# Unadulterated expression of tropomyosin isoforms and their interactions with F-actin, tropomodulin and leiomodin

> **NIH NIH F31** · UNIVERSITY OF PENNSYLVANIA · 2021 · $33,410

## Abstract

Project summary/abstract:
The goal of this proposal is to uncover how tropomyosin (Tpm) isoforms regulate actin filament function and
how Tpm mutations cause muscle diseases such as cardiomyopathy, nemaline myopathy, and congenital
myopathy. Progress in this area has been stalled by the inability to obtain physiologically relevant Tpm
isoforms. Tpm isoforms play non-overlapping roles, and to function properly and self-associate head-to-tail
along actin filaments, they must carry native post-translational modifications (PTMs) and be free of extra amino
acids from purification tags. Traditional E. coli and yeast expression systems are not adequate in this case. In
a major breakthrough, I have developed a system to express Tpm isoforms in human cells that overcomes
these obstacles. In contrast to previous methods, my expressed Tpm isoforms (1) are N-terminally acetylated,
(2) do not contain extra tag amino acids at the N- or C- termini, and (3) form natural heterodimers with other
Tpm isoforms. In aim 1, I will optimize this expression method, including transfection efficiency and protein
yields, so that it can be readily adopted by other scientists. I will further characterize the native state of each
Tpm isoform, including the identification of natural heterodimer pairs and PTMs. In aim 2, I will address what
are the biochemical differences among Tpm isoforms that determine their specialized cellular roles. I will focus
on each isoform’s (or heterodimer’s) interactions with actin, and two regulatory proteins that must bind Tpm for
their functions: tropomodulin (Tmod, a pointed end capping protein) and leiomodin (Lmod, a filament
nucleator). In aim 3, I will study point mutations in Tpm responsible for muscle diseases, and address how
these mutations disrupted the biochemical properties of Tpm, including its interactions with actin filaments,
Tmod and Lmod. This knowledge should inform treatment strategies.
The success of my aims depends mainly on the ability to express native Tpm isoforms, which I demonstrate in
preliminary studies. My novel expression system uniquely positions me to address long-standing questions
about Tpm function, and can be readily adopted by researchers wishing to study proteins in their native state.
Accomplishment of the aims should have a far-reaching impact, providing both a biochemical understanding of
Tpm isoform-specific cellular roles and information regarding heart and muscle disease mutations. This
research will also serve to further my training in the fields of biochemistry, physiology, and molecular
biophysics.

## Key facts

- **NIH application ID:** 10140807
- **Project number:** 1F31HL156431-01
- **Recipient organization:** UNIVERSITY OF PENNSYLVANIA
- **Principal Investigator:** PETER J CARMAN
- **Activity code:** F31 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $33,410
- **Award type:** 1
- **Project period:** 2021-04-25 → 2023-02-15

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10140807

## Citation

> US National Institutes of Health, RePORTER application 10140807, Unadulterated expression of tropomyosin isoforms and their interactions with F-actin, tropomodulin and leiomodin (1F31HL156431-01). Retrieved via AI Analytics 2026-05-22 from https://api.ai-analytics.org/grant/nih/10140807. Licensed CC0.

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