SHORT DISORDERED PEPTIDES ARE SUFFICIENT TO CONVERT PROTEINS INTO MECHANOSENSORS THAT RESPOND TO PHYSIOLOGICAL CELLULAR FORCES

Mechanical forces regulate many biological processes and a handful of mechanosensor domains have been identified in proteins that respond to cellular forces. The Notch signaling pathway, a key regulator of cell communication processes ranging from animal development to cancer, is activated by mechanical forces generated by cell-cell interactions. The mechanosensing element of Notch is thought to be a 300 amino acid extracellular domain called negative regulatory region (NRR). Ligand binding exerts mechanical forces that are believed to partially unfold NRR, rendering it susceptible to cleavage by extracellular metalloproteases that leads to activation of signaling. However, some engineered notch-like receptors lacking the NRR can be activated upon ligand binding, although the mechanisms underlying this activation are not known. Here we observe that Notch molecules without the NRR, but with a 12 amino acid sequence present in the extracellular juxta-transmembrane domain (eJTMD) are also cleaved and activated upon interaction with their ligands in cultured cells and in transgenic animals. Furthermore, the ∼12 aa eJTMD from notch genes from other species (chicken, xenopus, zebrafish and Drosophila ), from other unrelated transmembrane proteins (erbB4, N-CAM, and E-Cadherin), or multiple artificial sequences of ∼12 amino acids without any apparent structure or specific sequence can also convert proteins into ligand-dependent mechanosensors. These results indicate that short, disordered amino acid sequences that are commonly found in many proteins are capable of imparting mechanosensing capabilities into proteins, suggesting that mechanical force may regulate many more cellular processes than previously suspected.