Winfield S. Sale
Professor Ph.D. 1977, University of California, Berkeley

Department of Cell Biology
Emory University School of Medicine
1648 Pierce Dr.
Atlanta, Georgia 30322

Phone: 404/727-6265 FAX: 404/727-6256 Email: win@cellbio.emory.edu

People:

Laura Fox
Pinfen Yang
Anne Roush Gaillard
Kristin W. Millions
Rip Finst
Ravi Bodiwala
Farah Bader

Research Support:

Work is supported by grants from the National Institutes of Health and the March of Dimes.

Research:

[1] Control of the Molecular Motor Dynein by Phosphorylation

[2] Signal Transduction, Protein Scaffolds and Anchoring of Kinases and Phosphatases that Control Motility

Our research focuses on two generally important issues: [1] the mechanisms and regulation of the molecular motor dynein; and [2] the signal transduction pathway that controls dynein, with particular emphasis on molecular mechanisms that target and anchor protein kinases and phosphatases in position to selectively control motor activity.  The significance of the work is founded on the universal importance of the molecular motor dynein in vital cell functions and development.  The significance is also founded on importance of precise targeting of signaling molecules including protein kinases and phosphatases for control of cell function.  Genetic, molecular, biochemical, functional and structural approaches are applied.
 
Dyneins are a family of ATPases responsible for ciliary and flagellar motility, retrograde cytoplasmic transport of organelles, assembly of the Golgi and mitotic apparatus during the cell division cycle and function of the mitotic spindle.  Among the most important questions are included: How do molecular motors generate force?  How are molecular motors targeted and anchored to specific cellular cargo?  How is motor activity regulated?  What are the signal transduction pathways and mechanisms that control molecular motor activity?

We are focused particularly on regulation of dynein by phosphorylation, and the molecular mechanism of anchoring the kinases and phosphatases that regulate dynein phosphorylation and activity.  To address these questions, we take advantage of the experimental strengths of Chlamydomonas and its paired flagella  (see Figure 1).  Chlamydomonas offers genetic, biochemical, molecular, and structural advantages for study of dynein.  Moreover, the flagellar axonemes (Figure 2) can be characterized by genetics and conveniently isolated for biochemical analysis and “reactivation” studies: addition of MgATP results in in vitro reactivation of motility.
 

Figure 1.  Phase contrast light-micrograph of Chlamydomonas reinhardtii taken by Steve L’Hernault of Emory University.

Figure 2.  Cross-section diagram of a Chlamydomonas flagellar axoneme illustrating selected structures and a few of the mutations used to define structures and their functions.

 Moreover, Chlamydomonas mutants are critical for identifying conserved components of the axoneme and dyneins, and offer a new opportunity to define scaffolds that anchor regulatory enzymes, including kinases, phosphatases and calcium binding proteins, in positions to control motility.  For example, the axoneme provides an ideal experimental system for study of targeted assembly of dynein motors since the axoneme contains several dyneins isoforms, each localized to distinct positions (Smith and Sale, 1992; Yang and Sale, 1998; Porter and Sale, 2000). The axoneme also offers an ideal and fresh system for study of the targeting and anchoring highly conserved enzymes such as PKA (Howard et al., 1994; Roush Gaillard et al., 2001), CK1 (Yang and Sale, 2000) and PP1 and PP2A (Habermacher and Sale, 1996; 1997; Yang et al., 2000; Porter and Sale, 2000 and Figure 3).

Figure 3.  The 9+2 axoneme is a highly conserved scaffold that localizes conserved kinases and phosphatases in positions to control motility.  The diagram summarizes many studies, providing a new picture of axonemal chemistry and illustrating the power of the axoneme, and Chlamydomonas, for defining molecules and mechanisms responsible for anchoring common enzymes in the cell (adapted from Porter and Sale, 2000).  

Selected Papers:

Piperno, G., Z. Ramanis, W. S. Sale, and E. Smith. 1990.  Three distinct inner dynein arms of Chlamydomas flagella:  molecular composition and location in the axoneme.  J. Cell Biol. 110: 379?389.

Smith, E. F. and W. S. Sale. 1991.  Microtubule binding and translocation by inner dynein arm subtype I1. Cell Motility and the Cytoskel. 18: 258-268.

Smith, E.F. and W.S. Sale. 1992.  Structural and functional reconstitution of inner dynein arms in Chlamydomonas flagellar axonemes, J. Cell Biol. 117:573-581.

Smith, E. F. and W. S. Sale. 1992. Regulation of dynein driven microtubule sliding by the radial spokes in flagella. Science 257: 1557-1559.

Moss, A. G., W. S. Sale, L. A. Fox, and G. B. Witman. 1992. The a-subunit of sea urchin sperm outer arm dynein mediates structural and rigor binding to microtubules. J. Cell Biol. 118: 1189-1200.

Sale, W. S., L. A. Fox, and E. F. Smith. 1993. Assays of axonemal dynein driven motility. In: Methods in Cell Biology.Jon Scholey, ed.  Academic Press.

Smith, E.F. and W.S. Sale. 1993. Molecular Basis for eukaryotic flagellar motility. In: Microtubules.  J. S. Hyams and C. Lloyd, eds. Wiley-Liss.

Fox, L. A., K. Sawin, and W. S. Sale.  1994.  Kinesin-related proteins in eukaryotic flagella. J. Cell Sci. 107:1545-550.

Howard, D., G. Habermaker, D. Glass, E.F. Smith and W.S. Sale.  1994.  Regulation of Chlamydomonas flagellar dynein by an axonemal kinase.  J. Cell Biol.  127:1683-1692.

Sale, W.S. and D.R. Howard.  1995.  Microscopic assays of flagellar dynein activity.  Meth. Cell Biology 47:257-262.

Howard, D.R. and W.S. Sale.  1995.  Isolation of inner and outer arm dyneins.  Meth. Cell Biology 47:481-486.

Habermacher, G. and W.S. Sale.  1995.  Regulation of dynein driven microtubule sliding by an axonemal kinase and phosphatase in Chlamydomonas flagella.  Cell Motility and the Cytosk.  32:106-109.

Sale, W. S. and L. A. Fox. 1996.  Microtubule translocation and rotation are general properties of dynein ATPases (in preparation).

Habermacher, G. and W.S. Sale.  1996.  Regulation of Flagellar Dynein by an Axonemal Type-1 Phosphatase in Chlamydomonas.  Journal of Cell Science 109:1899-1907.

Habermacher, G. and W.S. Sale. 1997.  Regulation of flagelar dynein by phosphorylation o a 138kDa dynein intermediate chain  J. Cell Biol. 136:167-176.

Yang, P., and W.S. Sale. 1998.  The 140,000 Mr Intermediate Chain of Chlamydomonas flagella inner arm dynein is a WD-repeat protein implicated in dynein arm anchoring.  Mol. Biol. Cell. 9:3335-3349.

Perrone, D., P. Yang, E. O'Toole, W. Sale, M. Porter. 1998.  The Chlamydomonas IDA7 locus encodes a 140-kDa dynein intermediate chain required to assemble the I1 inner arm complex.  Mol. Cell Biol. 9:3351-3365.

Mitchell, D. and W.S. Sale.  1999.  Characterization of a Chlamydomonas insertional mutant that disrupts flagellar central pair microtubule structures.  J. Cell Biol. 144:293-304.

Yang, P., L. Fox, R. Colbran, and W.S. Sale.  2000.  Protein phosphatases PP1 and PP2A are anchored in distinct locations in the flagellar axoneme. J. Cell Sci. 113:91-102.

Yang, P., and W.S. Sale.  2000.  CK1 is anchored on axonemal doublet microtubules and regulates dynein phosphorylation and activity.  J. Biol. Chem. 275:18905-18912.

Porter, M. and W. S. Sale.  2000.  The 9+2 axoneme anchors multiple dyneins and a network of kinases and phosphatases that control motility.  J. Cell Biol. (Nov 2000)

Roush, A., D. Diener, J. Rosenbaum and W.S. Sale.  2001.  Radial spoke protein 3 (RSP3) is an AKAP.   (In Press: J. Cell Biol.)

Yang, P., D. Diener, J. Rosenbaum and W. Sale 2001. Localization of calmodulin and dynein light chain LC8 in  flagellar radial spokes.  Submitted to JCB

Positions Available:

Postdoctoral fellow(s).

Anyone interested in doing postdoctoral research in the lab should contact Win Sale. Be prepared to supply a curriculum vitae, statement of research interests, and three people who can write letters on your behalf.

PhD student(s).

Any student who is interested in pursuing their thesis research in the lab who is in good standing in any of the PhD programs in bio/medical sciences at Emory University should contact Win Sale. Anyone interested in applying to the Emory University Graduate Division of Biological and Biomedical Sciences should follow this link.

Undergraduates.

Any undergraduate student at Emory (or any other university) who is interested in spending their summer in the lab, in an independent research project or work-study job should contact Win Sale. Be prepared to apply a curriculum vitae, statement of research interests, and three people who can write letters on your behalf.