Rutilio "Rudy" A Fratti
Associate Head of Biochemistry
Associate Professor of Biochemistry
Associate Professor, Center for Biophysics and Quantitative Biology
Drug Discovery, Host-Pathogen Interactions, Ion Channels, Membrane Biology, Protein Dynamics, Receptor Biochemistry
B.S. 1992 California State University, Long Beach
Ph.D. 2002 University of Michigan
Postdoc. 2002-2006 Dartmouth Medical School
Regulatory Lipids and Protein Function
Eukaryotic cells are compartmentalized by membrane-bound organelles that communicate through the trafficking of transport vesicles. The transfer of vesicular cargo between organelles is finalized by the fusing of two membrane bilayers into a continuous membrane. Membrane fusion is catalyzed by proteins called SNAREs that are present on both donor and acceptor membranes. Although seemingly straight forward, membrane fusion is highly orchestrated and tightly controlled to prevent unchecked fusion when unwanted, or missorting cargo to the wrong organelle. Thus, it is essential to regulate membrane trafficking and fusion to maintain homeostasis. Many proteins control these pathways, yet the role of the membrane itself remains underappreciated.
Membranes contain a small, yet critical group of “Regulatory Lipids” that aid in signal transduction, recruit proteins to their site of action, or alter membrane curvature and fluidity to modulate protein function. In the context of membrane fusion, regulatory lipids are essential for the fusion of synaptic vesicles to the plasma membrane, autophagosomes-to-lysosome fusion, phagosome-endosome fusion, et cetera. Regulatory lipids are essential throughout the different stages of membrane fusion and their effects can be altered though modification. Phosphoinositides, for example, can be differentially phosphorylated and dephosphorylated by specific kinase and phosphatases, or hydrolyzed by various lipases.
My lab focuses on how lipid modification affects protein function in the fusion pathway. It is clear that dysregulation of lipid modification has severe deleterious effects on fusion and overall cellular upkeep. In many cases pathologies have been mapped to lipid modifying enzymes, however, their effects on membrane function and cellular homeostasis is often unknown. Our long term goal is to bridge the gap between mapping diseases to genes and understanding the underlying mechanisms of the corresponding pathologies.
Current projects include:
I. Phosphatidic acid and Diacylglycerol Interconversion • Regulation of SNARE activation • Endolysosomal maturation • Autophagy
II. PI3P binding by Vam7 • Autoregulation of PI3P binding by Vam7 • Role of PI3P-Vam7 interaction in pathogenic yeast
III. PI(3,5)P2 production during fusion and fission • Reciprocal modulation of Ca2+ during fusion and fission • Role in regulating vacuole pH
IV. ABC transporters and vacuole fusion • Regulation of Ca2+ efflux • Interplay with PI(3,5)P2 production
Lipids and proteins that mediate membrane fusion are co-enriched at identical vertices. (Top) A schematic view of vacuoles showing the three morphological features that are developed during docking and fusion. (Bottom) Epifluorescence images of purified vacuoles in an in vitro docking assay. Vacuoles isolated from yeast harboring Vps33p-GFP were incubated with fluorescent Cy3-labeled FYVE domain to specifically localize PI(3)P. Cy3-FYVE is shown in red. To view the entire cluster of membranes, phosphatidylserine (PS), which is evenly distributed on vacuoles, was labeled with the fluorescent probe PSS-380 and is shown in blue. Solid arrows show examples of vertex sites enriched in Vps33p and PI(3)P. Hollow arrows specify outer membrane microdomains. Bar, 2 µm.
Hurst LR, Zhang C, Kazmirchuk TDD, Rivera-Kohr DA, Brett CL, Fratti RA. 2020. Sphingolipids with Very Long-chain Fatty Acids Regulate Vacuole Fusion During Tethering and Docking. bioRxiv.
Sparks RP, Arango AS, Aboff ZL, Jenkins JL, Guida WC, Tajkhorshid E, Sparks CE, Sparks JD, Fratti RA. 2019. Non-Canonical Binding of a Small Molecule to Sortilin Alters Cellular Trafficking of ApoB and PCSK9 in Liver Derived Cells. bioRxiv.
Sparks RP, Arango AS, Tajkhorshid E, Fratti RA. 2020. Use of Microscale Thermophoresis to Measure Protein-Lipid Interactions. J Vis Exp. in press.
Sparks R, Lui A, Bader D, Patel R, Murr M, Guida W, Fratti R, Patel N. 2019. A specific small-molecule inhibitor of Protein Kinase C-delta I activity improves metabolic dysfunction in human adipocytes from obese individuals. J Biol. Chem. 294, 14896-14910.
Sparks RP, Arango AS, Starr ML, Aboff ZL, Hurst LR, Rivera-Kohr DA, Zhang C, Harden KA, Jenkins JL, Guida WC, Tajkhorshid E, Fratti RA. 2019. A small-molecule competitive inhibitor of phosphatidic acid binding by the AAA+ protein NSF/Sec18 blocks the SNARE-priming stage of vacuole fusion. J Biol. Chem. 294, 17168-17185. * Featured Article
Miner GE, Sullivan KD, Jones BC, Guo A, Ellis EC, Starr ML, Fratti RA. 2019. Phosphatidylinositol 3,5-bisphosphate regulates Ca2+ transport during yeast vacuole fusion through activating the Ca2+ ATPase Pmc1. bioRxiv.
Miner GE, Sullivan KD, Zhang C, Hurst LR, Starr ML, Rivera-Kohr DA, Jones BC, Guo A, Fratti RA. 2019. Copper blocks V-ATPase activity and SNARE complex formation to inhibit yeast vacuole fusion. Traffic. 20:841-850.
Starr ML, Sparks RP, Hurst LR, Zhao Z, Arango A, Lihan M, Jenkins JL, Tajkhorshid E, Fratti RA. 2019. Phosphatidic acid induces conformational changes in Sec18 protomers that prevent SNARE priming. J. Biol. Chem. 294:3100-3116.
Starr ML, Fratti RA. 2019. The Participation of Regulatory Lipids in Vacuole Homotypic Fusion. Trends Biochem. Sci. 44:546-554 . * Featured Article
Miner GE, Fratti R. 2019. Real-Time Fluorescence Detection of Calcium Efflux During Vacuolar Membrane Fusion. Methods Mol. Biol. 1860:323-331.
Starr ML, Fratti R. 2019. Determination of Sec18-Lipid Interactions by Liposome-Binding Assay. Methods Mol. Biol. 1860:211-220.
Sparks RP, Jenkins JL, Fratti R. 2019. Use of Surface Plasmon Resonance (SPR) to Determine Binding Affinities and Kinetic Parameters Between Components Important in Fusion Machinery. Methods Mol. Biol. 1860:199-210.
Sparks RP, Fratti R. 2019. Use of Microscale Thermophoresis (MST) to Measure Binding Affinities of Components of the Fusion Machinery. Methods Mol. Biol. 1860:191-198.
Miner GE, Sullivan KD, Guo A, Jones BC, Hurst LR, Ellis EC, Starr ML, Fratti RA. 2019. Phosphatidylinositol 3,5-Bisphosphate Regulates the Transition between trans-SNARE Complex Formation and Vacuole Membrane Fusion. Mol. Biol. Cell. 30:201-208.
Miner GE, Starr ML, Hurst LR, Fratti RA. 2017. Deleting the DAG Kinase Dgk1 Augments Yeast Vacuole Fusion Through Increased Ypt7 Activity and Altered Membrane Fluidity. Traffic. 18:315-329.
Sparks RP, Jenkins LJ, Miner GE, Wangm Y, Guida WC, Sparks CE, Fratti RA, Sparks JD. 2016. Phosphatidylinositol (3,4,5)-trisphosphate binds to sortilin and competes with neurotensin: Implications for very low density lipoprotein binding. Biochem. Biophys. Res. Commun. 479:551-556.
Starr ML, Hurst LR, Fratti RA. 2016. Phosphatidic acid sequesters Sec18p from cis-SNARE complexes to inhibit priming. Traffic. 17:1091-1109.
Sparks RP, Guida WC, Sowden MP, Jenkins JL, Starr ML, Fratti RA, Sparks CE, Sparks JD. 2016. Sortilin facilitates VLDL-B100 secretion by insulin sensitive McArdle RH7777 cells. Biochem. Biophys. Res. Commun. 478:546-552.
Miner G, Starr ML, Hurst LR, Sparks RP Padolina M, Fratti RA. 2016. The Central Polybasic Region of the Soluble SNARE (Soluble N-Ethylmaleimide-sensitive Factor Attachment Protein Receptor) Vam7 Affects Binding to Phosphatidylinositol 3-Phosphate by the PX (Phox Homology) Domain. J. Biol. Chem. 291:17651-17663.
Sasser TL, Fratti RA. 2014. Class C ABC transporters and Saccharomyces cerevisiae vacuole fusion. Cell. Logist. 2014 Jul 3;4(3):e943588.
Lawrence G, Flood B, Brown C, Karunakaran S, Cabrera M, Nordmann M, Ungermann C, Fratti RA. 2014. Dynamic association of the PI3P-interacting Mon1-Ccz1 GEF with vacuoles is controlled through its phosphorylation by the type-1 casein kinase Yck3. Mol. Biol. Cell. 25:1608-1619.
Sasser TL, Lawrence G, Karunakaran S, Brown C, Fratti RA. 2013. The Yeast ATP-binding Cassette (ABC) Transporter Ycf1p Enhances the Recruitment of the Soluble SNARE Vam7p to Vacuoles for Efficient Membrane Fusion. J. Biol. Chem. 288(25):18300-10.
Karunankaran S, Fratti RA. 2013. The Lipid Composition and Physical Properties of the Yeast Vacuole Affect the Hemifusion-Fusion Transition. Traffic. 14(6):650-62.
Sasser TL, Padolina M, Fratti RA. 2012. The yeast vacuolar ABC transporter Ybt1p regulates membrane fusion through Ca2+ transport modulation. Biochem. J. 448(3):365-72.
Karunakaran S, Sasser T, Rajalekshmi S, Fratti RA. 2012. SNAREs, HOPS and regulatory lipids control the dynamics of vacuolar actin during homotypic fusion in S. cerevisiae. J. Cell Sci. 125(Pt 7):1683-92.
Sasser T, Karunakaran S, Qiu Q, Padolina M, Reyes A, Flood B, Smith S, Fratti RA. 2012 The Yeast Lipin 1 Orthologue Pah1p Regulates Vacuole Homeostasis and Membrane Fusion. J. Biol. Chem. 287:2221-2236.
Qiu Q, Fratti RA. 2010. The Na+/H+ exchanger Nhx1p Regulates the Initiation of Saccharomyces cerevisiae Vacuole Fusion. J. Cell Science. 123:3266-3275.