Unique role for the periplakin tail in intermediate filament association: specific binding to keratin 8 and vimentin

Abstract

Plectin, desmoplakin, and the 230-kDa bullous pemphigoid antigen (BPAG1), members of the plakin family of proteins, are multifunctional cytolinkers, connecting the cytoskeletal structures to the cell adhesion complexes. Envoplakin and periplakin are components of the cornified envelope, but less is known about their role in tissues other than the stratified epithelium. Our tissue-wide survey utilizing RT-PCR revealed that periplakin, like plectin and desmoplakin, has a wide tissue distribution, but envoplakin expression is limited to certain tissues only, and BPAG1 is clearly specific for epidermal keratinocytes. Plectin, desmoplakin and BPAG1 are known to bind to the intermediate filaments through their C-terminal domains. The short C-terminal domain of periplakin is composed only of the linker domain, a region highly homologous between the plakin proteins. Here we demonstrate, through the use of yeast two-hybrid assay, a specific interaction of the periplakin linker domain with keratin 8 and vimentin. Co-expression of each plakin linker domain with keratin 8 revealed that periplakin and BPAG1 linkers co-localize with keratin signals in HaCaT cells, plectin and desmoplakin linkers were detected both in the nucleus and in cytoplasm together with the overexpressed keratin 8, while envoplakin linker localized independently into the nucleus. These results suggest that, in spite of its high homology and structural similarity with envoplakin, periplakin is functionally closer to the well-characterized plakin proteins plectin and desmoplakin, and thus may function tissue-wide as a scaffolding protein in intermediate filament assembly.

Periplakin Interferes with G Protein Activation by the Melanin-concentrating Hormone Receptor-1 by Binding to the Proximal Segment of the Receptor C-terminal Tail

“…[ 35 S]GTP␥S Binding Assays-Membranes expressing equivalent levels of VSV-G-hMCH-1 full-length receptor (as determined by densitometric scanning of blots of immunodetected receptors using the anti-VSV-G antibody) were incubated with an assay mix (20 mM HEPES, pH 7.4, 5 mM MgCl 2 , 100 mM NaCl, 1 mM EDTA, 1 mM dithiothreitol, 0.1% bovine serum albumin, 50 M guanosine 5Ј-diphosphate, 100 nCi of [ 35 S]GTP␥S) containing the indicated concentrations of MCH. Nonspecific binding was determined in the presence of 100 M GTP␥S.…”

“…In brief, rat postnatal and adult brains were fixed overnight with 4% paraformaldehyde in PBS and sectioned using a cryomicrotome. Radiolabeled antisense and sense RNA probes were generated by in vitro transcription using [␣- 35 S]UTP and linearized cDNAs with nucleotides 1114 -1705 of human PPL (cloned from yeast clone L16) and with the full-length open reading frame of rMCH-1 receptor, respectively. Sections were hybridized overnight at 54°C in a humid chamber, washed with decreasing salt concentrations, dehydrated using increasing ethanol concentrations, and exposed to BetaMax x-ray film (Amersham Biosciences) for 3-7 days.…”

Direct interaction between FcγRI (CD64) and periplakin controls receptor endocytosis and ligand binding capacity

“…PPL is associated with desmosomes in keratinocytes and is involved in cornified envelope assembly (14)(15)(16)). PPL's central rod domain mediates homo-and heterodimerization and separates the N terminus (which can associate with actin filaments) from the vimentin-interacting C terminus (17)(18)(19). Additionally, recent studies suggested that PPL is involved in signaling through protein kinase B and G proteins in nonimmune cells (20,21).…”

Periplakin-dependent re-organisation of keratin cytoskeleton and loss of collective migration in keratin-8-downregulated epithelial sheets

“…The periplakin C-terminus is the shortest among the family members comprising only the linker domain (Ruhrberg et al, 1997). However, this minimal IF-binding domain is able to interact with intermediate filaments both in vivo and in vitro (DiColandrea et al, 2000;Kazerounian et al, 2002;Karashima and Watt, 2002). Unexpectedly, periplakin associates with cortical actin network in keratinocytes even though the protein lacks a classical actin-binding domain (DiColandrea et al, 2000).…”