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Growth factor stimulation promotes multivesicular endosome biogenesis by prolonging recruitment of the late-acting ESCRT machinery.

ABSTRACT: The formation of multivesicular endosomes (MVEs) mediates the turnover of numerous integral membrane proteins and has been implicated in the down-regulation of growth factor signaling, thereby exhibiting properties of a tumor suppressor. The endosomal sorting complex required for transport (ESCRT) machinery plays a key role in MVE biogenesis, enabling cargo selection and intralumenal vesicle (ILV) budding. However, the spatiotemporal pattern of endogenous ESCRT complex assembly and disassembly in mammalian cells remains poorly defined. By combining CRISPR/Cas9-mediated genome editing and live cell imaging using lattice light sheet microscopy (LLSM), we determined the native dynamics of both early- and late-acting ESCRT components at MVEs under multiple growth conditions. Specifically, our data indicate that ESCRT-0 accumulates quickly on endosomes, typically in less than 30 seconds, and its levels oscillate in a manner dependent on the downstream recruitment of ESCRT-I. Similarly, levels of the ESCRT-I complex also fluctuate on endosomes, but its average residency time is more than fivefold shorter compared with ESCRT-0. Vps4 accumulation is the most transient, however, suggesting that the completion of ILV formation occurs rapidly. Upon addition of epidermal growth factor (EGF), both ESCRT-I and Vps4 are retained at endosomes for dramatically extended periods of time, while ESCRT-0 dynamics are only modestly affected. Our findings are consistent with a model in which growth factor stimulation stabilizes late-acting components of the ESCRT machinery at endosomes to accelerate the rate of ILV biogenesis and attenuate signal transduction initiated by receptor activation.

SUBMITTER: Quinney KB

PROVIDER: S-EPMC6452702 | biostudies-literature | 2019 Apr

REPOSITORIES: biostudies-literature

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Growth factor stimulation promotes multivesicular endosome biogenesis by prolonging recruitment of the late-acting ESCRT machinery.

Quinney Kyle B KB Frankel Elisa B EB Shankar Raakhee R Kasberg William W Luong Peter P Audhya Anjon A

Proceedings of the National Academy of Sciences of the United States of America 20190320 14

The formation of multivesicular endosomes (MVEs) mediates the turnover of numerous integral membrane proteins and has been implicated in the down-regulation of growth factor signaling, thereby exhibiting properties of a tumor suppressor. The endosomal sorting complex required for transport (ESCRT) machinery plays a key role in MVE biogenesis, enabling cargo selection and intralumenal vesicle (ILV) budding. However, the spatiotemporal pattern of endogenous ESCRT complex assembly and disassembly i ...[more]

PMID: 30894482

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Project description:C-type lectins are key players in immune regulation by driving distinct functions of antigen-presenting cells. The C-type lectin CLEC16A gene is located at 16p13, a susceptibility locus for several autoimmune diseases, including multiple sclerosis. However, the function of this gene and its potential contribution to these diseases in humans are poorly understood. In this study, we found a strong upregulation of CLEC16A expression in the white matter of multiple sclerosis patients (n = 14) compared to non-demented controls (n = 11), mainly in perivascular leukocyte infiltrates. Moreover, CLEC16A levels were significantly enhanced in peripheral blood mononuclear cells of multiple sclerosis patients (n = 69) versus healthy controls (n = 46). In peripheral blood mononuclear cells, CLEC16A was most abundant in monocyte-derived dendritic cells, in which it strongly co-localized with human leukocyte antigen class II. Treatment of these professional antigen-presenting cells with vitamin D, a key protective environmental factor in multiple sclerosis, downmodulated CLEC16A in parallel with human leukocyte antigen class II. Knockdown of CLEC16A in distinct types of model and primary antigen-presenting cells resulted in severely impaired cytoplasmic distribution and formation of human leucocyte antigen class II-positive late endosomes, as determined by immunofluorescence and electron microscopy. Mechanistically, CLEC16A participated in the molecular machinery of human leukocyte antigen class II-positive late endosome formation and trafficking to perinuclear regions, involving the dynein motor complex. By performing co-immunoprecipitations, we found that CLEC16A directly binds to two critical members of this complex, RILP and the HOPS complex. CLEC16A silencing in antigen-presenting cells disturbed RILP-mediated recruitment of human leukocyte antigen class II-positive late endosomes to perinuclear regions. Together, we identify CLEC16A as a pivotal gene in multiple sclerosis that serves as a direct regulator of the human leukocyte antigen class II pathway in antigen-presenting cells. These findings are a first step in coupling multiple sclerosis-associated genes to the regulation of the strongest genetic factor in multiple sclerosis, human leukocyte antigen class II.

Dataset Information

Growth factor stimulation promotes multivesicular endosome biogenesis by prolonging recruitment of the late-acting ESCRT machinery.

Publications

Growth factor stimulation promotes multivesicular endosome biogenesis by prolonging recruitment of the late-acting ESCRT machinery.

Similar Datasets

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