PMID- 18413253
OWN - NLM
STAT- MEDLINE
DCOM- 20080619
LR  - 20191210
IS  - 0076-6879 (Print)
IS  - 0076-6879 (Linking)
VI  - 438
DP  - 2008
TI  - Regulation of endosome dynamics by Rab5 and Huntingtin-HAP40 effector complex in 
      physiological versus pathological conditions.
PG  - 239-57
LID - 10.1016/S0076-6879(07)38017-8 [doi]
AB  - Vesicular transport of signaling molecules, specifically neurotrophins, in 
      neurons is essential for their differentiation, survival, and plasticity. 
      Neurotrophins such as neuron growth factor (NGF) and brain-derived neurotrophic 
      factor (BDNF) are internalized by receptor-mediated endocytosis at synaptic 
      terminals and loaded into endosomes for microtubule-based transport along axons 
      to the cell body where they exert their signaling function in the nucleus. The 
      molecular mechanisms underlying this intracellular transport are not only 
      relevant from a basic knowledge viewpoint, but have also important implications 
      for neurodegenerative diseases. Defects in trafficking are increasingly 
      implicated in the pathology of Huntington's disease (HD) and other 
      neurodegenerative disorders. The small GTPases Rab5 and Rab7 play important roles 
      in the endocytic trafficking of neurotrophins. We have recently identified 
      Huntingtin (Htt) and Huntingtin associated protein of 40 kDa (HAP40) as a novel 
      Rab5 effector complex that regulates endosome motility. In HD, we detected higher 
      HAP40 protein levels compared with normal cells. Such increase causes an 
      augmented recruitment of Htt onto Rab5-positive early endosomes that drastically 
      reduces their motility by "switching" these organelles from microtubules to 
      F-actin. These findings suggest a mechanism by which impaired Rab5-mediated 
      trafficking of neurotrophic factors may be a key event of the pathogenetic 
      process leading to neurodegeneration in HD. To dissect the mechanisms by which 
      Htt, HAP40, and Rab5 function in early endosome interactions with the 
      cytoskeleton, we developed assays to investigate endosome-cytoskeleton 
      interactions that can be applied to normal and pathological conditions. We 
      provide here detailed protocols for, first, an assay that measures binding of 
      early endosomes to microtubules and F-actin. Second, we describe an improved 
      protocol for a cell-free assay that recapitulates the motility of early endosomes 
      along microtubules in vitro. These assays provide mechanistic insights into the 
      dysfunction of endosome motility occurring in HD as well as other 
      neurodegenerative disorders.
FAU - Pal, Arun
AU  - Pal A
AD  - Max Planck Institute of Molecular Cell Biology and Genetics, Dresden, Germany.
FAU - Severin, Fedor
AU  - Severin F
FAU - Hopfner, Sebastian
AU  - Hopfner S
FAU - Zerial, Marino
AU  - Zerial M
LA  - eng
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
PL  - United States
TA  - Methods Enzymol
JT  - Methods in enzymology
JID - 0212271
RN  - 0 (Actins)
RN  - 0 (Carrier Proteins)
RN  - 0 (F8a protein, mouse)
RN  - 0 (HTT protein, human)
RN  - 0 (Huntingtin Protein)
RN  - 0 (Intracellular Signaling Peptides and Proteins)
RN  - 0 (Nerve Tissue Proteins)
RN  - 0 (Nuclear Proteins)
RN  - EC 3.6.5.2 (rab5 GTP-Binding Proteins)
SB  - IM
MH  - Actins/metabolism
MH  - Animals
MH  - Blotting, Western
MH  - Carrier Proteins/*physiology
MH  - Cell Movement/drug effects
MH  - Endocytosis/*drug effects/physiology
MH  - Endosomes/metabolism
MH  - HeLa Cells
MH  - Humans
MH  - Huntingtin Protein
MH  - Intracellular Signaling Peptides and Proteins
MH  - Mice
MH  - Microtubules/metabolism
MH  - Nerve Tissue Proteins/physiology
MH  - Nuclear Proteins/physiology
MH  - Protein Transport
MH  - rab5 GTP-Binding Proteins/*physiology
EDAT- 2008/04/17 09:00
MHDA- 2008/06/20 09:00
CRDT- 2008/04/17 09:00
PHST- 2008/04/17 09:00 [pubmed]
PHST- 2008/06/20 09:00 [medline]
PHST- 2008/04/17 09:00 [entrez]
AID - S0076-6879(07)38017-8 [pii]
AID - 10.1016/S0076-6879(07)38017-8 [doi]
PST - ppublish
SO  - Methods Enzymol. 2008;438:239-57. doi: 10.1016/S0076-6879(07)38017-8.