Different modes of translation for hid, grim and sickle mRNAs in Drosophila

Abstract

Protein synthesis is inhibited during apoptosis. However, the translation of many mRNAs still proceeds driven by internal ribosome entry sites (IRESs). Here we show that the 5 0 UTR of hid and grim mRNAs promote translation of uncapped-mRNA reporters in cell-free embryonic extracts and that hid and grim mRNA 5 0 UTRs drive IRES-mediated translation. The translation of capped-reporters proceeds in the presence of cap competitor and in extracts where cap-dependent translation is impaired. We show that the endogenous hid and grim mRNAs are present in polysomes of heat-shocked embryos, indicating that cap recognition is not required for translation. In contrast, sickle mRNA is translated in a cap-dependent manner in all these assays. Our results show that IRES-dependent initiation may play a role in the translation of Drosophila proapoptotic genes and suggest a variety of regulatory pathways.

Strong Eukaryotic IRESs Have Weak Secondary Structure

“…Previous publications [8], [11] investigated the IRES activity of the 5′-UTR sequences in five Drosophila melanogaster protein-coding genes, rpr, hsp70, hid, grim and skl , as well as the IRES activity of the reverse complement of the 5′-UTR sequences. We analyzed the 60 nt segment immediately upstream of the initiation codon of these genes and their reverse complements.…”

“…We found that the strength of IRES activity of the fruit fly IRESs is also strongly associated with weak secondary structure (Figure 2, r = −0.8461, p = 0.001), consistent with the pattern observed with yeast data in Figure 1. The reverse complements of the 60 nt 5′-UTR sequences in the five fruit fly genes showed little or no IRES activity [8], [11] and all exhibited relatively stable secondary structure (Figure 2). The observation that the skl sequence with the least IRES activity also has the most stable secondary structure supports the general pattern (Figure 2).…”

“…This result is perhaps not surprising. The yeast IRESs were identified in response to the same stress (starvation induced differentiation) [15], and the Drosophila IRESs are found within a group of genes with similar function [8], [11]. It is therefore possible that these IRESs evolved under similar selective pressures and share a common feature such as weak secondary structure.…”

“…The annotated Drosophila melanogaster genome, dated May 14, 2008, was also downloaded from GenBank. For the five D. melanogaster whose 5′-UTR and the associated reverse complements have been studied for IRES activity [8], [11], we extracted the 60 nt immediately upstream of the initiation AUG and obtained their reverse complements. The sequences (60 nt immediately upstream of the initiation AUG and their reverse complements), are shown in Table S2 in supplementary online material.…”

Alternative Mechanisms to Initiate Translation in Eukaryotic mRNAs

“…Other features such as long length of the 5′UTR (200 to 500 nucleotides), heavy RNA structure, high GC content, initiation at non-AUG codons, ignored AUGs upstream of the functional start codon are often but not universally found [46, 53, 76, 77]. For instance, some IRES elements found in plant RNA viruses, D. melanogaster and S. cerevisiae, have a high AU content [37, 62, 63, 73]. In this regard, the 5′UTR sequences of mRNAs are highly divergent [78].…”