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Related Experiment Video

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Analysis of Endocytic Uptake and Retrograde Transport to the Trans-Golgi Network Using Functionalized Nanobodies in Cultured Cells

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A FAM21-containing WASH complex regulates retromer-dependent sorting.

Timothy S Gomez¹, Daniel D Billadeau

¹Department of Immunology, Division of Oncology Research and Schulze Center for Novel Therapeutics, College of Medicine, Mayo Clinic, Rochester, MN 55905, USA. gomez.timothy@mayo.edu

Developmental Cell

|November 20, 2009

Summary

This summary is machine-generated.

WASH (Wiskott-Aldrich Syndrome Protein and SCAR Homolog) protein regulates endosomal sorting and trafficking by interacting with the Arp2/3 complex and microtubules. It is crucial for proper retromer-mediated transport and actin-driven tubule scission.

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The Microscopy-Based Assay to Study and Analyze the Recycling Endosomes using SNARE Trafficking

The Microscopy-Based Assay to Study and Analyze the Recycling Endosomes using SNARE Trafficking

Published on: February 12, 2022

Area of Science:

Cell Biology
Molecular Biology
Protein Trafficking

Background:

The Arp2/3 complex is vital for endocytosis, sorting, and trafficking, but its activators are not fully understood.
WASH (Wiskott-Aldrich Syndrome Protein and SCAR Homolog) is an Arp2/3 activator that evolved during primate radiation, with its specific function remaining unclear.

Purpose of the Study:

To elucidate the function of WASH in cellular trafficking pathways.
To investigate the molecular mechanisms by which WASH regulates endosomal sorting and transport.

Main Methods:

Immunofluorescence microscopy to determine WASH localization and interactions.
Biochemical assays to analyze WASH complex formation and Arp2/3 activation.
Functional assays to assess the role of WASH in retromer-mediated trafficking and actin dynamics.

Main Results:

WASH localizes to early endosomes, associating with tubulin, Arp2/3, F-actin, and retromer components.
WASH is not essential for endocytosis but regulates retromer-mediated retrograde CI-MPR trafficking.
WASH functions within a multiprotein complex including FAM21, linking endosomes to Arp2/3-driven actin regulation and tubulin for Golgi-directed transport.

Conclusions:

WASH plays a critical role in linking retromer-mediated tubules to microtubules for Golgi trafficking.
WASH utilizes Arp2/3-directed F-actin dynamics to generate force for tubule scission during retrograde transport.
The FAM21-WASH complex is essential for regulating retromer-mediated sorting and cargo trafficking.