Characterization of eIF3k

Abstract

Mammalian translation initiation factor 3 (eIF3) is a multisubunit complex containing at least 12 subunits with an apparent aggregate mass of ≈ 700 kDa. eIF3 binds to the 40S ribosomal subunit, promotes the binding of methionyl‐tRNA i and mRNA, and interacts with several other initiation factors to form the 40S initiation complex. Human cDNAs encoding 11 of the 12 subunits have been isolated previously; here we report the cloning and characterization of a twelfth subunit, a 28‐kDa protein named eIF3k. Evidence that eIF3k is present in the eIF3 complex was obtained. A monoclonal anti‐eIF3a (p170) Ig coimmunoprecipitates eIF3k with the eIF3 complex. Affinity purification of histidine‐tagged eIF3k from transiently transfected COS cells copurifies other eIF3 subunits. eIF3k colocalizes with eIF3 on 40S ribosomal subunits. eIF3k coexpressed with five other ‘core’ eIF3 subunits in baculovirus‐infected insect cells, forms a stable, immunoprecipitatable, complex with the ‘core’. eIF3k interacts directly with eIF3c, eIF3g and eIF3j by glutathione S ‐transferase pull‐down assays. Sequences homologous with eIF3k are found in the genomes of Caenorhabitis elegans , Arabidopsis thaliana and Drosophila melanogaster , and a homologous protein has been reported to be present in wheat eIF3. Its ubiquitous expression in human tissues, yet its apparent absence in Saccharomyces cerevisiae and Schizosaccharomyces pombe , suggest a unique regulatory role for eIF3k in higher organisms. The studies of eIF3k complete the characterization of mammalian eIF3 subunits.