Cell motility plays a central role in a wide variety of normal and pathological processes. Examples of cell motility in the nervous system include the movement of growth cones and the migration of neuroblasts and glial cells that occur in development. A striking example of how motility leads to disease can be seen in malignant gliomas. Cells from these tumors migrate considerable distances from the primary site, sometimes crossing from one cerebral hemisphere to the other. It is likely that motility is important in making these tumors so invasive and difficult to treat. There is a growing body of evidence that two cytoskeletal proteins--actin and myosin--compose the motor that drives motility. Myosins can be divided into two groups, called myosins I and II. These two forms of myosin are found in different regions of motile cells, and they clearly have different roles. These different functions must be reflected in differences in enzymology, structure, and mechanism of regulation. Data to be presented in this application will show that myosin I and II isoforms are present in several primary CNS malignancies, including gliomas. I will propose in this application to first expand on this observation by screening a wider variety of glioma cell lines as well as developing, embryonic brain in order to see if both myosin isoforms are present. I will also examine the intracellular distribution of these isoforms and their response to motility-stimulating growth factors. Second, I will isolate myosins I and II and measure the rate and equilibrium constants that describe their interactions with actin and ATP. Finally, I will examine the unique structural aspects of myosin I in detail, using a variety of biochemical and biophysical techniques. These latter experiments will use myosins I and II from Acanthameoba castellanii, a unicellular motile organism, as this source can provide quantities of protein large enough to perform the measurements that I propose. The high degree of sequence homology between Acanthamoeba myosins I and II and their vertebrate counterparts means that these experiments should allow me to make valid conclusions about CNS and glioma myosins. Results of these studies will be used to construct a detailed molecular model of how the actin-myosin interaction produces motility. These studies may ultimately allow the development of pharmacologic modulators of cell motility that could alter the biological aggressiveness of malignant glial tumors.
| Rosenfeld, S S; Rener, B (1994) The GPQ-rich segment of Dictyostelium myosin IB contains an actin binding site. Biochemistry 33:2322-8 |
