A novel role for snapin in dendrite patterning: interaction with cypin.
Temporal and spatial assembly of signal transduction machinery determines dendrite branch patterning, a process crucial for proper synaptic transmission. Our laboratory previously cloned and characterized cypin, a protein that decreases PSD-95 family member localization and regulates dendrite number. Cypin contains zinc binding, collapsin response mediator protein (CRMP) homology, and PSD-95, Discs large, zona occludens-1 binding domains. Both the zinc binding and CRMP homology domains are needed for dendrite patterning. In addition, cypin binds tubulin via its CRMP homology domain to promote microtubule assembly. Using a yeast two-hybrid screen of a rat brain cDNA library with cypin lacking the carboxyl terminal eight amino acids as bait, we identified snapin as a cypin binding partner. Here, we show by affinity chromatography and coimmunoprecipitation that the carboxyl-terminal coiled-coil domain (H2) of snapin is required for cypin binding. In addition, snapin binds to cypin's CRMP homology domain, which is where tubulin binds. We also show that snapin competes with tubulin for binding to cypin, resulting in decreased microtubule assembly. Subsequently, overexpression of snapin in primary cultures of hippocampal neurons results in decreased primary dendrites present on these neurons and increased probability of branching. Together, our data suggest that snapin regulates dendrite number in developing neurons by modulating cypin-promoted microtubule assembly.
Chen
Maxine
M
Department of Cell Biology and Neuroscience, Rutgers University, Piscataway, NJ 08854-8082, USA.
Lucas
Kenyatta G
KG
Akum
Barbara F
BF
Balasingam
Gaithri
G
Stawicki
Tamara M
TM
Provost
Janine M
JM
Riefler
Gary M
GM
Jörnsten
Rebecka J
RJ
Firestein
Bonnie L
BL
D000818:Animals; D001667:Binding, Competitive; D019521:Body Patterning / Q000502:physiology*; D019556:COS Cells; D002352:Carrier Proteins / Q000378:metabolism*; D018929:Cell Culture Techniques; D002522:Chlorocebus aethiops; D002846:Chromatography, Affinity; D003712:Dendrites / Q000502:physiology*; D006148:Guanine Deaminase / Q000378:metabolism*; D006624:Hippocampus / Q000196:embryology; D008870:Microtubules / Q000378:metabolism / Q000502:physiology*; D008954:Models, Biological; D009474:Neurons / Q000378:metabolism; D017434:Protein Structure, Tertiary; D051381:Rats; D013574:Synaptosomes / Q000378:metabolism; D014162:Transfection; D033921:Vesicular Transport Proteins / Q000378:metabolism*
D002352:Carrier Proteins; C498452:Snapin protein, rat; D033921:Vesicular Transport Proteins; C485728:Gda protein, rat; D006148:Guanine Deaminase