The Roles of eIF4G2 in Leaky Scanning and Reinitiation on the Human Dual-Coding POLG mRNA.

Upstream open reading frames (uORFs) are a frequent feature of eukaryotic mRNAs. Upstream ORFs govern main ORF translation in a variety of ways, but, in a nutshell, they either filter out scanning ribosomes or allow downstream translation initiation via leaky scanning or reinitiation. Previous reports concurred that eIF4G2, a long-known but insufficiently studied eIF4G1 homologue, can rescue the downstream translation, but disagreed on whether it is leaky scanning or reinitiation that eIF4G2 promotes. Here, we investigated a unique human mRNA that encodes two highly conserved proteins (POLGARF with unknown function and POLG, the catalytic subunit of the mitochondrial DNA polymerase) in overlapping reading frames downstream of a regulatory uORF. We show that the uORF renders the translation of both POLGARF and POLG mRNAs reliant on eIF4G2. Mechanistically, eIF4G2 enhances both leaky scanning and reinitiation, and it appears that ribosomes can acquire eIF4G2 during the early steps of reinitiation. This emphasizes the role of eIF4G2 as a multifunctional scanning guardian that replaces eIF4G1 to facilitate ribosome movement but not ribosome attachment to an mRNA.

Specific mechanisms of translation initiation in higher eukaryotes: the eIF4G2 story.

The eukaryotic initiation factor 4G2 (eIF4G2, DAP5, Nat1, p97) was discovered in 1997. Over the past two decades, dozens of papers have presented contradictory data on eIF4G2 function. Since its identification, eIF4G2 has been assumed to participate in noncanonical translation initiation mechanisms, but recent results indicate that it can be involved in scanning as well. In particular, eIF4G2 provides leaky scanning through some upstream open reading frames (uORFs), which are typical for long 5' UTRs of mRNAs from higher eukaryotes. It is likely the protein can also help the ribosome overcome other impediments during scanning of the 5' UTRs of animal mRNAs. This may explain the need for eIF4G2 in higher eukaryotes, as many mRNAs that encode regulatory proteins have rather long and highly structured 5' UTRs. Additionally, they often bind to various proteins, which also hamper the movement of scanning ribosomes. This review discusses the suggested mechanisms of eIF4G2 action, denotes obscure or inconsistent results, and proposes ways to uncover other fundamental mechanisms in which this important protein factor may be involved in higher eukaryotes.

Ribosomal leaky scanning through a translated uORF requires eIF4G2.

eIF4G2 (DAP5 or Nat1) is a homologue of the canonical translation initiation factor eIF4G1 in higher eukaryotes but its function remains poorly understood. Unlike eIF4G1, eIF4G2 does not interact with the cap-binding protein eIF4E and is believed to drive translation under stress when eIF4E activity is impaired. Here, we show that eIF4G2 operates under normal conditions as well and promotes scanning downstream of the eIF4G1-mediated 40S recruitment and cap-proximal scanning. Specifically, eIF4G2 facilitates leaky scanning for a subset of mRNAs. Apparently, eIF4G2 replaces eIF4G1 during scanning of 5' UTR and the necessity for eIF4G2 only arises when eIF4G1 dissociates from the scanning complex. In particular, this event can occur when the leaky scanning complexes interfere with initiating or elongating 80S ribosomes within a translated uORF. This mechanism is therefore crucial for higher eukaryotes which are known to have long 5' UTRs with highly frequent uORFs. We suggest that uORFs are not the only obstacle on the way of scanning complexes towards the main start codon, because certain eIF4G2 mRNA targets lack uORF(s). Thus, higher eukaryotes possess two distinct scanning complexes: the principal one that binds mRNA and initiates scanning, and the accessory one that rescues scanning when the former fails.

eIF4G2 balances its own mRNA translation via a PCBP2-based feedback loop.

Poly(rC)-binding protein 2 (PCBP2, hnRNP E2) is one of the most abundant RNA-binding proteins in mammalian cells. In humans, it exists in seven isoforms, which are assumed to play similar roles in cells. The protein is shown to bind 3'-untranslated regions (3'-UTRs) of many mRNAs and regulate their translation and/or stability, but nothing is known about the functional consequences of PCBP2 binding to 5'-UTRs. Here we show that the PCBP2 isoform f interacts with the 5'-UTRs of mRNAs encoding eIF4G2 (a translation initiation factor with a yet unknown mechanism of action, also known as DAP5) and Cyclin I, and inhibits their translation in vitro and in cultured cells, while the PCBP2 isoform e only affects Cyclin I translation. Furthermore, eIF4G2 participates in a cap-dependent translation of the PCBP2 mRNA. Thus, PCBP2 and eIF4G2 seem to regulate one another's expression via a novel type of feedback loop formed by the translation initiation factor and the RNA-binding protein.