Two Arabidopsis Loci Encode Novel Eukaryotic Initiation Factor 4E Isoforms That Are Functionally Distinct from the Conserved Plant Eukaryotic Initiation Factor 4E    

Abstract

Canonical translation initiation in eukaryotes begins with the Eukaryotic Initiation Factor 4F (eIF4F) complex, made up of eIF4E, which recognizes the 7-methylguanosine cap of messenger RNA, and eIF4G, which serves as a scaffold to recruit other translation initiation factors that ultimately assemble the 80S ribosome. Many eukaryotes have secondary EIF4E genes with divergent properties. The model plant Arabidopsis (Arabidopsis thaliana) encodes two such genes in tandem loci on chromosome 1, EIF4E1B (At1g29550) and EIF4E1C (At1g29590). This work identifies EIF4E1B/EIF4E1C-type genes as a Brassicaceae-specific diverged form of EIF4E. There is little evidence for EIF4E1C gene expression; however, the EIF4E1B gene appears to be expressed at low levels in most tissues, though microarray and RNA Sequencing data support enrichment in reproductive tissue. Purified recombinant eIF4E1b and eIF4E1c proteins retain cap-binding ability and form functional complexes in vitro with eIF4G. The eIF4E1b/eIF4E1c-type proteins support translation in yeast (Saccharomyces cerevisiae) but promote translation initiation in vitro at a lower rate compared with eIF4E. Findings from surface plasmon resonance studies indicate that eIF4E1b and eIF4E1c are unlikely to bind eIF4G in vivo when in competition with eIF4E. This study concludes that eIF4E1b/eIF4E1c-type proteins, although bona fide cap-binding proteins, have divergent properties and, based on apparent limited tissue distribution in Arabidopsis, should be considered functionally distinct from the canonical plant eIF4E involved in translation initiation.

Mechanism of Cytoplasmic mRNA Translation

“…A number of forms of eIF4E have evolved in other organisms with special functions such as recognition of tri-methylated caps or tissuespecific regulation; however, some of these proteins have lost either their ability to bind m 7 G or eIF4G poising them as potential inhibitors or repressors of initiation (Joshi et al, 2005;Rhoads, 2009). eIF4E1b and eIF4E1c from Arabidopsis were found to bind eIF4G and to m 7 G-affinity resin and thus function biochemically in vitro as canonical cap-binding proteins (Patrick et al, 2014;Kropiwnicka et al, 2015). However, the double mutant eIF4E eIFiso4E is lethal suggesting that eIF4E1b and eIF4E1c are not able to replace eIF4E in vivo (Callot and Gallois, 2014;Patrick et al, 2014).…”

“…eIF4E1b and eIF4E1c from Arabidopsis were found to bind eIF4G and to m 7 G-affinity resin and thus function biochemically in vitro as canonical cap-binding proteins (Patrick et al, 2014;Kropiwnicka et al, 2015). However, the double mutant eIF4E eIFiso4E is lethal suggesting that eIF4E1b and eIF4E1c are not able to replace eIF4E in vivo (Callot and Gallois, 2014;Patrick et al, 2014).…”

“…Arabidopsis thaliana has three genes for the class 1 eIF4E (eIF4E, eIF4E1b, eIF4E1c) and one gene for eIFiso4E (Patrick and Browning, 2012;Patrick et al, 2014). eIF4E is the most highly expressed; the other two eIF4E genes (eIF4E1b, eIF4E1c) correspond to a tandem duplication event within the Brassicaceae with evidence that only eIF4E1b generates a transcript (Patrick and Browning, 2012) (see Table 1 for accession numbers).…”

“…All higher plants have three forms of the cap-binding proteins, eIF4E, eIFiso4E (plant-specific) and 4EHP (4E homologous protein) which all bind to m 7 GTP-Sepharose (Ruud et al, 1998;Patrick et al, 2014;Kropiwnicka et al, 2015). eIF4E and eIFiso4E are both class 1 cap-binding proteins (Joshi et al, 2005) and form the eIF4F and eIFiso4F complexes with their respective subunits, eIF4G and eIFiso4G (Mayberry et al, 2011;Patrick and Browning, 2012).…”

Eukaryotic Initiation Factor eIFiso4G1 and eIFiso4G2 Are Isoforms Exhibiting Distinct Functional Differences in Supporting Translation in Arabidopsis

“…As the eif4e triple null mutant is not viable (26), it could not be included. Progeny homozygous for the Luc transgene and null for the initiation factor were isolated.…”