Control of receptor internalization and recycling to the plasma membrane is central to normal cell function, and dysregulation of these processes is the underlying cause for diseases as diverse as atherosclerosis, diabetes, and cancer. Among the key endocytic regulatory proteins are more than 50 Rab-GTP binding proteins and their effectors that control vesicle transport and fusion events. Over the past five years, we have focused on understanding the role of a novel endocytic regulatory protein family, known as the C-terminal Eps15 Homology Domain proteins (EHD). Despite a growing number of studies demonstrating roles for EHD proteins in the regulation of transport of a variety of receptors, thus far a unifying hypothesis for the actual mechanistic function of EHD proteins has remained elusive. EH-domains interact with proteins containing the tripeptide motif asparagine-proline- phenylalanine (NPF). Most significantly for this renewal application, we have discovered that the protein known as Molecule Interacting with CasL-Like 1 (MICAL-L1) is a novel NPF-containing interaction partner of EHD1. MICAL-L1 acts as unusual Rab effectors because it is critical for the recruitment of both Rab8a and EHD1 to tubular recycling endosomes that are enriched in phosphatidylinositol-4-phosphate and phosphatidylinositol-4, 5-bisphosphate. Depletion of MICAL-L1 simulates EHD1 depletion, causing a delay in recycling to the plasma membrane and an accumulation of internalized receptors at the endocytic recycling compartment (ERC). Moreover, new preliminary data show that MICAL-L1 also interacts with the membrane-bending BAR-domain protein, Syndapin II. Collectively, our data indicate a key role for MICAL-L1 in regulating the generation of tubular endosomes, and in recruiting EHD1, which appears to be responsible for their subsequent vesiculation. Our central hypothesis is that EHD1 plays a critical role in tubular membrane scission and facilitates recycling of membrane and proteins to the plasma membrane. Our first specific aim is to determine the mechanism by which EHD1 functions in tubule membrane scission and recycling. Our working hypothesis is that ATP hydrolysis by EHD1 promotes scission of vesicles from EHD1- containing tubules, thus supporting recycling to the plasma membrane. Our second Specific Aim is to characterize the roles of select EHD1 interaction partners in tubule membrane generation and in the modulation of EHD1 function. We hypothesize that BAR-domain-containing EHD1 interaction partners (such as Syndapin II and/or Bin1) generate the tubular membranes to which EHD1 is recruited. We further hypothesize that MICAL-L1 plays a crucial role in regulating EHD1 function by maintaining it on tubular membranes. Ultimately, these studies will facilitate the development of new strategies to treat the many diseases that arise as a result of aberrant endocytic events.

Public Health Relevance

Control of receptor localization to the plasma membrane is central to normal cell function, and dysregulation is the underlying cause for diseases as diverse as atherosclerosis, diabetes and cancer. A comprehensive understanding of the mechanisms that control membrane tubulation, transport and fission is of fundamental importance and will likely lead to the development of new therapeutic targets.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM074876-08
Application #
8706895
Study Section
Membrane Biology and Protein Processing (MBPP)
Program Officer
Ainsztein, Alexandra M
Project Start
2006-05-01
Project End
2016-07-31
Budget Start
2014-08-01
Budget End
2015-07-31
Support Year
8
Fiscal Year
2014
Total Cost
$281,118
Indirect Cost
$91,118
Name
University of Nebraska Medical Center
Department
Biochemistry
Type
Schools of Medicine
DUNS #
168559177
City
Omaha
State
NE
Country
United States
Zip Code
68198
Xie, Shuwei; Naslavsky, Naava; Caplan, Steve (2014) Diacylglycerol kinase ? regulates tubular recycling endosome biogenesis and major histocompatibility complex class I recycling. J Biol Chem 289:31914-26
Reinecke, James; Caplan, Steve (2014) Endocytosis and the Src family of non-receptor tyrosine kinases. Biomol Concepts 5:143-55
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
Cai, Bishuang; Xie, Shuwei; Liu, Fengming et al. (2014) Rapid degradation of the complement regulator, CD59, by a novel inhibitor. J Biol Chem 289:12109-25
Cai, Bishuang; Giridharan, Sai Srinivas Panapakkam; Zhang, Jing et al. (2013) Differential roles of C-terminal Eps15 homology domain proteins as vesiculators and tubulators of recycling endosomes. J Biol Chem 288:30172-80
Giridharan, Sai Srinivas Panapakkam; Cai, Bishuang; Vitale, Nicolas et al. (2013) Cooperation of MICAL-L1, syndapin2, and phosphatidic acid in tubular recycling endosome biogenesis. Mol Biol Cell 24:1776-90, S1-15
Simone, Laura C; Caplan, Steve; Naslavsky, Naava (2013) Role of phosphatidylinositol 4,5-bisphosphate in regulating EHD2 plasma membrane localization. PLoS One 8:e74519
Zhang, Jing; Reiling, Calliste; Reinecke, James B et al. (2012) Rabankyrin-5 interacts with EHD1 and Vps26 to regulate endocytic trafficking and retromer function. Traffic 13:745-57
Giridharan, Sai Srinivas Panapakkam; Rohn, Jennifer L; Naslavsky, Naava et al. (2012) Differential regulation of actin microfilaments by human MICAL proteins. J Cell Sci 125:614-24
Zhang, Jing; Naslavsky, Naava; Caplan, Steve (2012) Rabs and EHDs: alternate modes for traffic control. Biosci Rep 32:17-23

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