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

Abstract

The eukaryotic translation initiation factor (eIF) 4G is required during protein synthesis to promote the assembly of several factors involved in the recruitment of a 40S ribosomal subunit to an mRNA. Although many eukaryotes express two eIF4G isoforms that are highly similar, the eIF4G isoforms in plants, referred to as eIF4G and eIFiso4G, are highly divergent in size, sequence, and domain organization but both can interact with eIF4A, eIF4B, eIF4E isoforms, and the poly(A)-binding protein.Nevertheless, eIF4G and eIFiso4G from wheat exhibit preferences in the mRNAs they translate optimally. For example, mRNA containing the 5-leader (called ⍀) of tobacco mosaic virus preferentially uses eIF4G in wheat germ lysate. In this study, the eIF4G isoform specificity of ⍀ was used to examine functional differences of the eIF4G isoforms in Arabidopsis. As in wheat, ⍀-mediated translation was reduced in an eif4g null mutant. Loss of the eIFiso4G1 isoform, which is similar in sequence to wheat eIFiso4G, did not substantially affect ⍀-mediated translation. However, loss of the eIFiso4G2 isoform substantially reduced ⍀-mediated translation. eIFiso4G2 is substantially divergent from eIFiso4G1 and is present only in the Brassicaceae, suggesting a recent evolution. eIFiso4G2 isoforms exhibit sequence-specific differences in regions representing partner protein and RNA binding sites. Loss of any eIF4G isoform also resulted in a substantial reduction in reporter transcript level. These results suggest that eIFiso4G2 appeared late in plant evolution and exhibits more functional similarity with eIF4G than with eIFiso4G1 during ⍀-mediated translation.The synthesis of proteins from cellular mRNAs requires several translation initiation factors that assist in recruiting the 40S ribosomal subunit to an mRNA as well as in the recognition of the initiation codon and in the assembly of the 80S ribosome (1-3). Eukaryotic initiation factor (eIF) 4F is essential to promote 40S subunit binding to an mRNA and to assist in 40S subunit scanning of the 5Ј-leader in search of the initiation codon. eIF4F is a multisubunit factor composed of eIF4E, which binds the 5Ј-cap structure; the RNA helicase, eIF4A, which unwinds the secondary structure in a 5Ј-leader that would otherwise inhibit 40S subunit scanning; and the scaffolding protein, eIF4G, which interacts with eIF4E, eIF4A, eIF4B (which stimulates the RNA helicase activity of eIF4A), eIF3 (required for 40 S binding to an mRNA), and the poly(A)-binding protein (PABP) 2 (2, 4 -6). The interactions between eIF4G and eIF4E and between eIF4G and PABP results in mRNA circularization that stimulates translation by promoting 40S subunit recruitment (7,8). An interaction between eIF4B and PABP further increases the affinity for poly(A) RNA of PABP (9 -11). These factors, therefore, are necessary for the efficient translation of most mRNAs.Although plants express two forms of eIF4G as do many eukaryotes (12), the plant isoforms are more divergent from each other than observed between eIF4G isoforms in othe...

Translation regulation in plants: an interesting past, an exciting present and a promising future

“…Finally, plants also have an alternative version of eIF4G, eIFiso4G, so in addition to the universal eIF4F, plants make a second, plant-specific, cap-binding complex called eIFiso4F (Patrick and Browning, 2012). Research suggests that the two complexes have different affinities for certain mRNAs, providing a potential mechanism to regulate the translation of specific mRNA populations (Mayberry et al, 2009;Mart ınez-Silva et al, 2012;Gallie, 2016).…”

“…There are five PAO genes in Arabidopsis, and three of them, PAO2, PAO3 and PAO4, possess uORFs that are conserved in a wide array of plant species, ranging from mosses to monocots and dicots (Vaughn et al, 2012;Guerrero-Gonz alez et al, 2014). PAO2 has been studied in more detail, and the peptide encoded by its uORF was found to exert control by repressing translation of the mORF in the presence of exogenously added polyamines (Guerrero-Gonz alez et al, 2014, 2016.…”

Non-canonical Translation in Plant RNA Viruses

“…is recognized by the heat shock protein 101 (HSP101), mediating translational activity (Wells et al, 1998) and interacts with eIF4F via eIF4G (Gallie, 2002(Gallie, , 2016. Similarly, the Brassicaceae-specific eIFiso4G2 isoform also contributes in -mediated translation, unlike eIFiso4G which did not affectdependent translation (Gallie, 2016).…”

Class II members of the poly(A) binding protein family exhibit distinct functions during Arabidopsis growth and development

“…12 To determine the extent to which any class II PABP member might contribute to gene expression, the expression from the firefly luciferase (Luc) reporter gene was examined in each class II PABP mutant. To this end, Arabidopsis containing the Luc cDNA under the control of the CaMV promoter as described previously 26 was crossed with the class II single and double PABP mutants. Progeny homozygous for the Luc transgene and null for each class II PABP mutation, either as single or double mutants, were isolated.…”

“…27,28 Arabidopsis expresses 2 eIFiso4G isoforms (eIFiso4G1 and eIFiso4G2) as do other species throughout the Brassicaceae whereas species outside the Brassicaceae express only eIFiso4G1 isoforms. 26 eIFiso4G1 and eIFiso4G2 exhibit substantial sequence divergence in those species that express both isoforms. Translation from Luc mRNA containing the 5 0 -leader sequence from tobacco mosaic virus (TMV) (known as V) preferentially uses eIF4G and eIFisoG2 over eIFisoG1 in Arabidopsis.…”