# Mechanisms of septin-actin cytoskeletal crosstalk

> **NIH NIH F32** · BRANDEIS UNIVERSITY · 2024 · $74,284

## Abstract

PROJECT SUMMARY
This proposal addresses how the cellular functions of two major cytoskeletal polymer systems, septins and actin,
are coordinated and influence each other. To address this question, I will use the budding yeast S. cerevisiae,
where septins were first discovered, and where I am readily able to combine a ‘bottom up’ in vitro reconstitution
and single molecule imaging approach with a ‘top down’ genetics and live imaging approach. The proposal builds
off of recent discoveries made in the Goode lab, which reveal that septins are organized at the yeast bud neck
into 8-10 evenly-spaced bars, or “pillars”, which co-align with F-actin cables used for intracellular transport. While
work in a number of model organisms has closely linked the in vivo functions of septins and actin, we still have
only a limited understanding of the molecular mechanisms underlying this septin-actin crosstalk. The goal of this
proposal is to define these mechanisms. S. cerevisiae express 5 different septin proteins, which co-polymerize
into filaments and are further organized into higher order structures. My preliminary data show that one of the
septins (Shs1) mediates direct binding to F-actin in vitro, and that loss of SHS1 disrupts actin cable architecture
and function in vivo. In Aim1, I will use targeted mutagenesis to generate new shs1 separation-of-function
mutants disrupting F-actin binding. I will then use these mutants to investigate how Shs1-mediated F-actin
binding contributes to the alignment of actin cables with septin pillars in vivo, and intracellular transport of
secretory vesicles. In Aim2, I will reconstitute purified septin oligomers and filaments decorated with actin-
nucleating formins (Bnr1) and formin-regulatory proteins (e.g., Gin4, Bud6, and the Mlc1-Iqg1-Hof1 complex),
and define their effects on F-actin assembly and organization by TIRF microscopy and single molecule imaging.
These experiments will test several important hypotheses, including: (1) whether septins and Gin4 activate full-
length Bnr1 from autoinhibition to promote actin assembly; (2) whether Iqg1 has regulatory effects on F-actin
and Bnr1, like its human counterpart IQGAP1 (based on a recent study from the Goode lab; Hoeprich et al.,
2022); and (3) whether septin oligomers/filaments themselves (via Shs1) directly influence F-actin bundling and
dynamics. In parallel to these in vitro experiments, I will acutely deplete the same proteins in vivo (using degron
tags) to determine how each contributes to the assembly and alignment of actin cables at the bud neck. My
preliminary data already point to an exciting new role for Iqg1 in controlling actin cable formation during polarized
cell growth. Together, the in vitro and in vivo work outlined in this proposal will: (i) clarify how septins, formins,
and formin-regulatory proteins work in concert to shape actin networks, (ii) define new subunit-specific roles for
septins in actin regulation, (iii) lay a strong foundation for lau...

## Key facts

- **NIH application ID:** 10843758
- **Project number:** 5F32GM150204-02
- **Recipient organization:** BRANDEIS UNIVERSITY
- **Principal Investigator:** Joseph O Magliozzi
- **Activity code:** F32 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $74,284
- **Award type:** 5
- **Project period:** 2023-06-01 → 2025-05-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10843758

## Citation

> US National Institutes of Health, RePORTER application 10843758, Mechanisms of septin-actin cytoskeletal crosstalk (5F32GM150204-02). Retrieved via AI Analytics 2026-05-25 from https://api.ai-analytics.org/grant/nih/10843758. Licensed CC0.

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