Modulation of mechanical tissue properties resulting from changes in splice variant expression of the Drosophila titin ortholog, sallimus
Authors: Tierney Patrick , Krajewski Dylan , Bonacker Kimberly , Goodreau Jordan , Kinney Shannon , Krans Jacob 
Published: June 2021

We are broadly interested in the many nuances of neuromuscular transduction. Work performed over the past two decades has demonstrated important non‐acto‐myosin means of modulating muscle stiffness ‐ elasticity and viscosity ‐ that can be (a) relatively rapid and long lasting, whilst (b) remaining somewhat independent of neuronal signaling. We hope to provide mechanistic explanations for historical phenomena like catch tension, the Blashko effect, and generally, muscle hysteresis. Here we report on the potential mechanical role(s) of alternative splice variants of the sallimus gene / protein in muscle tissue. Sallimus (sls) is an ortholog of vertebrate titin and like it's counterparts across Animalia (e.g. twitchin), sls possesses both compliant (i.e. Ig) and stiffer elastic domains (i.e. PEVK). There are at least ten sls isoforms described in fruit fly and several possess notably different proportions of compliant and stiff domains, potentially offering a diversity of mechanical tissue properties that could be critical to developmentally shifting motor demands. Indeed, Drosophila larvae must escape the dense, thick rinds of fruit (e.g. citrus), but have only hydrostatic soft bodies with which to do so. Stiffening the muscle could be achieved by either (a) increased neuronal output, or (b) modulation of splice variants / sls proteins. We hypothesize that modulation of sls variants, either by short‐term signalling (i.e. Ca2+) or longer‐term modified splice variant expression, could yield stiffness changes in a manner that mitigates the need to modulate neuronal output. In such a system, varied developmental motor demands could be accommodated more so by intramuscular protein modulation than motoneuronal output.We use two approaches to evaluate this hypothesis: (1) molecular and (2) physiologic / mechanical. We characterize the expression of different sls variants using several molecular approaches: qPCR, Western blots, and super resolution immunohistochemistry. Tissue mechanics are examined using a custom voice coil actuator, silicon based force beams, and motoneuronal electrical activation. Longitudinal bodywall muscles of larval Drosophila were stretched cyclically within previously described behavioral parameters. Motoneuronal stimuli were presented concurrent with various portions of the tissue length duty cycle. In some experiments, blebbistatin derivatives were utilized to interrupt acto‐myosin interactions. Tissue mechanics data suggest that short‐term modulation, independent of acto‐myosin interactions, has significant and long‐lasting effects on muscle stiffness. We used a transient anti‐sls RNAi paradigm to further evaluate our hypothesis. Interestingly, rather than knocking down all sls splice variants, molecular data suggest that transient RNAi against sls reduced overall sls expression but also induced the expression of different variants. Such a shift in variant expression, perhaps driven by neurohormonal modulation like the pre‐puparian octopaminergic surge, could be physiologically utilized to increase bodywall stiffness to a value greater than that of the fruit rind that a larva must escape.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.