involved in the folding of key proteins related to cytoskeleton dynamics, intracellular signalling and cell cycle regulation. CCT implication in T cell metabolism, organelle positioning and cell activation through the T cell receptor (TCR) has remained unexplored until now. We found that all the CCT subunits and associated co-chaperones are loaded into EVs of T cells by profiling the proteome of these vesicles. By limiting CCT cell-content and functionality employing specific siRNAs, we studied the proteome of lymphoblastoid T cells and their released EVs. We showed that CCT controls the proteins that are loaded into EVs and regulates the proteomic profile of these cells. Our lipidomic and metabolic analyses demonstrated that CCT silencing also causes an altered lipid composition and metabolic rewiring towards a lipid-dependent metabolism. Moreover, we detected an increased glycolysis rate and peroxisome number and activity under CCT limited-content conditions. Lipid droplets, other relevant organelles involved in lipid metabolism, were also smaller and in lesser amounts in these conditions. These changes were accompanied by a dysregulation of the dynamics of interorganelle contacts between lipid droplets, mitochondria, peroxisomes and the ELS. This process accelerates the maturation of multivesicular bodies, leading to higher EVs production. CCT was involved in the dynamic regulation of microtubule-based kinesin motors through the posttranslational modification acetylation. Due to this microtubule and kinesin dynamics dysregulation, an altered organelle distribution was observed. These findings relate chaperonin functioning with organelle positioning and cellular metabolism regulation. Since organelle redistribution and metabolic reprogramming are key events to a successful T cell activation, we studied the role of CCT on this process. We observed an altered centriolar and microtubule organization, and an impaired TCR downstream signalling, including mTORC1-depending pathways dysregulation upon T cell activation when cells expressed limited amounts of CCT. Moreover, CD3 clustering at the immune synapse area was deficient under CCT limited functioning. Altogether, our findings reveal an unexpected role of CCT in organelle distribution, cellular metabolism regulation and, consequently, T cell activation through cytoskeleton dynamics control