# Cell shape and maturation impacts α-actinin-2 tension in iPSC-derived cardiomyocytes

## TL;DR

This study shows how cell shape and maturation affect the tension on a key protein in heart muscle cells, which is important for understanding heart function.

## Contribution

The study introduces a FRET-based sensor to measure tension on α-actinin-2 in cardiomyocytes and reveals how cell shape and maturation influence this tension.

## Key findings

- α-actinin-2 tension increases in rectangular-shaped cardiomyocytes after 5 days of culture.
- Cell shape and maturation modulate tension on α-actinin-2, a load-bearing sarcomeric protein.
- FRET index spread increases in both cell shapes after 5 days, indicating greater variability in tension.

## Abstract

The contractile activity of cardiomyocytes (CMs) critical to heart function emerges from the collective shortening of sarcomeres. However, how these sarcomeric forces are transmitted within CMs during this process remains poorly understood. Traction force microscopy has been used to measure overall forces exerted by CMs, but it falls short in providing insights into which specific proteins within sarcomeres transmit and whether cell shape influences forces within each sarcomere. Here, we aimed to characterize force generation on α-actinin-2, a z-disk protein that crosslinks anti-parallel actin filaments from adjacent sarcomeres and transmits force within a cell. By incorporating a Förster resonance energy transfer (FRET)-based molecular tension sensor in α-actinin-2, we measured contraction-induced deformation of the α-actinin-2 sensor in human-induced pluripotent stem cell-derived cardiomyocytes cultured on rectangular and circular adhesive patterns. We observed α-actinin-2 localized within sarcomeres, and actinin-2 loading correlated with sarcomere maturation and organization. α-actinin-2 tension increased in contracting rectangular-shaped cells, but not in circular cells. Moreover, the increase in tension was only observed in rectangular cardiomyocytes that were cultured for 5 days, and not after 24 h. Interestingly, the spread of FRET index values was increased in both rectangular and circular cells after 5 days in culture, compared to cells that were kept for 24 h in culture. Together, these data suggest that cell shape and maturation modulates tension on a load-bearing sarcomeric protein, α-actinin-2, and highlights the importance of characterizing tension across sarcomeric structures to understand cardiomyocyte contractile activity.