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Retrofusion of intralumenal MVB membranes parallels viral infection and coexists with exosome release.

ABSTRACT: The endosomal system constitutes a highly dynamic vesicle network used to relay materials and signals between the cell and its environment.¹ Once internalized, endosomes gradually mature into late acidic compartments and acquire a multivesicular body (MVB) organization through invagination of the limiting membrane (LM) to form intraluminal vesicles (ILVs).² Cargoes sequestered into ILVs can either be delivered to lysosomes for degradation or secreted following fusion of the MVB with the plasma membrane.³ It has been speculated that commitment to ILVs is not a terminal event, and that a return pathway exists, allowing "back-fusion" or "retrofusion" of intraluminal membranes to the LM.⁴ The existence of retrofusion as a way to support membrane equilibrium within the MVB has been widely speculated in various cell biological contexts, including exosome uptake⁵ and major histocompatibility complex class II (MHC class II) antigen presentation.^6-9 Given the small physical scale, retrofusion of ILVs cannot be measured with conventional techniques. To circumvent this, we designed a chemically tunable cell-based system to monitor retrofusion in real time. Using this system, we demonstrate that retrofusion occurs as part of the natural MVB lifestyle, with attributes parallel to those of viral infection. Furthermore, we find that retrofusion and exocytosis coexist in an equilibrium, implying that ILVs inert to retrofusion comprise a significant fraction of exosomes destined for secretion. MVBs thus contain three types of ILVs: those committed to lysosomal degradation, those retrofusing ILVs, and those subject to secretion in the form of exosomes. VIDEO ABSTRACT.

SUBMITTER: Perrin P

PROVIDER: S-EPMC8445322 | biostudies-literature |

REPOSITORIES: biostudies-literature

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ISGylation controls exosome secretion by promoting lysosomal degradation of MVB proteins.

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Phosphorylation coexists with O-GlcNAcylation in a plant virus protein and influences viral infection.

Project description:Phosphorylation and O-GlcNAcylation are two widespread post-translational modifications (PTMs), often affecting the same eukaryotic target protein. Plum pox virus (PPV) is a member of the genus Potyvirus which infects a wide range of plant species. O-GlcNAcylation of the capsid protein (CP) of PPV has been studied extensively, and some evidence of CP phosphorylation has also been reported. Here, we use proteomics analyses to demonstrate that PPV CP is phosphorylated in vivo at the N-terminus and the beginning of the core region. In contrast with the 'yin-yang' mechanism that applies to some mammalian proteins, PPV CP phosphorylation affects residues different from those that are O-GlcNAcylated (serines Ser-25, Ser-81, Ser-101 and Ser-118). Our findings show that PPV CP can be concurrently phosphorylated and O-GlcNAcylated at nearby residues. However, an analysis using a differential proteomics strategy based on iTRAQ (isobaric tags for relative and absolute quantitation) showed a significant enhancement of phosphorylation at Ser-25 in virions recovered from O-GlcNAcylation-deficient plants, suggesting that crosstalk between O-GlcNAcylation and phosphorylation in PPV CP takes place. Although the preclusion of phosphorylation at the four identified phosphotarget sites only had a limited impact on viral infection, the mimicking of phosphorylation prevents PPV infection in Prunus persica and weakens infection in Nicotiana benthamiana and other herbaceous hosts, prompting the emergence of potentially compensatory second mutations. We postulate that the joint action of phosphorylation and O-GlcNAcylation in the N-proximal segment of CP allows a fine-tuning of protein stability, providing the amount of CP required in each step of viral infection.

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2022-03-03 | PXD006159 | Pride

Exosome Release Is Regulated by mTORC1.

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