Protein 4.1R binding to eIF3-p44 suggests an interaction between the cytoskeletal network and the translation apparatus

Blood. 2000 Jul 15;96(2):747-53.

Abstract

Erythroid protein 4.1 (4.1R) is an 80-kd cytoskeletal protein that stabilizes the membrane-skeletal network structure underlying the lipid bilayer. Using the carboxyl terminal domain (22/24 kd) of 4.1R as bait in a yeast 2-hybrid screen, we isolated cDNA clones encoding a polypeptide of eIF3-p44, which represents a subunit of a eukaryotic translation initiation factor 3 (eIF3) complex. The eIF3 complex consists of at least 10 subunits that play an essential role in the pathway of protein translation initiation. Northern blot analysis revealed that eIF3-p44 (approximately 1.35 kb) is constitutively expressed in many tissues. The essential sequence for this interaction was mapped to the carboxyl-terminus of 4.1R (residues 525-622) and a region (residues 54-321) of eIF3-p44. The direct association between 4.1R and eIF3-p44 was further confirmed by in vitro binding assays and coimmunoprecipitation studies. To characterize the functions of eIF3-p44, we depleted eIF3-p44 from rabbit reticulocyte lysates either by anti-eIF3-p44 antibody or by GST/4.1R-80 fusion protein. Our results show that the eIF3-p44 depleted cell-free translation system was unable to synthesize proteins efficiently. The direct association between 4.1R and elF3-p44 suggests that 4.1R may act as an anchor protein that links the cytoskeleton network to the translation apparatus. (Blood. 2000;96:747-753)

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Antibodies / pharmacology
  • Binding Sites
  • Blotting, Northern
  • Cell-Free System
  • Cloning, Molecular
  • Cytoskeletal Proteins*
  • Cytoskeleton / metabolism*
  • Gene Expression
  • Glutathione Transferase / genetics
  • Humans
  • Immunosorbent Techniques
  • Membrane Proteins / chemistry
  • Membrane Proteins / metabolism*
  • Neuropeptides*
  • Nucleic Acid Hybridization
  • Peptide Initiation Factors / chemistry
  • Peptide Initiation Factors / genetics
  • Peptide Initiation Factors / metabolism*
  • Peptide Mapping
  • Prokaryotic Initiation Factor-3
  • Protein Biosynthesis*
  • Rabbits
  • Recombinant Fusion Proteins / metabolism
  • Saccharomyces cerevisiae / genetics

Substances

  • Antibodies
  • Cytoskeletal Proteins
  • Membrane Proteins
  • Neuropeptides
  • Peptide Initiation Factors
  • Prokaryotic Initiation Factor-3
  • Recombinant Fusion Proteins
  • erythrocyte membrane band 4.1 protein
  • erythrocyte membrane protein band 4.1-like 1
  • Glutathione Transferase