The human genome contains 8 different isoforms of myosin I, making it the largest family of unconventional myosins expressed in humans. Myosin Ic is perhaps the best studied myosin I isoform due to its proposed roles in dynamic adaptation in hair cells, insulin stimulated GLUT-4 transport, compensatory excocytosis, and nuclear transcription. Despite its association with these cellular processes and disease such as congenital deafness, the molecular role of myo1c in the cell is unknown. It has been proposed that myosin Ic may act as a transporter, a force generator, or a strain-sensing tether. The ability of myosin Ic to function in these roles depends on its abilities to generate force and to modulate its biochemistry in response to load, with each of these roles placing very different and specific mechanical requirements on the myosin. Thus, by understanding the mechanics of myosin Ic, the molecular role of myosin Ic can be deduced. Despite the fact that myosin mechanics are at the heart of its cellular function, very little is known about the mechanics and load dependent mechanochemistry of myosin Ic. Furthermore, it has been proposed that calcium binding to calmodulins on the myosin Ic regulatory domain may play a central role in modulating its mechanics and response to load;however this notion has never been tested experimentally. Also, it is unknown whether calcium induced changes in myosin Ic mechanics and kinetics are relevant to cellular function since the response of myosin Ic to transient increases in calcium has never been studied. This research will address these gaps in our knowledge. Using a battery of single molecule techniques, the specific aims of this research are: 1. Determine the mechanical properties of the myosin Ic working stroke 2. Determine the load sensitivity of myosin Ic 3. Directly measure the effects of transient calcium on myosin Ic mechanics and kinetics All of these experiments will be conducted in the presence and the absence of calcium to test how calcium affects myosin kinetics and mechanics. Besides elucidating the molecular role that myosin Ic plays in the cell and how this role is regulated by calcium and force, this research will also shed light on the diversity within the myosin I superfamily.

Public Health Relevance

Molecular motors within the human body are responsible for generating and responding to forces with malfunction of these motors leading to a wide array of diseases including deafness, hypertension, cardiac failure, and developmental defects. While the ability to respond to forces is central to the function of these motors, little is known about how this is accomplished. Using cutting-edge single molecule techniques, this research will examine force generation and tension sensing in of one of these motors, myosin Ic, helping us to understand both its cellular function and the role that tension sensing plays in molecular motors in both health and disease.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
Postdoctoral Individual National Research Service Award (F32)
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Special Emphasis Panel (ZRG1-F04B-D (20))
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Sakalian, Michael
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University of Pennsylvania
Schools of Medicine
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Shuman, Henry; Greenberg, Michael J; Zwolak, Adam et al. (2014) A vertebrate myosin-I structure reveals unique insights into myosin mechanochemical tuning. Proc Natl Acad Sci U S A 111:2116-21
Greenberg, Michael J; Ostap, E Michael (2013) Regulation and control of myosin-I by the motor and light chain-binding domains. Trends Cell Biol 23:81-9
Lewis, John H; Greenberg, Michael J; Laakso, Joseph M et al. (2012) Calcium regulation of myosin-I tension sensing. Biophys J 102:2799-807
Greenberg, Michael J; Lin, Tianming; Goldman, Yale E et al. (2012) Myosin IC generates power over a range of loads via a new tension-sensing mechanism. Proc Natl Acad Sci U S A 109:E2433-40