# Mechanisms of endocytic recycling

> **NIH NIH R01** · BRIGHAM AND WOMEN'S HOSPITAL · 2024 · $417,070

## Abstract

ABSTRACT
 Endocytic recycling is critical for a broad range of physiologic events, including nutrient uptake, cell motility
and polarity, intracellular signaling, and cytokinesis. We have been studying a coat complex that forms
transport vesicles in a recycling pathway, which involves ACAP1 (Arfgap with Coil-coil, Ankyrin repeat and PH
domain type 1) acting as the inner component and clathrin as the outer coating. Recently, we have made a
remarkable discovery, finding that the protein kinase Akt acts as another component of this coat complex. Akt
also possesses a direct ability to bend membrane, a finding that is unprecedented, as no kinase is known to
possess this capability. Our recent discovery has also led us to reconstitute vesicle formation by this coat
complex. Altogether, these findings lead us to propose three major goals. First, an ultimate understanding of
how a protein bends membrane is being achieved through a high-resolution cryo-electron microscopy (EM)
approach that solves the structure of the protein assembled on membrane. As this is the functional form of
coat proteins, we will collaborate with a group having demonstrated expertise in the EM-based approach to
elucidate in molecular detail how coat factors assemble into a protein lattice structure on membrane to achieve
membrane bending. Second, whereas protein-based mechanisms of vesicular transport are being intensely
investigated, lipid-based mechanisms have been far less explored. Addressing this fundamental shortcoming,
we have recently pursued the vesicle reconstitution system to identify lipid enzymes needed for vesicle
formation by the ACAP1 coat complex. Thus, we will elucidate the specific stage of vesicle formation that
requires a particular enzyme. Moreover, to achieve a more complete understanding of how the lipid product of
a particular enzyme acts, we will explore whether the geometry of the produced lipid affects ACAP1 vesicle
formation, and also whether a particular lipid geometry promotes the ability of the ACAP1 coat factors to bend
membrane. Third, we have recently performed mass spectrometry on the reconstituted ACAP1 vesicles to
implicate many cargoes of the ACAP1 pathway. To validate this finding, we will focus on unexpected cargoes
for further scrutiny, as confirmation that they use the ACAP1 pathway will provide particularly compelling
support that our approach has identified true cargoes of the ACAP1 pathway. Specifically, we will mutate the
sequence in these unexpected cargoes recognized by the ACAP1 coat complex and then confirm that sorting
into the ACAP1 pathway is inhibited. We will also pursue an unifying explanation for the unexpected cargoes
using the ACAP1 pathway by determining whether this transport results in their delivery to invadopodia, which
are localized cell-surface structures that concentrate key factors for matrix degradation, a process needed for
cell invasion into tissue. We anticipate that the completion of these studies will not o...

## Key facts

- **NIH application ID:** 10833614
- **Project number:** 5R01GM145618-02
- **Recipient organization:** BRIGHAM AND WOMEN'S HOSPITAL
- **Principal Investigator:** VICTOR W HSU
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $417,070
- **Award type:** 5
- **Project period:** 2023-05-01 → 2027-04-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10833614

## Citation

> US National Institutes of Health, RePORTER application 10833614, Mechanisms of endocytic recycling (5R01GM145618-02). Retrieved via AI Analytics 2026-05-25 from https://api.ai-analytics.org/grant/nih/10833614. Licensed CC0.

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