# Structural-Functional Basis of Actin Cytoskeleton Dynamics

> **NIH NIH R01** · UNIVERSITY OF PENNSYLVANIA · 2021 · $360,364

## Abstract

Project Summary:
The long-term goal is to understand the structural and functional mechanisms that control actin cytoskeleton
dynamics in health and disease. In its fourth cycle, this grant addresses important gaps of knowledge in
previous focus areas, while also expanding into new areas in response to recent developments in the field.
Building upon the recent discovery by this lab in collaboration with a group in Norway of actin's dedicated N-
terminal acetyltransferase (Naa80), Aim-1 will focus on the mechanism and function of actin N-terminal
acetylation (N-acetylation) and isoform variations. Aim-1a will determine the mechanism of N-acetylation,
including biochemical and cellular studies of the Naa80-actin interaction and a series of crystal structures of
intermediate reaction steps, and using different actin isoforms. Aim-1b will test the hypothesis that myosin
activity is strongly dictated by actin isoform variations and N-acetylation. This will resolve an enduring
deficiency in the field; most myosin studies have used α-skeletal actin, overlooking the fact that differences
among actin isoforms concentrate at the N-terminus, which is also acetylated and forms part of the myosin-
binding site. Aim-1c will explore the role of actin N-acetylation and profilin/actin isoform variations on
formin function. Formins are the most important actin filament elongation factors in cells, and this group has
found that actin N-acetylation has a profound effect on filament elongation. It is further postulated that actin
isoform variations that concentrate at the N-terminus will have an equally strong effect on formin activity.
Aim-2 proposes new strategies to tackle persisting questions in the area of actin nucleation, which constitutes a
long-standing interest of this lab. Aim-2a will study in parallel Tmods and Lmods, which despite having a
common fold have evolved different activities – pointed-end capping and nucleation, respectively. The source
of the functional differences between these two subfamilies will be studied with a focus on Lmod3, implicated
in nemaline myopathy. An innovative strategy is proposed to determine the structure of Tmod at the pointed
end. It is finally investigated, in vitro and in cells, how specific Tmod and Tropomyosin isoforms interact with
each other to assemble morphologically and functionally distinct actin networks. Aim-2b will tackle the long-
standing problem of how WASP-family Nucleation Promoting Factors interact with and activate branch
formation by the Arp2/3 complex. The plans build upon the ability to perform biochemical and structural
studies on the baculovirus-expressed Arp2/3 complex and subcomplexes. Extensive preliminary and
published work provide the scientific premise and support feasibility.

## Key facts

- **NIH application ID:** 10116413
- **Project number:** 5R01GM073791-16
- **Recipient organization:** UNIVERSITY OF PENNSYLVANIA
- **Principal Investigator:** ROBERTO DOMINGUEZ
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $360,364
- **Award type:** 5
- **Project period:** 2005-04-01 → 2023-02-28

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10116413

## Citation

> US National Institutes of Health, RePORTER application 10116413, Structural-Functional Basis of Actin Cytoskeleton Dynamics (5R01GM073791-16). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/10116413. Licensed CC0.

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