Vinculin Tension and YAP Activation are Altered in Dystrophic Muscle Cells

Abstract

Duchenne muscular dystrophy is a lethal muscle wasting disease caused by the absence of dystrophin, a protein that links the actin cytoskeleton to the extracellular matrix. Cells generate, sense, and respond to forces via mechanosensitive protein complexes, and altered force transmission via these complexes such as focal adhesions (FAs) are known to contribute to the development and progression of muscular dystrophy. However, the interplay between loss of dystrophin and altered force transmission via FAs remains unknown. Here we show that FAs of dystrophin deficient muscle cells are under less mechanical load and have decreased activation of yes-associated protein 1 (YAP). Using a vinculin bioluminescent tension sensor, we measured FAs tension in transgenic muscle cells expressing wild type (WT) dystrophin, two patient derived dystrophin missense mutants (L172H and L54R), and a non-disease-causing mutation (I232M). We found that cells harboring WT dystrophin showed higher average tension across vinculin compared to the tension in dystrophic cells. I232M vinculin tension was the same as WT tension. YAP is a known FA mechano-target protein that functions in the nucleus as a transcriptional co-activator and regulates several cellular processes including cellular migration. Dystrophic cells had higher levels of cytoplasmic YAP when compared to nuclear YAP, increased YAP S127 inactivating phosphorylation and decreased migration rates, compared to WT and I23M. Our results suggest that the Hippo pathway may regulate YAP activation via LATS1/2 activity. LATS1/2 phosphorylates YAP in S172 sequestering it in the cytoplasm and preventing its translocation to the nucleus, decreasing the expression of YAP target genes in migration. Altogether, our results show that expression of disease-causing mutant dystrophins leads to decreased tension transmission and aberrant mechanotransduction via FAs. We propose that decreased vinculin tension leads to increased Hippo pathway signaling which causes decreased YAP activation that results in impaired migration. Therefore, our cell lines could be used as a platform to characterize the effectiveness of DMD therapies for patients with missense mutations by means of monitoring YAP and migration. Furthermore, they could also be used to explore novel therapeutic targets in the Hippo pathway.