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Adenofection: A Method for Studying the Role of Molecular Chaperones in Cellular Morphodynamics by Depletion-Rescue Experiments
Published on: September 16, 2016
Subversion of CtBP1-controlled macropinocytosis by human adenovirus serotype 3
Beat Amstutz1, Michele Gastaldelli, Stefan Kälin
1Institute of Zoology, University of Zürich, Zürich, Switzerland.
Abstract:
Endocytosis supports cell communication, growth, and pathogen infection. The species B human adenovirus serotype 3 (Ad3) is associated with epidemic conjunctivitis, and fatal respiratory and systemic disease. Here we show that Ad3 uses dynamin-independent endocytosis for rapid infectious entry into epithelial and haematopoietic cells. Unlike Ad5, which uses dynamin-dependent endocytosis, Ad3 endocytosis spatially and temporally coincided with enhanced fluid-phase uptake. It was sensitive to macropinocytosis inhibitors targeting F-actin, protein kinase C, the sodium-proton exchanger, and Rac1 but not Cdc42. Infectious Ad3 macropinocytosis required viral activation of p21-activated kinase 1 (PAK1) and the C-terminal binding protein 1 of E1A (CtBP1), recruited to macropinosomes. These macropinosomes also contained the Ad3 receptors CD46 and alpha v integrins. CtBP1 is a phosphorylation target of PAK1, and is bifunctionally involved in membrane traffic and transcriptional repression of cell cycle, cancer, and innate immunity pathways. Phosphorylation-defective S147A-CtBP1 blocked Ad3 but not Ad5 infection, providing a direct link between PAK1 and CtBP1. The data show that viruses induce macropinocytosis for infectious entry, a pathway used in antigen presentation and cell migration.
Insights
Human adenovirus serotype 3 (Ad3) rapidly infects cells via macropinocytosis, a dynamin-independent pathway. This process requires viral activation of p21-activated kinase 1 (PAK1) and C-terminal binding protein 1 (CtBP1) for infectious entry.
Area of Science:
- Cell Biology
- Virology
- Immunology
Background:
- Endocytosis is crucial for cellular processes including communication, growth, and pathogen invasion.
- Human adenovirus serotype 3 (Ad3) causes severe respiratory and systemic diseases, including epidemic conjunctivitis.
Purpose of the Study:
- To elucidate the mechanism of Ad3 entry into host cells.
- To investigate the role of specific cellular pathways and proteins in Ad3 infection.
Main Methods:
- Studied Ad3 entry into epithelial and haematopoietic cells, comparing it with Ad5.
- Utilized macropinocytosis inhibitors and analyzed the involvement of F-actin, protein kinase C, sodium-proton exchanger, Rac1, Cdc42, p21-activated kinase 1 (PAK1), and C-terminal binding protein 1 (CtBP1).
- Investigated the interaction of Ad3 receptors (CD46, alpha v integrins) within macropinosomes.
Main Results:
- Ad3 utilizes a rapid, dynamin-independent endocytosis pathway that coincides with enhanced fluid-phase uptake (macropinocytosis).
- Ad3 infection depends on viral activation of PAK1 and CtBP1, which are recruited to macropinosomes containing Ad3 receptors.
- A phosphorylation-defective CtBP1 mutant (S147A-CtBP1) specifically blocked Ad3, but not Ad5, infection, confirming a direct link between PAK1 and CtBP1 in Ad3 entry.
Conclusions:
- Viruses, like Ad3, can hijack and induce macropinocytosis for efficient infectious entry.
- This study reveals a novel mechanism of viral entry involving PAK1 and CtBP1 activation during macropinocytosis.
- The findings highlight macropinocytosis as a critical pathway for viral pathogenesis, with implications for antigen presentation and cell migration.
