Tetraspanins interweave EV secretion, endosomal network dynamics and cellular metabolism

Tetraspanin proteins organize membrane nanodomains related to cell adhesion and migration. An essential feature conserved along the superfamily is their cone-shaped tertiary structure, which allows tetraspanins to be enriched in highly curved membrane structures. Their conical shape, together with their ability to associate to transmembrane receptors and to bind to cystoskeletal and signaling scaffolds, are key in their ability to regulate endosomal network dynamics and Extracellular Vesicle biogenesis and cargo selection. Recent evidence suggests that tetraspanins have a relevant impact in mitochondria turnover and regulation of cellular metabolism. In this review we highlight those reports that point to tetraspanins as key regulators in the communication between the endosomal network, EVs and the cellular metabolismWe thank Dr. Cabañas for critical reading of the manuscript. This work has been supported by grants BIO2017-86500-R, PID2020- 119627GB-I00, and RED2018-102411-T TeNTaCLES from Ministerio Español de Ciencia e Innovación (Spain) to MY-M. VT is supported by an FPI-UAM predoctoral fellowshi

ALCAM/CD166 is involved in the binding and uptake of cancer-derived extracellular vesicles

Colorectal cancer (CRC) and ovarian cancer (OvC) patients frequently develop peritoneal metastasis, a condition associated with a very poor prognosis. In these cancers, tumor-derived extracellular vesicles (EVs) cause immunosuppression, facilitate the direct attachment and invasion of cancer cells through the mesothelium, induce the conversion of peritoneal mesothelial cells (PMCs) into cancer-associated fibroblasts (CAFs) and transfer a more aggressive phenotype amongst cancer cells. Although the promoting role of EVs in CRC and OvC peritoneal metastasis is well established, the specific molecules that mediate the interactions between tumor-derived EVs and immune and non-immune target cells remain elusive. Here, we employed the SKOV-3 (ovarian adenocarcinoma) and Colo-320 (colorectal adenocarcinoma) human cell lines as model systems to study the interactions and uptake of EVs produced by ovarian carcinoma and colorectal carcinoma cells, respectively. We established that the adhesion molecule ALCAM/CD166 is involved in the interaction of cancerderived EVs with recipient cancer cells (a process termed “EV binding” or “EV docking”) and in their subsequent uptake by these cells. The identification of ALCAM/CD166 as a molecule mediating the docking and uptake of CRC and OvC-derived EVs may be potentially exploited to block the peritoneal metastasis cascade promoted by EVs in CRC and OvC patient

El papel de las tetraspaninas en la relación del aparato endosomal con el metabolismo celular y la identificación de nuevos mecanismos moleculares implicados en la captación de vesículas

Tesis Doctoral inédita leída en la Universidad Autónoma de Madrid, Facultad de Ciencias, Departamento de Biología Molecular. Fecha de Lectura: 28-03-2023Esta tesis tiene embargado el acceso al texto completo hasta el 28-09-2024Las tetraspaninas son proteínas transmembrana que por sus características estructurales se relacionan con regiones de membrana con una alta curvatura, como los compartimentos endosomales, estructuras migratorias o las vesículas extracelulares. Es por ello, que tienen un importante papel en la regulación de la red endosomal. Dicha red está a su vez estrechamente ligada al estado metabólico celular, lo cual plantea la posibilidad de que las tetraspaninas jueguen un papel fundamental en la regulación del metabolismo celular. En la presente tesis se explora el papel de distintas tetraspaninas (CD9, CD63 y CD81) como reguladores del sistema endosomal, de la biogénesis de vesículas extracelulares y del estado metabólico celular a través de la fisiología mitocondrial. Las vesículas extracelulares son nanopartículas de origen celular muy importantes en la comunicación intercelular. Debido a la juventud de este campo de investigación hay todavía pocas técnicas para su estudio y los mecanismos moleculares que están detrás de su fisiología todavía no están descritos en su mayor parte. En esta tesis doctoral nos interesamos por el proceso de captación de dichas vesículas extracelulares por parte de las células diana. Hemos desarrollado una técnica para su cuantificación y caracterización utilizando reporteros moleculares fusionados a las tetraspaninas y realizado un cribado de amplio espectro basado en la tecnología CRISPR para identificar nuevos mecanismos moleculares implicados en este proceso. Estos estudios revelaron la implicación del complejo COMMANDER en la captación de vesículas extracelularesTetraspanins are transmembrane proteins that due to their structural characteristics are related to membrane regions with a high curvature, such as endosomal compartments, migratory structures or extracellular vesicles. For this reason, they play an important role in the regulation of the endosomal network. This network is, in turn, closely linked to the cellular metabolic state, which raises the possibility that tetraspanins play a fundamental role in the regulation of cellular metabolism. The present thesis explores the role of different tetraspanins (CD9, CD63 and CD81) as regulators of the endosomal system, extracellular vesicle biogenesis and cellular metabolic state through mitochondrial physiology. Extracellular vesicles are cell-derived nanoparticles that are very important in intercellular communication. Due to the youth of this research field, there are still few techniques for their study and the molecular mechanisms behind their physiology are still largely undescribed. In this doctoral thesis we are interested in the uptake process of these extracellular vesicles by target cells. We have developed a technique for their quantification and characterization using molecular reporters fused to tetraspanins and performed a broad-spectrum screening based on CRISPR technology to identify new molecular mechanisms involved in this process. These studies revealed the involvement of the COMMANDER complex in extracellular vesicle uptak

CD9 inhibition reveals a functional connection of extracellular vesicle secretion with mitophagy in melanoma cells

Tetraspanins are often used as Extracellular Vesicle (EV) detection markers because of their abundance on these secreted vesicles. However, data on their function on EV biogenesis are controversial and compensatory mechanisms often occur upon gene deletion. To overcome this handicap, we have compared the effects of tetraspanin CD9 gene deletion with those elicited by cytopermeable peptides with blocking properties against tetraspanin CD9. Both CD9 peptide or gene deletion reduced the number of early endosomes. CD9 peptide induced an increase in lysosome numbers, while CD9 deletion augmented the number of MVB and EV secretion, probably because of compensatory CD63 expression upregulation. In vivo, CD9 peptide delayed primary tumour cell growth and reduced metastasis size. These effects on cell proliferation were shown to be concomitant with an impairment in mitochondrial quality control. CD9 KO cells were able to compensate the mitochondrial malfunction by increasing total mitochondrial mass reducing mitophagy. Our data thus provide the first evidence for a functional connection of tetraspanin CD9 with mitophagy in melanoma cells.Ministerio Espanol de Economia y Competitividad (MINECO), grant from the Fundacion Ramon Areces and Leonardo Grant from fBBVA to Maria Yanez-M

CD9 inhibition reveals a functional connection of extracellular vesicle secretion with mitophagy in melanoma cells

Tetraspanins are often used as Extracellular Vesicle (EV) detection markers because of their abundance on these secreted vesicles. However, data on their function on EV biogenesis are controversial and compensatory mechanisms often occur upon gene deletion. To overcome this handicap, we have compared the effects of tetraspanin CD9 gene deletion with those elicited by cytopermeable peptides with blocking properties against tetraspanin CD9. Both CD9 peptide or gene deletion reduced the number of early endosomes. CD9 peptide induced an increase in lysosome numbers, while CD9 deletion augmented the number of MVB and EV secretion, probably because of compensatory CD63 expression upregulation. In vivo, CD9 peptide delayed primary tumour cell growth and reduced metastasis size. These effects on cell proliferation were shown to be concomitant with an impairment in mitochondrial quality control. CD9 KO cells were able to compensate the mitochondrial malfunction by increasing total mitochondrial mass reducing mitophagy. Our data thus provide the first evidence for a functional connection of tetraspanin CD9 with mitophagy in melanoma cells.Ministerio Espanol de Economia y Competitividad (MINECO), grant from the Fundacion Ramon Areces and Leonardo Grant from fBBVA to Maria Yanez-M