Direct demonstration of NCAM cis-dimerization and inhibitory effect of palmitoylation using the BRET2 technique.
Biological activity of the neural cell adhesion molecule (NCAM) depends on both adhesion and activation of intra-cellular signaling. Based on in vitro experiments with truncated extra-cellular domains, several models describing homophilic NCAM trans- and cis-interactions have been proposed. However, cis-dimerization in living cells has not been shown directly and the role of the cytoplasmic part in NCAM dimerization is poorly understood. Here, we used the bioluminescence resonance energy transfer (BRET(2)) technique to directly demonstrate that full-length NCAM cis-homodimerizes in living cells. Based on BRET(2)50 values we suggest that the intra-cellular part of NCAM inhibits cis-dimerization, an effect mainly dependent on the palmitoylation sites.
Department of Neuroscience and Pharmacology, Institute of Health Sciences, University of Copenhagen, Copenhagen, Denmark. niku@hagedorn.dk
Nikolaj
N
Kulahin
Lars Groth
LG
Grunnet
Morten
M
Lundh
Dan Ploug
DP
Christensen
Rasmus
R
Jorgensen
Anders
A
Heding
Nils
N
Billestrup
Vladimir
V
Berezin
Elisabeth
E
Bock
Thomas
T
Mandrup-Poulsen
D000818:Animals; D019556:COS Cells; D002522:Chlorocebus aethiops; D004735:Energy Transfer; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D054878:Lipoylation; D008163:Luminescent Measurements / Q000379:methods*; D019006:Neural Cell Adhesion Molecules / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D055503:Protein Multimerization; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D014162:Transfection
D019006:Neural Cell Adhesion Molecules; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins