# Regulating Microtubule Severing Physically and Chemically

> **NIH NIH R15** · SYRACUSE UNIVERSITY · 2022 · $13,249

## Abstract

PROJECT SUMMARY
The goal of this new proposal application is to uncover the physical and chemical regulatory
schemes to control the microtubule severing enzyme, katanin. Katanin is a AAA+ enzyme that
hexamerizes in order to remove tubulin diimers from the microtubule filament resulting in
filament severing. When at high levels, and unregulated in cells, katanin can destroy the entire
microtubule network, thus turning it off is an essential control knob. Overactivity of katanin can
lead to complete loss of microtubule polymer, but underactivity is linked with developmental
defects in the brain and ciliopathies. The central hypothesis of the proposed work is that the
mechanisms to control katanin actually regulate katanin hexamer oligomerization. Our prior
work indicated that oligomerization is the a rate-limiting step for katanin. We seek to use
quantitative fluorescence microscopy to directly test the hypothesis that oligomerization controls
severing through physical and chemical means. Specifically, we will explore the regulation of
katanin concentration in live cells using a novel light-sensitive dimerization domain to drive
katanin concentration locally by exploring the following aims: (1) Quantification of both the
katanin concentration and the microtubule filament density as a function of time will enable
biochemistry in the cell for the first time for katanin. (2) Using in vitro reconstitution of
microtubule severing and a novel single molecule counting technique, we will examine the effect
of the phosphorylation state of serine 131 on binding, oligomerization, and severing by katanin.
(3) The tubulin carboxy-terminal tail has been shown to act as a code to control many
microtubule-associated proteins and enzymes. Severing enzymes are no different and are
known to require the carboxy- terminal tail to sever microtubules. Our preliminary data shows
that katanin’s regulation is distinct from other severing enzymes. We will use a severing
inhibition assay and single molecule counting to quantify the ability to katanin to bind and act on
microtubules of various carboxy-terminal tail sequences.
Accomplishing the proposed aims will create a novel microtubule disruption tool that could be
used in a variety of cellular and organismal assays to control the location and density of the
microtubule network. Further, the proposed studies will reveal new information on how
microtubule severing can be regulated in cells through controlling the katanin oligomerization
state. This crucial step for katanin activity may be an entry-point for creating small molecule
inhibitors for microtubule severing enzymes and other AAA+ enzymes that are important for a
host of essential cellular functions including protein homeostasis, DNA recombination,
replication, and repair.

## Key facts

- **NIH application ID:** 10580392
- **Project number:** 3R15GM141722-01S1
- **Recipient organization:** SYRACUSE UNIVERSITY
- **Principal Investigator:** JENNIFER L ROSS
- **Activity code:** R15 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2022
- **Award amount:** $13,249
- **Award type:** 3
- **Project period:** 2021-05-01 → 2025-04-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10580392

## Citation

> US National Institutes of Health, RePORTER application 10580392, Regulating Microtubule Severing Physically and Chemically (3R15GM141722-01S1). Retrieved via AI Analytics 2026-05-24 from https://api.ai-analytics.org/grant/nih/10580392. Licensed CC0.

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