Inhibition of the Expression of the Small Heat Shock Protein αB-Crystallin Inhibits Exosome Secretion in Human Retinal Pigment Epithelial Cells in Culture
Open accessAbstract
Exosomes carry cell type-specific molecular cargo to extracellular destinations and therefore act as lateral vectors of intercellular communication and transfer of genetic information from one cell to the other. We have shown previously that the small heat shock protein αB-crystallin (αB) is exported out of the adult human retinal pigment epithelial cells (ARPE19) packaged in exosomes. Here, we demonstrate that inhibition of the expression of αB via shRNA inhibits exosome secretion from ARPE19 cells indicating that exosomal cargo may have a role in exosome biogenesis (synthesis and/or secretion). Sucrose density gradient fractionation of the culture medium and cellular extracts suggests continued synthesis of exosomes but an inhibition of exosome secretion. In cells where αB expression was inhibited, the distribution of CD63 (LAMP3), an exosome marker, is markedly altered from the normal dispersed pattern to a stacked perinuclear presence. Interestingly, the total anti-CD63(LAMP3) immunofluorescence in the native and αB-inhibited cells remains unchanged suggesting continued exosome synthesis under conditions of impaired exosome secretion. Importantly, inhibition of the expression of αB results in a phenotype of the RPE cell that contains an increased number of vacuoles and enlarged (fused) vesicles that show increased presence of CD63(LAMP3) and LAMP1 indicating enhancement of the endolysosomal compartment. This is further corroborated by increased Rab7 labeling of this compartment (RabGTPase 7 is known to be associated with late endosome maturation). These data collectively point to a regulatory role for αB in exosome biogenesis possibly via its involvement at a branch point in the endocytic pathway that facilitates secretion of exosomes.
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RPE-derived exosomes participate in inflammatory signaling and immune modulation; exosomes released by healthy RPE decrease inflammation and downregulate the immune response, whereas exosomes released by stressed RPE promote inflammation and activate immune cells.
“a common theme that has emerged from these studies suggests that RPE-derived exosomes participate in inflammatory signaling and in immune modulation. Exosomes released by healthy RPE serve to decrease inflammation and downregulate the immune response, whereas exosomes released by stressed RPE promote inflammation and activate immune cells (Biasutto et al., 2013; Gangalum et al., 2016; Knickelbein”
From The cell biology of the retinal pigment epithelium (2020) · cites this paper as Gangalum et al., 2016
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The cell biology of the retinal pigment epithelium
Lakkaraju, A., Umapathy, A., Tan, L. X., Daniele, L., Philp, N. J., Boesze-Battaglia, K., & Williams, D. S. (2020) · Progress in Retinal and Eye Research · doi:10.1016/j.preteyeres.2020.100846 · Cited by 422 · see citing works
- Authors
- Gangalum, R. K., Bhat, A. M., Kohan, S. A., & Bhat, S. P.
- Affiliations
- University of California, Los Angeles (US)
- Corresponding author
- Bhat, Suraj P.
- Venue
- Journal of Biological Chemistry
- Publisher
- Elsevier BV
- Published
- 2016-06
- Volume
- 291
- Issue
- 25
- Pages
- 12930-12942
- ISSN
- 0021-9258 (ISSN-L)
- PMC
- PMC4933212
- OpenAlex
- W2342693488
- Cited by
- 36
- References
- 47
- Citations by year
- 2025: 2 · 2023: 4 · 2022: 1 · 2021: 6 · 2020: 5 · 2019: 6 · 2018: 6 · 2017: 5 · 2016: 1
- FWCI
- 3.12
- Type
- journal article
- Language
- en
- Open access
- hybrid
- doi.org
- Licence
- Locations
- Journal of Biological Chemistry (publishedVersion, cc-by)
- Europe PMC (PubMed Central) (publishedVersion)
- PubMed Central (publishedVersion)
- PubMed (publishedVersion)
- eScholarship (California Digital Library) (submittedVersion, cc-by)
- Funders
- National Institutes of Health (1R01EY024929)
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Extracellular vesicles in disease
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Cell Biology › Cellular transport and secretion
- Health Sciences › Medicine › Ophthalmology › Retinal Diseases and Treatments
- Keywords
- alphaB-crystallin, small heat shock proteins, exosome secretion, exosome biogenesis, retinal pigment epithelial cells, ARPE-19 cells, CD63, LAMP3, LAMP1, Rab7, endolysosomal compartment, endocytic pathway, shRNA knockdown
- MeSH terms
- rab7 GTP-Binding Proteins, Cell Line, Epithelial Cells, Humans, Lysosomes, Endosomes, Vacuoles, Gene Expression, rab GTP-Binding Proteins, Protein Transport, RNA, Small Interfering, alpha-Crystallin B Chain, Retinal Pigment Epithelium, Exosomes, Gene Knockdown Techniques, Lysosomal-Associated Membrane Protein 3
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