Plectin belongs to the plakin family of high molecular weight cytolinkers. Plectin has a tripartite structure with N- and C-terminal regions separated by a central rod domain. The N-terminal region contains an actin binding domain (ABD) and a plakin domain. The ABD binds to integrin α6ß4 in hemidesmosomes and to the nuclear envelope protein nesprin-3. The plakin domain is formed by nine spectrin repeats (SR1-SR9) and an SH3 domain. Each SR consists of three α-helices (A-B-C) arranged in a left-handed bundle. Helix C of a SR is fussed to helix A of the downstream repeat; hence, the plakin domain has an elongated shape. Downstream of SR9, the rod forms a parallel coiled-coil that mediates dimerization. Previous results of our group suggested that short regions of the rod domain do not form stable dimers in vitro. Here, we have combined hybrid methods to elucidate the structure of a dimeric fragment of plectin that includes the SR7-SR9, a short region of the rod, and the GCN4 coiled-coil that stabilizes dimerization. The 2.1 Å resolution crystal structure revealed contacts along the plakin domain between two protomers. Small angle X-ray scattering (SAXS) supports that the closed arrangement of the dimer also occurs in solution. This was further confirmed by measuring 8 inter-monomer distances combining site directed spin labelling and double electron-electron resonance spectroscopy (EPR-DEER). Collectively, our data suggest that the plakin domain contributes to the stabilization of the plectin dimer. Moreover, the rigid structure of the SR3-SR9 constraints the maximum spacing between the binding-sites for integrin ß4 and nesprin-3 in the dimer, suggesting that avidity-driven recruitment of plectin might depend on the density of the interaction partners.

Resumen del póster presentado al XXXVIII Congreso de la Sociedad Española de Bioquímica y Biología Molecular, celebrado en Valencia (España) del 7 al 10 de septiembre de 2015.

Peer Reviewed