This application responds to a sponsored program announcement (NIH/CHDI) to validate potential therapeutic targets in Huntington's disease (HD). A major goal in finding treatments for (HD) is to identify early events causing neuronal dysfunction. We have identified an early brain dysfunction in the trafficking of cargoes from recycling endosomes back to the plasma membrane in HD neurons. This deficit is due to interference by mutant htt with nucleotide exchange on Rab11, a small GTPase which functions at recycling endosomes. We identify the impaired Rab11 GEF activity in brain of pre-symptomatic HD knock in mice. Defects in membrane trafficking in neurons cause cell death. Rab11 activity is required for cellular homeostasis and survival. Numerous cargoes involved in cellular homeostasis recycle using a Rab11 dependent pathway. These cargoes can be cell specific. So far we found that the recycling of three important Rab11 dependent cargoes is slowed in HD neurons expressing endogenous full-length mutant htt. One neuron specific cargo affected in HD neurons is the cysteine/glutamate transporter. In HD neurons, EAAC1 dependent uptake of cysteine is impaired leading to insufficient glutathione synthesis, elevated ROS and cell death. Lentivirus delivery of dominant active Rab11 in HD primary cortical neurons rescues these deficits. Our in vitro data provide compelling evidence for the trophic effect of dominant active Rab11 and point to Rab11 as a superb target needing in vivo validation. We will validate Rab11 activity as a novel therapeutic target in two HD mouse models that express full length mutant htt-- HD140Q/140Q knock-in and HD YAC 128 transgenic. We have generated transgenic mice that constitutively express dominant active Rab11 (TgRab11 mice). The Tg mice at 9 weeks show no difference in behavior to littermate controls and will be further studied using in vivo and in vitro studies to rule out side effects.
In Aim 1, we will cross the HD mouse models with the TgRab11 mice. Since inducible expression simulates the condition for treatment of patients with HD, we will also cross HD mice with two transgenic lines that together incorporate the elements for a Cre-loxP system and will allow for tightly regulated inducible expression of dARab11 in presymptomatic and symptomatic HD mice.
Aim 2 will be to determine if increased Rab11 activity rescues HD phenotypes. We will evaluate in vivo motor behavior and biochemical and anatomical readouts and in vitro endosome recycling and neuronal survival. These studies will demonstrate that raising levels of Rab11 activity increases neuronal survival and ameliorates or delays motor deficits and neuropathology in HD. In vivo validation of increasing Rab11 activity will justify a drug discovery effort to screen for small molecules that raise the activity of Rab11 as a therapy for HD.

Public Health Relevance

A mutation in huntingtin causes Huntington's disease (HD). We have shown that mutant huntingtin interferes with the function of a small Rab GTPase called Rab11, which is required for the normal function of recycling endosomes. This dysfunction leads to disturbances in neuronal homeostasis that cause neurodegeneration. We propose to validate Rab11 as a therapeutic target for HD. We will genetically introduce a dominant active Rab11 into two mouse models of HD and show that an increase in the activity of Rab11 is beneficial and can rescue neurodegeneration and motor dysfunction in these mice.

National Institute of Health (NIH)
National Institute of Neurological Disorders and Stroke (NINDS)
Research Project (R01)
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Cell Death in Neurodegeneration Study Section (CDIN)
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Sutherland, Margaret L
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Massachusetts General Hospital
United States
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McClory, Hollis; Wang, Xiaolong; Sapp, Ellen et al. (2018) The COOH-terminal domain of huntingtin interacts with RhoGEF kalirin and modulates cell survival. Sci Rep 8:8000
McClory, Hollis; Williams, Dana; Sapp, Ellen et al. (2014) Glucose transporter 3 is a rab11-dependent trafficking cargo and its transport to the cell surface is reduced in neurons of CAG140 Huntington's disease mice. Acta Neuropathol Commun 2:179
Lu, Boxun; Al-Ramahi, Ismael; Valencia, Antonio et al. (2013) Identification of NUB1 as a suppressor of mutant Huntington toxicity via enhanced protein clearance. Nat Neurosci 16:562-70
Valencia, Antonio; Sapp, Ellen; Kimm, Jeffrey S et al. (2013) Elevated NADPH oxidase activity contributes to oxidative stress and cell death in Huntington's disease. Hum Mol Genet 22:1112-31
Valencia, Antonio; Sapp, Ellen; Kimm, Jeffrey S et al. (2013) Striatal synaptosomes from Hdh140Q/140Q knock-in mice have altered protein levels, novel sites of methionine oxidation, and excess glutamate release after stimulation. J Huntingtons Dis 2:459-75
Li, Xueyi; Valencia, Antonio; McClory, Hollis et al. (2012) Deficient Rab11 activity underlies glucose hypometabolism in primary neurons of Huntington's disease mice. Biochem Biophys Res Commun 421:727-30