ABSTRACT
Major efforts are underway to harness motor proteins for technical applications. Yet how to best attach cargo to microtubules that serve as kinesin-driven "molecular shuttles" without compromising transport performance remains challenging. Furthermore, microtubule-associated proteins (MAPs) can block motor protein-powered transport in neurons, which can lead to neurodegenerative diseases. Again it is unclear how different physical roadblock parameters interfere with the stepping motion of kinesins. Here, we employ a series of MAPs, tailored (strept)avidins, and DNA as model roadblocks and determine how their geometrical, nanomechanical, and electrochemical properties can reduce kinesin-mediated transport. Our results provide insights into kinesin transport regulation and might facilitate the choice of appropriate cargo linkers for motor protein-driven transport devices.
Subject(s)
Avidin/metabolism , DNA/metabolism , Kinesins/antagonists & inhibitors , Microtubule-Associated Proteins/metabolism , Microtubules/metabolism , Streptavidin/metabolism , Avidin/chemistry , DNA/chemistry , Electrochemical Techniques , Kinesins/chemistry , Kinesins/metabolism , Mechanical Phenomena , Microtubule-Associated Proteins/chemistry , Microtubules/chemistry , Streptavidin/chemistryABSTRACT
Transforming growth factor beta (TGFbeta) promotes epithelial cell differentiation but induces Schwann cell proliferation. We show that the protooncogene Ski (Sloan-Kettering viral oncogene homologue) is an important regulator of these effects. TGFbeta down-regulates Ski in epithelial cells but not in Schwann cells. In Schwann cells but not in epithelial cells, retinoblastoma protein (Rb) is up-regulated by TGFbeta. Additionally, both Ski and Rb move to the cytoplasm, where they partially colocalize. In vivo, Ski and phospho-Rb (pRb) appear to interact in the Schwann cell cytoplasm of developing sciatic nerves. Ski overexpression induces Rb hyperphosphorylation, proliferation, and colocalization of both proteins in Schwann cell and epithelial cell cytoplasms independently of TGFbeta treatment. Conversely, Ski knockdown in Schwann cells blocks TGFbeta-induced proliferation and pRb cytoplasmic relocalization. Our findings reveal a critical function of fine-tuned Ski levels in the control of TGFbeta effects on the cell cycle and suggest that at least a part of Ski regulatory effects on TGFbeta-induced proliferation of Schwann cells is caused by its concerted action with Rb.
