טכניון מכון טכנולוגי לישראל
הטכניון מכון טכנולוגי לישראל - בית הספר ללימודי מוסמכים  
Ph.D Thesis
Ph.D StudentBoris Shneyer
SubjectNew Function for the Actin-Based Molecular Motor
Myo19 in Mitochondrial Motility and Filopodia
Formation
DepartmentDepartment of Biology
Supervisor Professor Henn Arnon
Full Thesis textFull thesis text - English Version


Abstract

Eukaryotic cells are exposed to many environmental cues and stress conditions which have profound effects on the physiology of the cells and specifically on the dynamics of the mitochondrial network. This is apparent in the adaptation of the mitochondrial network by regulating its fusion -fission balance and moving to distinct regions within the cells. The importance of correct mitochondrial homeostasis and distribution is underlined by the findings that its disruption is implicated in neurodegenerative diseases. The dynamics and distribution of the mitochondrial network are mainly based on microtubules, whereas the contribution of the actin cytoskeleton is only recently beginning to be appreciated, revealing new functions for actin-based motors. Myo19 is an actin-based motor that is associated with the mitochondria and affects its motility when ectopically expressed. However, how Myo19 is associated with the mitochondria, what are the extracellular cues that affect Myo19 activity, what is the function of Myo19 in mitochondrial dynamics and distribution, and how it is enzymatically adapted to its function are unknown. We show that Myo19 is stably anchored to the outer mitochondrial membrane (OMM), being the only known myosin to directly interact with a membrane, suggesting that it is regulated whilst bound to the membrane. Myo19 exhibited a dramatic effect on mitochondrial dynamics in response to starvation stress, promoting their localization to starvation-induced filopodia. By tracing the effect that glucose-starvation exerts on the cells we found that Myo19 promotes the localization of mitochondria to filopodia in response to reactive oxygen species (ROS) and EGF. Time-lapse fluorescent microscopy reveals that ROS and EGF-induced Myo19 motility is a highly dynamic process which is coupled to filopodia elongation and retraction, exhibiting a complex pattern of intrafilopodial movements. Interestingly, Myo19 filopodial motility is inhibited by back-to-consensus-mutation of a unique residue of class XIX myosins in the motor domain, indicating that it is uniquely adapted to transport a large cargo as the mitochondria through the dense filopodia. RNAi mediated knockdown of Myo19 diminished the formation of the glucose-starvation induced filopodia without evident effects on the mitochondrial network, indicating that Myo19 promotes the formation of filopodia, most likely by the transport of mitochondria to their tips. Recent studies reveal that increased mitochondria density in neurons is correlated with increased branching of actin-based protrusions and dendrites, mainly by replenishing local pools of ATP. In accordance with these findings, we show that the function of mitochondria at EGF-induced filopodia is, at least in part, to provide localized ATP synthesis. In summary, our study demonstrates the contribution of actin-based motility to the mitochondrial localization to filopodia by specific cellular cues and stress conditions, and suggests that the function of the mitochondrial molecular motor is to transport mitochondria to sites that require localized ATP synthesis.