The in vitro assembly of hair follicle keratins: comparison of cortex and companion layer keratins.
The hair follicle consists of a complex system of multiple tissue compartments that are clearly distinguishable by their morphology and type of differentiation. We have synthesized hair follicle-specific keratins from the companion layer (K6hf, K17) and the hair cortex (Ha1, Hb3, Hb6) in Escherichia coli. The assembly of purified keratins in mixtures of K6hf/K17 and in mixtures of hair cortex keratins was compared in urea solutions, low ionic strength and physiological strength buffers, by urea melting gels, electron microscopy and analytical ultracentrifugation. Both types of keratin mixtures, keratins from the companion layer and keratins from the hair cortex, formed heterotypic complexes at 5 M urea. In low ionic strength buffers, the keratins from the companion layer were assembled to bona fide intermediate filaments. In contrast, mixtures of hair cortex keratins stayed in an oligomeric state with a mean s value of 9 as determined in sedimentation velocity experiments. Hair cortex keratins were, however, assembled into intermediate filaments at physiological salt conditions. A point mutated hair cortex keratin [Hb6(Glu402Lys)] formed no long filaments when mixed with Ha1; instead, the assembled structures showed a length distribution of 50.8 +/- 13.4 nm, comparable to the size distribution of assembly intermediates called 'unit-length' filaments.
Hofmann
Ilse
I
Division of Cell Biology, German Cancer Research Center, Heidelberg.
Winter
Hermelita
H
Mücke
Norbert
N
Langowski
Jörg
J
Schweizer
Jürgen
J
D003001:Cloning, Molecular; D015180:Electrophoresis, Gel, Two-Dimensional; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D018859:Hair Follicle / Q000378:metabolism* / Q000648:ultrastructure; D006801:Humans; D007382:Intermediate Filaments / Q000378:metabolism / Q000648:ultrastructure; D007633:Keratins / Q000378:metabolism*; D008854:Microscopy, Electron; D012313:RNA / Q000737:chemistry / Q000235:genetics; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014461:Ultracentrifugation
D011994:Recombinant Proteins; D012313:RNA; D007633:Keratins