Characterization of human epiplakin: RNAi-mediated epiplakin depletion leads to the disruption of keratin and vimentin IF networks.
Epiplakin is a member of the plakin family with multiple copies of the plakin repeat domain (PRD). We studied the subcellular distribution and interactions of human epiplakin by immunostaining, overlay assays and RNAi knockdown. Epiplakin decorated the keratin intermediate filaments (IF) network and partially that of vimentin. In the binding assays, the repeat unit (PRD plus linker) showed strong binding and preferentially associated with assembled IF over keratin monomers. Epiplakin knockdown revealed disruption of IF networks in simple epithelial but not in epidermal cells. In rescue experiments, the repeat unit was necessary to prevent the collapse of IF networks in transient knockdown; however, it could only partially restore the keratin but not the vimentin IF network in stably knocked down HeLa cells. We suggest that epiplakin is a cytolinker involved in maintaining the integrity of IF networks in simple epithelial cells. Furthermore, we observed an increase of epiplakin expression in keratinocytes after the calcium switch, suggesting the involvement of epiplakin in the process of keratinocyte differentiation.
Jang
Shyh-Ing
SI
Laboratory of Skin Biology, NIAMS, National Institutes of Health, Bethesda, MD 20892-8023, USA.
Kalinin
Alexandr
A
Takahashi
Kaoruko
K
Marekov
Lyuben N
LN
Steinert
Peter M
PM
D001324:Autoantigens / Q000032:analysis / Q000737:chemistry / Q000502:physiology*; D002454:Cell Differentiation; D002478:Cells, Cultured; D006367:HeLa Cells; D006801:Humans; D007382:Intermediate Filaments / Q000737:chemistry / Q000648:ultrastructure*; D015603:Keratinocytes / Q000166:cytology / Q000378:metabolism / Q000648:ultrastructure*; D007633:Keratins / Q000032:analysis*; D017434:Protein Structure, Tertiary; D034622:RNA Interference; D014746:Vimentin / Q000032:analysis*
D001324:Autoantigens; D014746:Vimentin; C426678:epiplakin; D007633:Keratins