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Huntingtin-Associated Protein HAP1 trafficking and orexin neuronal function

Huntingtin-Associated Protein HAP1 trafficking and orexin neuronal function. Yung-Feng Lin Postdoctoral Fellow Li Lab. References. HAP1 and Htt in trafficking. 1995 1997, 1998 1997 2002 2004 2003 2003 2006 2006 2007 2007 2008. Kinesin light chain (KLC).

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Huntingtin-Associated Protein HAP1 trafficking and orexin neuronal function

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  1. Huntingtin-Associated Protein HAP1 trafficking and orexin neuronal function Yung-Feng Lin Postdoctoral Fellow Li Lab

  2. References HAP1 and Htt in trafficking 1995 1997, 1998 1997 2002 2004 2003 2003 2006 2006 2007 2007 2008 Kinesin light chain (KLC) Microtubule-dependent transport Androgen receptor (AR) Membrane receptor 14-3-3 Protein trafficking complex assembly Salinas S et al, Curr Opin Cell Biol 2008 TBP Transcription factor AHI1 Intracellular trafficking

  3. 100 75 % Survival 50 25 0 P1 P2 P3 P15 P4 P6 P9 HAP1 expression in hypothalamus P3 P10 P15 Li et al., J. Neurosci, 2003

  4. Question: Dose reducing HAP1 expression in hypothalamic neurons in adult mice also lead to neurological phenotypes? Approach: Use conditional HAP1 knockout mice to examine HAP1 deficiency-related phenotypes

  5. Orexin neurons are located in the lateral hypothalamic area (LHA) and project to almost all parts of the brain except the cerebellum Orexin expression HAP1 expression Coronal sections LH DMH VMH ARC Orexin neuronal function: Feeding Locomotor Activity Sleep/wakefulness My study focuses on orexin neurons because of their importance and availability of orexin-Cre mice

  6. Cre-LoxP system and the Orexin-Hap1 conditional knockout mouse 3951 6957 Hap1 TK E1 Hap1-LoxP 2 kb 2 kb TK E1 Neo Cre recombinase Hap1-CKO M C T Tc Hc TK 5 Kb HAP1-loxP WT 3 Kb KO 1 Kb

  7. Homozygous orexin-HAP1 knockout selectively depletes HAP1 in orexin neurons Het Hom HAP1 Orexin A Merged

  8. Orexin-HAP1 KO mice show decreased body weight Body weight (g) Body weight (% of WT)

  9. Orexin-HAP1 KO mice eat less Food intake

  10. Orexin-HAP1 KO mice also have lower locomotor activity activity

  11. HAP1 knockout reduces neurite extension of orexin neurons in mouse brain WT KO

  12. Question: Does lack of HAP1 affect neuronal intracellular trafficking in mouse brain? Approach: Sucrose gradient fractionation to examine the distribution of organelle marker proteins in the brains of HAP1 null and WT pups.

  13. Sucrose gradient 5-45% Input Htt (2166) Hap1 pHap1A Ahi1 Early endosome EEA1 Golgi complex GM130 Post-synaptic density PSD95 Synaptic vesicle Syntaxin Autophagisome Beclin-1 Mitochondria SOD2

  14. 5 ~ 45% Sucrose Input WT Tubulin KO WT Dynactin p150 KO WT Kinesin Light chain KO

  15. Lack of HAP1 affects the distribution of lysosomes and BDNF 5 ~ 45% Sucrose Input LAMP1 WT KO WT BDNF KO

  16. HAP1 deficiency Conclusion Intracellular trafficking defect Impaired neurite extension Orexin neuronal dysfunction Feeding and locomotor activity changes

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