An Isoform of the Giant Protein Titin is a Master Regulator of Human T Lymphocyte Trafficking

An Isoform of the Giant Protein Titin is a Master Regulator of Human T Lymphocyte Trafficking

During migration, circulating T lymphocytes encounter a multitude of environmental cues, forces, and cell deformations, which are processed by mechano-transduction mechanisms to elicit a cellular response.  Toffali et al. sought to identify key regulatory proteins that control these mechano-transduction mechanisms to regulate T lymphocyte trafficking.  They utilized mass spectrometry profiling and interactome studies to identify the giant titan protein as a potential target, which they validated with RNA sequencing analysis.  Interestingly they found three isoforms of the titan protein that were actively expressed in T lymphocytes.  These findings were validated with extensive flow cytometry analysis.  To further investigate the role of the first isoform of titan, LTTN1, they silenced the protein with a pool of four siRNAs which depleted the levels of the protein by 40%.  Surprisingly, this depletion of LTTN1 was sufficient to impair adhesive interactions at physiologic shear stress.  This impairment was attributed to collapsed microvilli in these T lymphocytes, which they determined using scanning electron microscopy studies. To investigate the mechanism by which LTTN1 affects microvilli morphology the group looked at several mechanisms, and determined that while it did not affect the phosphorylation status of ERM proteins, it did affect chemokine activation of integrins as LTTN1 silencing inhibited CXCL12-induced adhesion of LFA-1 to ICAM-1.  Importantly, LTTN1 silencing also suppressed CXCL12 activation of the small GTPases RhoA and Rac1 which the group hypothesized was contributing to the microvilli morphological changes.  LTTN1 was also shown to play a role in T lymphocyte polarization, migration, and resilience to passive cell deformation. Cytoskeleton Inc’s RhoA (Cat. # BK124) and Rac1 (Cat. # BK128) G-LISA activation assays were used to analyze the level of small G-protein activation in these T lymphocyte studies.