Biomedical Research Education & Training
Faculty Member

Graham, Todd R., Ph.D.
Professor of Biological Sciences
Professor of Cell and Developmental Biology

Lab Url: N/A

Phone Number: 615-343-1835

Email Address: tr.graham@vanderbilt.edu

Graham, Todd's picture
Academic history
BS, Maryville College, St. Louis, MO
PhD, St. Louis University, St. Louis, MO
postdoctoral training, CalTech, Pasadena, CA
postdoctoral training, UCSD, La Jolla, CA
N/A

Office Address   Mailing Address

2433 Stevenson Center

Department of Biological Sciences VU Station B Box 35-1634 37235-1634


Research Keywords
Protein transport, protein sorting, secretory pathway, endocytosis, Golgi complex, clathrin, ARF, yeast genetics, endocrinology, genetics, membrane biology, aminophospholipid translocase, Drs2, flippase, P4-ATPase,Yeast

Research Specialty
Protein transport and membrane biogenesis

Research Description
The research goals of the Graham laboratory are to understand the molecular mechanisms underpinning vesicle-mediated protein transport and membrane biogenesis. Most of our effort is focused on determining how type IV P-type ATPases (P4-ATPases) contribute to the establishment of membrane asymmetry and budding of transport vesicles from organelle membranes using the yeast model system.
The P4-ATPases flip specific phospholipid species, such as phosphatidylserine, from the extracellular leaflet of the plasma membrane to the cytosolic leaflet, thus producing an asymmetric membrane structure that is conserved among most eukaryotic cells. This phospholipid asymmetry has a major influence on the localization and activity of many different plasma membrane proteins. Moreover, regulated disruption of membrane asymmetry is a signaling device used in blood clotting reactions and for recognition of apoptotic cells. Humans have 14 P4-ATPases and members of this protein family are implicated in severe liver and neurological disease. Additionally, murine P4-ATPases are implicated in B-cell deficiency, obesity and type 2 diabetes, motor neuron degeneration, defects in bile secretion, and reduced male fertility. A current project in the laboratory is to define the mechanism of substrate recognition and translocation by P4-ATPases using molecular genetic and biochemical approaches. The best characterized P-type ATPases transport small cations across membranes to establish ion gradients and so phospholipid molecules are an unusual substrate for this protein family. Our work is suggesting a novel transport mechanism for the P4-ATPases and is providing insight into how these transporters evolved the ability to transport their "giant substrate".
In addition to establishing membrane asymmetry, we discovered that P4-ATPases play a crucial role in budding protein transport vesicles from Golgi and endosomal membranes. For example, a P4-ATPase called Drs2 translocates phosphatidylserine across the membrane of the trans-Golgi network and this flippase activity is required to bud AP-1/clathrin-coated vesicles that transport proteins from the Golgi to endosomes. Our work has uncovered both positive and negative regulators of Drs2 activity representing proteins and lipids known to have critical roles in vesicular transport. The ATP-powered, unidirectional translocation of phosphatidylserine to cytosolic leaflet should have a dramatic influence on the biophysical properties of the membrane; enhancing the anionic membrane potential of the cytosolic surface as well as inducing curvature through a bilayer couple mechanism. Another current project in the lab is to determine how these P4-ATPase effects on the membrane are coordinated with the vesicle budding machinery to sort and package cargo proteins into newly forming vesicles. The protein trafficking events dependent on P4-ATPase function have a major influence on the protein composition of the plasma membrane and organelles of the secretory and endocytic pathways.


Publications
Baldridge, RD, Graham, TR. Two-gate mechanism for phospholipid selection and transport by type IV P-type ATPases. Proc Natl Acad Sci U S A, 110(5), E358-67, 2013

Baldridge, RD, Xu, P, Graham, TR. Type IV P-type ATPases Distinguish Mono- versus Diacyl Phosphatidylserine Using a Cytofacial Exit Gate in the Membrane Domain. J Biol Chem, 288(27), 19516-27, 2013

Graham, TR. Arl1 gets into the membrane remodeling business with a flippase and ArfGEF. Proc Natl Acad Sci U S A, 110(8), 2691-2, 2013

Xu, P, Baldridge, RD, Chi, RJ, Burd, CG, Graham, TR. Phosphatidylserine flipping enhances membrane curvature and negative charge required for vesicular transport. J Cell Biol, 2013

Baldridge, RD, Graham, TR. Identification of residues defining phospholipid flippase substrate specificity of type IV P-type ATPases. Proc Natl Acad Sci U S A, 109(6), E290-8, 2012

Sebastian, TT, Baldridge, RD, Xu, P, Graham, TR. Phospholipid flippases: building asymmetric membranes and transport vesicles. Biochim Biophys Acta, 1821(8), 1068-77, 2012

Graham, TR, Burd, CG. Coordination of Golgi functions by phosphatidylinositol 4-kinases. Trends Cell Biol, 21(2), 113-21, 2011

Graham, TR, Kozlov, MM. Interplay of proteins and lipids in generating membrane curvature. Curr Opin Cell Biol, 22(4), 430-6, 2010

Muthusamy, BP, Natarajan, P, Zhou, X, Graham, TR. Linking phospholipid flippases to vesicle-mediated protein transport. Biochim Biophys Acta, 1791(7), 612-9, 2009

Muthusamy, BP, Raychaudhuri, S, Natarajan, P, Abe, F, Liu, K, Prinz, WA, Graham, TR. Control of protein and sterol trafficking by antagonistic activities of a type IV P-type ATPase and oxysterol binding protein homologue. Mol Biol Cell, 20(12), 2920-31, 2009 PMCID:2695799

Natarajan, P, Liu, K, Patil, DV, Sciorra, VA, Jackson, CL, Graham, TR. Regulation of a Golgi flippase by phosphoinositides and an ArfGEF. Nat Cell Biol, 11(12), 1421-6, 2009 PMCID:2787759

Zhou, X, Graham, TR. Reconstitution of phospholipid translocase activity with purified Drs2p, a type-IV P-type ATPase from budding yeast. Proc Natl Acad Sci U S A, 106(39), 16586-91, 2009 PMCID:2757829

Liu, K, Surendhran, K, Nothwehr, SF, Graham, TR. P4-ATPase Requirement for AP-1/Clathrin Function in Protein Transport from the trans-Golgi Network and Early Endosomes. Mol Biol Cell, 2008 PMCID:2488278

Liu, K, Hua, Z, Nepute, JA, Graham, TR. Yeast P4-ATPases Drs2p and Dnf1p Are Essential Cargos of the NPFXD/Sla1p Endocytic Pathway. Mol Biol Cell, 18, 487-500, 2007 PMCID:1783782

Chen, S, Wang, J, Muthusamy, BP, Liu, K, Zare, S, Andersen, RJ, Graham, TR. Roles for the Drs2p-Cdc50p complex in protein transport and phosphatidylserine asymmetry of the yeast plasma membrane. Traffic, 7(11), 1503-17, 2006

Natarajan, P, Graham, TR. Measuring translocation of fluorescent lipid derivatives across yeast Golgi membranes. Methods, 39(2), 163-8, 2006

Parsons, AB, Lopez, A, Givoni, IE, Williams, DE, Gray, CA, Porter, J, Chua, G, Sopko, R, Brost, RL, Ho, CH, Wang, J, Ketela, T, Brenner, C, Brill, JA, Fernandez, GE, Lorenz, TC, Payne, GS, Ishihara, S, Ohya, Y, Andrews, B, Hughes, TR, Frey, BJ, Graham, TR, Andersen, RJ, Boone, C. Exploring the mode-of-action of bioactive compounds by chemical-genetic profiling in yeast. Cell, 126(3), 611-25, 2006

Xiao, J, Kim, LS, Graham, TR. Dissection of Swa2p/auxilin domain requirements for cochaperoning Hsp70 clathrin-uncoating activity in vivo. Mol Biol Cell, 17(7), 3281-90, 2006 PMCID:1483056

Chantalat, S., S.-K. Park, Z. Hua, K. Liu, R. Gobin, A. Peyroche, A. Rambourg, T.R. Graham and C.L. Jackson. The Arf activator Gea2p and the P-type ATPase Drs2p interact at the Golgi in Saccharomyces cerevisiae. J Cell Sci, 117(Pt 5), 711-22, 2004

Chim, Nicholas, Gall, Walter E, Xiao, Jing, Harris, Mark P, Graham, Todd R, Krezel, Andrzej M. Solution structure of the ubiquitin-binding domain in Swa2p from Saccharomyces cerevisiae. Proteins, 54(4), 784-93, 2004

Graham, Todd R. Flippases and vesicle-mediated protein transport. Trends Cell Biol, 14(12), 670-7, 2004

Graham, Todd R. Membrane targeting: getting Arl to the Golgi. Curr Biol, 14(12), R483-5, 2004

Natarajan, Paramasivam, Wang, Jiyi, Hua, Zhaolin, Graham, Todd R. Drs2p-coupled aminophospholipid translocase activity in yeast Golgi membranes and relationship to in vivo function. Proc Natl Acad Sci U S A, 101(29), 10614-9, 2004 PMCID:489982

Hua, Zhaolin, Graham, Todd R. Requirement for neo1p in retrograde transport from the Golgi complex to the endoplasmic reticulum. Mol Biol Cell, 14(12), 4971-83, 2003 PMCID:284799

Gall, Walter E, Geething, Nathan C, Hua, Zhaolin, Ingram, Michael F, Liu, Ke, Chen, Sophie I, Graham, Todd R. Drs2p-dependent formation of exocytic clathrin-coated vesicles in vivo. Curr Biol, 12, 1623-7, 2002

Hua, Zhaolin, Fatheddin, Parvin, Graham, Todd R. An essential subfamily of Drs2p-related P-type ATPases is required for protein trafficking between Golgi complex and endosomal/vacuolar system. Mol Biol Cell, 13, 3162-77, 2002 PMCID:124150

Graham, TR. Metabolic labeling and immunoprecipitation of yeast proteins. Curr Protoc Cell Biol, Chapter 7, Unit 7.6, 2001

Brigance, W T, Barlowe, C, Graham, T R. Organization of the yeast Golgi complex into at least four functionally distinct compartments. Mol Biol Cell, 11, 171-82, 2000 PMCID:14766

Gall WE, MA Higginbotham, C-Y Chen, MF Ingram, DM Cyr, and TR Graham. The auxilin-like phosphoprotein Swa2p is required for clathrin function in yeast. Curr Biology , 10, 1394-1358, 2000

Gall, W E, Higginbotham, M A, Chen, C, Ingram, M F, Cyr, D M, Graham, T R. The auxilin-like phosphoprotein Swa2p is required for clathrin function in yeast. Curr Biol, 10(21), 1349-58, 2000

Hopkins, B D, Sato, K, Nakano, A, Graham, T R. Introduction of Kex2 cleavage sites in fusion proteins for monitoring localization and transport in yeast secretory pathway. Methods Enzymol, 327, 107-18, 2000

Chen, C Y, Ingram, M F, Rosal, P H, Graham, T R. Role for Drs2p, a P-type ATPase and potential aminophospholipid translocase, in yeast late Golgi function. J Cell Biol, 147(6), 1223-36, 1999 PMCID:2168089

Chen, C.-Y., M.F. Ingram, P. Rosal, and T.R. Graham. Role for Drs2p, a P-type ATPase and potential aminophospholipid translocase, in yeast late Golgi function. J. Cell Biol. , 147, 1223-1236, 1999

Chen, C Y, Graham, T R. An arf1Delta synthetic lethal screen identifies a new clathrin heavy chain conditional allele that perturbs vacuolar protein transport in Saccharomyces cerevisiae. Genetics, 150(2), 577-89, 1998 PMCID:1460353

Chen, C.-Y., and T. R. Graham. An arf1 synthetic lethal screen identifies a new clathrin heavy chain conditional allele that perturbs vacuolar protein transport. Genetics , 150, 577 - 589, 1998

Gaynor, E C, Chen, C Y, Emr, S D, Graham, T R. ARF is required for maintenance of yeast Golgi and endosome structure and function. Mol Biol Cell, 9(3), 653-70, 1998 PMCID:25294

Gaynor, E C, Graham, T R, Emr, S D. COPI in ER/Golgi and intra-Golgi transport: do yeast COPI mutants point the way. Biochim Biophys Acta, 1404(1-2), 33-51, 1998

Gaynor, E. C., C.-Y. Chen, S. D.Emr, and T. R.Graham. ARF is required for maintenance of yeast Golgi and endosome structure and function. Mol. Biol. Cell, 9, 653 - 670, 1998

Gaynor, E.C., T.R. Graham and S.D. Emr. COPI in ER/Golgi transport and intra-Golgi transport: do yeast COPs point the way?. Bioch. Biophys. Acta, 1404, 33 - 51, 1998

Reynolds, T B, Hopkins, B D, Lyons, M R, Graham, T R. The high osmolarity glycerol response (HOG) MAP kinase pathway controls localization of a yeast golgi glycosyltransferase. J Cell Biol, 143(4), 935-46, 1998 PMCID:2132948

Reynolds, T.B., B.D. Hopkins, M.R. Lyons and T.R. Graham. The high osmolarity glycerol response (HOG) MAP kinase pathway controls localization of a yeast Golgi glycosyltransferase. J. Cell Biol. , 143, 935-946, 1998

Graham, T R, Krasnov, V A. Sorting of yeast alpha 1,3 mannosyltransferase is mediated by a lumenal domain interaction, and a transmembrane domain signal that can confer clathrin-dependent Golgi localization to a secreted protein. Mol Biol Cell, 6(7), 809-24, 1995 PMCID:301242

Graham, T. R., and V. A. Krasnov. Sorting of yeast alpha1,3 mannosyltransferase is mediated by a lumenal domain interaction, and a transmembrane domain signal that can confer clathrin-dependent Golgi localization to a secreted protein. Mol. Biol. Cell, 6, 809-824 , 1995

Krasnov, V. and Graham, T.R. The Golgi complex of Saccharomyces cerevisiae. Can J Botany, 73, S343-S346, 1995

Gaynor, E.C., te Heesen, S., Graham, T.R., Aebi, M., and Emr, S.D. Signal-mediated retrieval of a membrane protein from the Golgi to the ER in yeast. J Cell Biology, 127, 653-665, 1994

Graham, T R, Seeger, M, Payne, G S, MacKay, V L, Emr, S D. Clathrin-dependent localization of alpha 1,3 mannosyltransferase to the Golgi complex of Saccharomyces cerevisiae. J Cell Biol, 127(3), 667-78, 1994 PMCID:2120240

Graham, T.R. and S.D. Emr. SEC18. In: Guidebook to the Secretory Pathway (J. Rothblatt, P. Novick, and T. Stevens, eds.) Oxford Univ. Press, NY, 132-133, 1994

Graham, T.R., Seegar, M., MacKay, V., Payne, G.S., and Emr, S.D. Clathrin-dependent localization of a, 3-mannosyltransferase to the Golgi complex of Saccharomyces cerevisiae. J Cell Biology, 127, 667-678, 1994

Graham, T.R., Scott, P., and Emr, Scott D. Brefeldin A reversibly blocks early but not late protein transport steps in the yeast secretory pathway . EMBO J, 12, 869-877, 1993

Horazdovsky, B.F., Graham, T.R., and Emr, S.D. ?Vacuolar protein sorting in yeast". Protein Synthesis and Targeting in Yeast (M.F. Tuite, J.E.G. McCarthy, A.J. Brown and F. Sherman, eds.) Springer Verlag, Berlin, 1992

Lacoste, H.C., Graham, T.R., and Kaplan, A. A sequence in b-hexosaminadase from Dictyostelium discoideum required for sorting of proteins to a compartment involved in developmentally induced secretion. J Biol Chem, 267, 5942-5948, 1992

Graham, T.R. and Emr, S.D. Compartmental organization of Golgi-specific protein modification and vacuolar protein sorting events defined in a yeast sec18 (NSF) mutant. J Cell Biology, 114, 207-218, 1991

Robinson, J.S., Graham, T.R.,and Emr, S.D. A putative zinc finger protein, Saccaromyces cerevisiae Vps18p, affects late Golgi functions required for vacuolar protein sorting and efficient a factor prohormone maturation. Mol Cell Biol, 12, 5813-5824, 1991

Vida, T.A., Herman, P.K., Emr, S.D., and Graham, T.R. Compartmentalized transport, modification and sorting of yeast vacuolar hydrolases. Biomed. Biochem. Acta, 50, 413-420, 1991


Postdoctoral Position Available
Yes

Postdoctoral Position Details
Applications are sought for a full time postdoctoral position to study the mechanism of substrate recognition and transport by P4-ATPases. Experience in molecular biology techniques such as cloning, mutagenesis, PCR, constructing gene fusions is essential. In addition, there is a preference for candidates familiar with protein expression and purification, microscopy, and bacterial and/or yeast microbial genetic techniques. Candidates must work well within a team environment and be willing to train graduate and undergraduate students.

Applicants should send a cover letter and curriculum vitae to Professor Todd Graham (tr.graham@vanderbilt.edu) in the Department of Biological Sciences at Vanderbilt University. Please include the names and addresses of three references who are willing to provide letters of recommendation. The position is open starting Sept 1, 2013 and will remain open until filled.

Updated Date
09/12/2013