14-3-3 proteins associate with phosphorylated simple epithelial keratins during cell cycle progression and act as a solubility cofactor

J Cell Biol. 1996 Apr;133(2):345-57. doi: 10.1083/jcb.133.2.345.

Abstract

14-3-3 is a ubiquitous protein family that interacts with several signal transduction kinases. We show that 14-3-3 proteins associate with keratin intermediate filament polypeptides 8 and 18 (K8/18) that are expressed in simple-type epithelia. The association is stoichiometrically significant (> or = one 14-3-3 molecule/keratin tetramer), occurs preferentially with K18, and is phosphorylation- and cell cycle-dependent in that it occurs during S/G2/M phases of the cell cycle when keratins become hyperphosphorylated. Binding of phospho-K8/18 to 14-3-3 can be reconstituted in vitro using recombinant 14-3-3 or using total cellular cytosol. Phosphatase treatment results in dissociation of 14-3-3, and dephosphorylation of phospho-K8/18 prevents reconstitution of the binding. Three cellular keratin subpopulations were analyzed that showed parallel gradients of keratin phosphorylation and 14-3-3 binding. Incubation of 14-3-3 with keratins during or after in vitro filament assembly results in sequestering of additional soluble keratin, only in cases when the keratins were hyperphosphorylated. Our results demonstrate a stoichiometrically significant cell cycle- and phosphorylation-regulated binding of 14-3-3 proteins to K18 and in vitro evidence of a simple epithelial keratin sequestering role for 14-3-3 proteins.

Publication types

  • Research Support, U.S. Gov't, Non-P.H.S.
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • 14-3-3 Proteins
  • Amino Acid Sequence
  • Animals
  • Cell Cycle / physiology*
  • Cells, Cultured
  • Colon / cytology
  • Colon / metabolism
  • Cytosol / metabolism
  • Dogs
  • Epithelium / metabolism
  • Humans
  • Intermediate Filaments / metabolism
  • Keratins / chemistry
  • Keratins / metabolism*
  • Molecular Sequence Data
  • Phosphorylation
  • Protein Binding
  • Proteins / chemistry
  • Proteins / metabolism*
  • Solubility
  • Tyrosine 3-Monooxygenase*

Substances

  • 14-3-3 Proteins
  • Proteins
  • Keratins
  • Tyrosine 3-Monooxygenase