CONTRIBUTIONS OF THE STRUCTURAL DOMAINS OF FILENSIN IN POLYMER FORMATION AND FILAMENT DISTRIBUTION

Filensin and phakinin constitute the subunits of a heteropolymeric, le ns-specific intermediate filament (IF) system known as the beaded-chai n filaments (BFs). Since the rod of filensin is four heptads shorter t han the rods of all other IF proteins, we decided to examine the speci fic contribution of this protein in filament assembly, For these purpo ses, we constructed chimeric proteins in which regions of filensin wer e exchanged with the equivalent ones of vimentin, a self-polymerizing IF protein, Our in vitro studies show that the filensin rod domain doe s not allow homopolymeric filament elongation, However, the filensin r od is necessary for co-polymerization of filensin with phakinin and se ems to counteract the inherent tendency of the latter protein to homop olymerize into large, laterally associated filament bundles, Apart fro m the rod domain, the presence of an authentic or substituted tail dom ain in filensin is also essential for co-assembly with the naturally t ail-less phakinin and formation of extended filaments in vitro, Finall y, transfection experiments in CHO and MCF-7 cells show that the rod d omain of filensin plays an important role in de novo filament formatio n and distribution, The same type of analysis further suggests that th e end-domains of filensin interact with cell-specific, assembly-modula ting factors.