Endocytic recycling is essential for the control of receptors on the plasma membrane in mammalian cells. Consequently, recycling impacts health by regulating crucial cellular events such as signal transduction and proliferation, cholesterol homeostasis, nutrient uptake, and insulin-dependent glucose transport. Thus, understanding the regulation of endocytic events is critical for a wide range of diseases, including malignancies, heart disease and diabetes. Endocytic transport and recycling are controlled by a number of small Rab GTP-binding proteins. Recently, a non-Rab protein called EHD1 has been ascribed a role in regulating recycling at the recycling compartment. However, the mode by which EHD1 coordinates its regulatory activity with Rab-family proteins is not understood. A biochemical approach has identified the divalent Rab4/Rab5 effector prtoein, Rabenosyn-5, as a binding partner for EHD1, and defined a role for it in recycling at the early endosome. The critical task at hand is to understand how trafficking events at the early endosome are linked to those at the recycling compartment, particularly what regulates transport and fusion of early endosome-derived vesicles with the recycling compartment.
The first aim will focus on identifying the mechanisms by which proteins are transported from early endosomes to the endocytic recycling compartment, en route to the plasma membrane. The working hypothesis is that the interaction between EHD1 and Rabenosyn-5 is critical for transport of internalized proteins from early endosomes to the recycling compartment.
The second aim i s based on new data elucidating a physical connection between EHD proteins and a Rab11 effector protein, and proposes to determine the mechanisms by which EHD proteins coordinate endocytic recycling and transport with Rab11 and SNARE proteins. The working hypothesis is that EHD proteins coordinate transport steps with Rab11 and its effectors, and that the SNARE proteins Syntaxin13 and SNAP29 play a critical role in fusion of early endosome-derived vesicles at the recycling compartment.
These aims will be accomplished using novel fibroblasts from EHD1-knock-out mice, RNAi- based 'knock-down/knock-in'strategy, and a series of biochemical, flow cytometry and microscopic assays. These studies will significantly enhance our fundamental understanding of the mechanisms regulating recycling and have an important bearing on diseases as diverse as cancer and atherosclerosis and diabetes.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM074876-05
Application #
7843430
Study Section
Cell Structure and Function (CSF)
Program Officer
Ainsztein, Alexandra M
Project Start
2006-05-01
Project End
2012-04-30
Budget Start
2010-05-01
Budget End
2012-04-30
Support Year
5
Fiscal Year
2010
Total Cost
$242,022
Indirect Cost
Name
University of Nebraska Medical Center
Department
Biochemistry
Type
Schools of Medicine
DUNS #
168559177
City
Omaha
State
NE
Country
United States
Zip Code
68198
Caplan, Steve (2017) Into the linker's DENN: A tyrosine's control of autophagy. J Biol Chem 292:7283-7284
Farmer, Trey; Reinecke, James B; Xie, Shuwei et al. (2017) Control of mitochondrial homeostasis by endocytic regulatory proteins. J Cell Sci 130:2359-2370
Xie, Shuwei; Bahl, Kriti; Reinecke, James B et al. (2016) The endocytic recycling compartment maintains cargo segregation acquired upon exit from the sorting endosome. Mol Biol Cell 27:108-26
Bahl, Kriti; Xie, Shuwei; Spagnol, Gaelle et al. (2016) EHD3 Protein Is Required for Tubular Recycling Endosome Stabilization, and an Asparagine-Glutamic Acid Residue Pair within Its Eps15 Homology (EH) Domain Dictates Its Selective Binding to NPF Peptides. J Biol Chem 291:13465-78
Reineke, James B; Xie, Shuwei; Naslavsky, Naava et al. (2015) Qualitative and quantitative analysis of endocytic recycling. Methods Cell Biol 130:139-55
Reinecke, James B; Katafiasz, Dawn; Naslavsky, Naava et al. (2015) Novel functions for the endocytic regulatory proteins MICAL-L1 and EHD1 in mitosis. Traffic 16:48-67
McAtee, Caitlin O; Berkebile, Abigail R; Elowsky, Christian G et al. (2015) Hyaluronidase Hyal1 Increases Tumor Cell Proliferation and Motility through Accelerated Vesicle Trafficking. J Biol Chem 290:13144-56
Bahl, Kriti; Naslavsky, Naava; Caplan, Steve (2015) Role of the EHD2 unstructured loop in dimerization, protein binding and subcellular localization. PLoS One 10:e0123710
Simone, Laura C; Naslavsky, Naava; Caplan, Steve (2014) Scratching the surface: actin' and other roles for the C-terminal Eps15 homology domain protein, EHD2. Histol Histopathol 29:285-92
Reinecke, James; Caplan, Steve (2014) Endocytosis and the Src family of non-receptor tyrosine kinases. Biomol Concepts 5:143-55

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