Margaret Titus, PhD

Research Instersts:

The unconventional myosins are a superfamily of at least fifteen distinct classes of actin-based motors whose functions range from organelle transport to generating forces required for cellular migration. The myosins all share a common motor domain, but each class of myosin has a distinct tail domain. Several of the classes of unconventional myosins, specifically the VI, VII and XV myosins, have been implicated in neurosensory disorders in humans and mice, but the molecular basis of their function remains unknown. The research in the laboratory is focussed on several questions: why do cells express so many myosins? what is the function of each individual class of myosin? do the properties of the tail domain direct the function of a given class of myosin? The cellular function of several classes of unconventional myosins in both the simple eukaryote Dictyostelium discoideum and the nematode Caenorhabditis elegans is being studied through a combined approach that includes genetic, molecular genetic, cell biological and biochemical techniques.

The lab's recent studies revealed that the class I myosins play a role in endocytosis. Dictyostelium myosin I mutants exhibit decreased endocytic activity and produce abnormal numbers of actin-rich membrane extensions. The working hypothesis is that the class I myosins play a role in the manipulation of the actin-rich cortex underlying the plasma membranes. A complementation approach is being employed to define regions of the myosin I important for this function, with a specific emphasis on the tail region. Molecular techniques are being employed to generate deletion mutants and re-express these in mutant cells to determine if they rescue mutant phenotypes and if they alter the subcellular localization of myosin I.

The laboratory has recently identified a Dictyostelium class VII myosin and analyzed its function by gene targeting. Interestingly, myosin VII mutants have a specific, and severe, defect in phagocytosis. Future experiments will be directed at identifying the step in phagocytosis that requires myosin VII through videomicroscopic analysis and immunolocalization. Complementation experiments will be carried out to rescue the phagocytosis defect with GFP tagged versions of myosin VII to follow the dynamics of myosin VII localization during phagocytosis. Regions of the tail domain required for function and/or localization of this myosin will be identified using a combined mutagenesis and complementation strategy.

Another goal of the lab's studies is to understand the role of unconventional myosins in development and in neurosensory function. A reverse genetic approach, employing a combined chemical mutagenesis and PCR strategy is being employed to obtain C. elegans lacking either the class VI or VII unconventional myosins. Mutant phenotypes will be analyzed in combination with immunolocalization studies. Future work will employ genetic methods to identify functionally important regions of either myosin VI or VII and identify interacting molecules. Together, studies of unconventional myosins in two diverse, yet complementary, systems will allow the lab to understand how cells use unconventional myosins to generate a range of actin-based motile behaviors.

Selected Publications:

Klein, J.C., Burr, A.R., Svensson, B., Kennedy, D.J., Titus, M.A., Rayment, I.,& Thomas, D.D.  (2008).  Actin-binding cleft closure in myosin II probed by site-directed spin labeling and pulsed EPR. P.N.A.S.  105:12867-12872

Agafonov, R.V, Titus, M.A., Thomas, D.D. & Nesmelov, Y.E.  (2008).  Muscle and nonmuscle myosins probed by a spin label at equivalent sites in the force-generating domain.  P.N.A.S.  105:13397-13402

Wang Q., Deloia M.A., Kang Y., Litchke C., Zhang, N., Titus M.A., & Walters, K.J.  (2007). The SH3 domain of a M7 interacts with its C-terminal proline rich region.  Prot. Sci.  16:189-196

Mini-Review - Titus, M.A.  (2006).  Myosin I and actin dynamics:  the frogs weigh in. Dev. Cell 11:594-595

Titus MA (2005) An unexpected role for myosins in adhesion. Novartis Foundation Symposium Proceedings, in press.

Titus MA (2004) Myosins meet microtubules. Nature 431:252-253.

Titus MA (2004) The role of talin and myosin VII in adhesion - A FERM connection. in: Cell Motility. From molecules to organisms., A. Ridley, M Peckham & P. Clark, eds. pp 19-37.

Cooper JA and Titus MA. co-editors, Curr.Op. Cell Biol. - Cell Structure & Dynamics Issue for 2004 (volume 16, pages 1-112).

Falk DL, Wessels D, Jenkins L, Pham T, Kuhl S, Titus MS, and Soll DR (2003) Shared, unique and redundant functions of three myosin Is (myoA, myoB and myoF) in motility and chemotaxis in Dictyostelium. J. Cell Sci. 116:3985-3999

Senda, S., Lee, S.-F., Côté, G.P. & Titus, M.A. (2001). Recruitment of a specific amoeboid myosin I isoform to the plasma membrane in chemotactic Dictyostelium cells. J. Biol. Chem. 276:2898-2904.

Gliksman, N.R., Santoyo, G., Novak, K.D. & Titus, M.A. (2001). In vivo phosphorylation of myosin I is increased by chemotactic stimulation. J. Biol. Chem. 276: 5235-5239.

Tuxworth, R.I., Weber, I., Wessels, D., Addicks, G.C. Gerisch, G., Soll, D.R. & Titus, M.A. (2001). A role for myosin VII in dynamic cell adhesion. Curr Biol. 11:318-329.

Senda, S. & Titus, M.A. (2000). A potential mechanism for regulating myosin I binding to membranes in vivo. FEBS Letters. 484:125-128.

Kelleher, J.F., Mandell, M.A., Moulder, G., Hill, K.L., L'Hernault, S.W., Barstead, R. & Titus, M.A. (2000). Myosin VI is required for the asymmetric segregation of cellular components during C. elegans spermatogenesis. Curr. Biol. 10:1489-1496.

Dai, J., Ting-Beall, P., Hochmuth, R., Sheetz, M.P. & Titus, M.A. (1999). Myosin I contributes to the generation of resting cortical tension. Biophys J. 77:1168-1176.

Titus, M.A. (1999). A class VII unconventional myosin is required for phagocytosis. Curr. Biol. 9:1297-1303.

 

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