α-Actinin-dependent cytoskeletal anchorage is important for ICAM-5-mediated neuritic outgrowth

University of Helsinki, Helsinki, Uusimaa, Finland
Journal of Cell Science (Impact Factor: 5.43). 09/2006; 119(Pt 15):3057-66. DOI: 10.1242/jcs.03045
Source: PubMed


Intercellular adhesion molecule-5 (ICAM-5, telencephalin) is a dendrite-expressed membrane glycoprotein of telencephalic neurons in the mammalian brain. By deletion of the cytoplasmic and membrane-spanning domains of ICAM-5, we observed that the membrane distribution of ICAM-5 was determined by the cytoplasmic portion. Therefore we have characterized the intracellular associations of ICAM-5 by using a bacterially expressed glutathione S-transferase (GST) fusion protein encompassing the cytoplasmic part of ICAM-5. One of the main proteins in the neuronal cell line Paju that bound to the ICAM-5 cytodomain was alpha-actinin. ICAM-5 expressed in transfected Paju cells was found in alpha-actinin immunoprecipitates, and ICAM-5 colocalized with alpha-actinin both in Paju cells and in dendritic filopodia and spines of primary hippocampal neurons. We were also able to coprecipitate alpha-actinin from rat brain homogenate. Binding to alpha-actinin appeared to be mediated mainly through the N-terminal region of the ICAM-5 cytodomain, as the ICAM-5(857-861) cytoplasmic peptide (KKGEY) mediated efficient binding to alpha-actinin. Surface plasmon resonance analysis showed that the turnover of the interaction was rapid. In a mutant cell line, Paju-ICAM-5-KK/AA, the distribution was altered, which implies the importance of the lysines in the interaction. Furthermore, we found that the ICAM-5/alpha-actinin interaction is involved in neuritic outgrowth and the ICAM-5(857-861) cytoplasmic peptide induced morphological changes in Paju-ICAM-5 cells. In summary, these results show that the interaction between ICAM-5 and alpha-actinin is mediated through binding of positively charged amino acids near the transmembrane domain of ICAM-5, and this interaction may play an important role in neuronal differentiation.

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    • "All PCR-derived clones were verified by sequencing. The GST-a-actinin and the GST- ICAM-5cyto fusion proteins representing the ICAM-5 cytoplasmic domain were purified by affinity chromatography as described previously (Gilmore et al., 1994; Nyman-Huttunen et al., 2006). "
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    • "The binding to -actinin [54] and ezrin [55] involves the sequence KKGEY in the membrane proximal part of the ICAM-5 cytoplasmic domain. When the cytoplasmic region was deleted or when the protein was anchored to the membrane through a glycophosphatidyl inositol-anchor, the distribution changed to a diffuse pattern in the plane of the membrane , whereas the full-length ICAM-5 molecule showed a more patchy distribution [54]. ICAM-5 co-localized with -actinin to the soma and dendritic shafts, and when this interaction was interrupted using a membrane-permeable KKGEY-containing peptide, it resulted in morphological changes of the cells and reduction in dendritic outgrowth. "
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