Identification of trichoplein, a novel keratin filament-binding protein.
Keratins 8 and 18 (K8/18) are major components of the intermediate filaments (IFs) of simple epithelia. We report here the identification of a novel protein termed trichoplein. This protein shows a low degree of sequence similarity to trichohyalin, plectin and myosin heavy chain, and is a K8/18-binding protein. Among interactions between trichoplein and various IF proteins that we tested using two-hybrid methods, trichoplein interacted significantly with K16 and K18, and to some extent with K5, K6a, K8 and K14. In in vitro co-sedimentation assays, trichoplein directly binds to K8/18, but not with vimentin, desmin, actin filaments or microtubules. An antibody raised against trichoplein specifically recognized a polypeptide with a relative molecular mass of 61 kDa in cell lysates. Trichoplein was immunoprecipitated using this antibody in a complex with K8/18 and immunostaining revealed that trichoplein colocalized with K8/18 filaments in HeLa cells. In polarized Caco-2 cells, trichoplein colocalized not only with K8/18 filaments in the apical region but also with desmoplakin, a constituent of desmosomes. In the absorptive cells of the small intestine, trichoplein colocalized with K8/18 filaments at the apical cortical region, and was also concentrated at desmosomes. Taken together, these results suggest that trichoplein is a keratin-binding protein that may be involved in the organization of the apical network of keratin filaments and desmosomes in simple epithelial cells.
Nishizawa
Miwako
M
Division of Biochemistry, Aichi Cancer Center Research Institute, 1-1 Kanokoden, Chikusa-ku, Nagoya 464-8681, Japan. minagaki@aichi-cc.jp
Izawa
Ichiro
I
Inoko
Akihito
A
Hayashi
Yuko
Y
Nagata
Koh-ichi
K
Yokoyama
Tomoya
T
Usukura
Jiro
J
Inagaki
Masaki
M
D000595:Amino Acid Sequence; D000818:Animals; D018938:Caco-2 Cells; D002352:Carrier Proteins / Q000737:chemistry / Q000502:physiology*; D045744:Cell Line, Tumor; D003001:Cloning, Molecular; D003598:Cytoskeletal Proteins / Q000378:metabolism; D018076:DNA, Complementary / Q000378:metabolism; D051180:Desmoplakins; D003896:Desmosomes / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D047468:Immunoprecipitation; D007381:Intermediate Filament Proteins / Q000737:chemistry; D007382:Intermediate Filaments / Q000378:metabolism*; D053538:Keratin-18; D053533:Keratin-8; D007633:Keratins / Q000737:chemistry* / Q000378:metabolism*; D051379:Mice; D008854:Microscopy, Electron; D008856:Microscopy, Fluorescence; D008870:Microtubules / Q000737:chemistry; D008969:Molecular Sequence Data; D018995:Myosin Heavy Chains / Q000737:chemistry; D010455:Peptides / Q000737:chemistry; D051190:Plectin; D016133:Polymerase Chain Reaction; D011485:Protein Binding; D011498:Protein Precursors / Q000737:chemistry; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000378:metabolism; D014162:Transfection; D014404:Tubulin / Q000737:chemistry; D020798:Two-Hybrid System Techniques
D002352:Carrier Proteins; D003598:Cytoskeletal Proteins; D018076:DNA, Complementary; C495294:DSP protein, human; D051180:Desmoplakins; C495295:Dsp protein, mouse; D007381:Intermediate Filament Proteins; C488421:KRT18 protein, human; C426203:KRT8 protein, human; D053538:Keratin-18; D053533:Keratin-8; C495332:PLEC protein, human; D010455:Peptides; C495333:Plec protein, mouse; D051190:Plectin; D011498:Protein Precursors; D011994:Recombinant Proteins; C502303:TCHP protein, human; D014404:Tubulin; C016159:trichohyalin; D049452:Green Fluorescent Proteins; D007633:Keratins; D018995:Myosin Heavy Chains