The long-term goal is to understand the physiological and pathological roles of Rab GTPases in cells and organisms. This project is focused on two closely related Rab GTPases (Rab22 and Rab5) that are localized in early endosomes and regulate endosomal sorting and endocytosis. Specifically the hypothesis that Rab22 is evolved from Rab5 to accommodate the need for regulation of specialized cell differentiation processes in higher eukaryotes, such as nerve growth factor (NGF)-mediated neuronal differentiation and survival, will be investigated. Malfunction in NGF signal transduction plays an important role in neurodegenerative diseases such as Alzheimer's disease and Down syndrome. We found that inhibition of Rab22 function via RNAi or dominant negative mutants dramatically reduces NGF-induced VGF expression and neurite outgrowth in PC12 cells, in contrast to the stimulatory effect of dominant negative Rab5 mutants. Furthermore, the activated NGF receptor (pTrkA) is endocytosed and localized to Rab22-containing endosomes. The preliminary data suggest an important role for Rab22 in the biogenesis and function of NGF-pTrkA signaling endosomes that promote cell differentiation and survival. This project contains three specific aims to investigate the mechanism of Rab22 in NGF signal transduction and cell differentiation in PC12 cells as well as neurons and to determine the structural differences that distinguish Rab22 from Rab5 in terms of promoting NGF signal transduction and cell differentiation.
Aim 1 will test if Rab22 knockdown via RNAi may abrogate the formation of NGF-pTrkA signaling endosomes and consequently diminish the level and duration of pTrkA in PC12 cells and the activities of downstream effectors (Rap1 and ERKs). Furthermore, the experiments will determine Rab22- specific effectors responsible for the function in NGF-induced activation of ERKs, VGF expression, and neurite outgrowth.
Aim 2 will test if Rab22 knockdown may abrogate the retrograde trafficking and signaling of NGF- pTrkA signaling endosomes in dorsal root ganglia (DRG) neurons and lead to apoptosis.
Aim 3 will test if and what mutations in Rab5 may give rise to Rab22 characteristics in terms of specificity in interactions with effectors and function in NGF signaling and cell differentiation. In summary, this project will contribute to understanding the evolution of the Rab GTPase family in terms of structural and functional specificity of Rab22 and Rab5. Furthermore, it develops a novel concept that the early endosome-associated Rab22 is critical for the biogenesis and function of signaling endosomes that promote NGF signaling and neuron differentiation, which will contribute to understanding the mechanism of neurodegenerative diseases and developing potential therapeutics.

Public Health Relevance

This project will investigate the mechanism of how endosomal Rab proteins may control nerve growth factor (NGF) signal transduction and neuronal differentiation and survival. Preliminary studies suggest that endosomal Rab proteins play an important role in regulation of endocytosis, sorting, and retrograde trafficking of NGF in neurons, which is required for NGF signaling processes. The results may have significant impact on the development of diagnosis and therapeutics against neurodegenerative disorders such as Alzheimer's disease, Down syndrome, and hereditary sensory and autonomic neuropathy.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
2R01GM074692-06A1
Application #
8237266
Study Section
Membrane Biology and Protein Processing (MBPP)
Program Officer
Dunsmore, Sarah
Project Start
2005-07-01
Project End
2016-01-31
Budget Start
2012-05-01
Budget End
2013-01-31
Support Year
6
Fiscal Year
2012
Total Cost
$279,729
Indirect Cost
$89,729
Name
University of Oklahoma Health Sciences Center
Department
Biochemistry
Type
Schools of Medicine
DUNS #
878648294
City
Oklahoma City
State
OK
Country
United States
Zip Code
73117
Zhang, Dongmei; Marlin, M Caleb; Liang, Zhimin et al. (2016) The Protein Tyrosine Phosphatase MEG2 Regulates the Transport and Signal Transduction of Tropomyosin Receptor Kinase A. J Biol Chem 291:23895-23905
Marlin, M Caleb; Li, Guangpu (2015) Differential effects of overexpression of Rab5 and Rab22 on autophagy in PC12 cells with or without NGF. Methods Mol Biol 1298:295-304
Qi, Yaoyao; Liang, Zhimin; Wang, Zonghua et al. (2015) Determination of Rab5 activity in the cell by effector pull-down assay. Methods Mol Biol 1298:259-70
Maskey, Dipak; Marlin, Matthew Caleb; Kim, Seokho et al. (2015) Cell cycle-dependent ubiquitylation and destruction of NDE1 by CDK5-FBW7 regulates ciliary length. EMBO J 34:2424-40
Qi, Yaoyao; Marlin, M Caleb; Liang, Zhimin et al. (2015) Expression and localization of exocytic and recycling Rabs from Magnaporthe oryzae in mammalian cells. Methods Cell Biol 130:35-45
Marlin, M Caleb; Li, Guangpu (2015) Biogenesis and function of the NGF/TrkA signaling endosome. Int Rev Cell Mol Biol 314:239-57
Wu, Kong-Yan; He, Miao; Hou, Qiong-Qiong et al. (2014) Semaphorin 3A activates the guanosine triphosphatase Rab5 to promote growth cone collapse and organize callosal axon projections. Sci Signal 7:ra81
Qi, Yaoyao; Marlin, M Caleb; Liang, Zhimin et al. (2014) Distinct biochemical and functional properties of two Rab5 homologs from the rice blast fungus Magnaporthe oryzae. J Biol Chem 289:28299-309
Dou, Zhixun; Pan, Ji-An; Dbouk, Hashem A et al. (2013) Class IA PI3K p110β subunit promotes autophagy through Rab5 small GTPase in response to growth factor limitation. Mol Cell 50:29-42
Li, Guangpu (2011) Rab GTPases, membrane trafficking and diseases. Curr Drug Targets 12:1188-93

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