Terminal oligopyrimidine (TOP) mRNAs (encoded by the TOP genes) are identified by a sequence of 6–12 pyrimidines at the 5′ end and by a growth-associated translational regulation. All vertebrate genes for the 80 ribosomal proteins and some other genes involved, directly or indirectly, in translation, are TOP genes. Among the numerous translation factors, only eEF1A and eEF2 are known to be encoded by TOP genes, most of the others having not been analyzed. Here, we report a systematic analysis of the human genes for translation factors. Our results show that: (1) all five elongation factors are encoded by TOP genes; and (2) among the initiation and termination factors analyzed, only eIF3e, eIF3f, and eIF3h exhibit the characteristics of TOP genes. Interestingly, these three polypeptides have been recently shown to constitute a specific subgroup among eIF3 subunits. In fact, eIF3e, eIF3f, and eIF3h are the part of the functional core of eIF3 that is not conserved in Saccharomyces cerevisiae. It has been hypothesized that they are regulatory subunits, and the fact that they are encoded by TOP genes may be relevant for their function.

The synthesis of ribosomal proteins (RPs) has long been known to be a process strongly linked to the growth status of the cell. In vertebrates, this coordination is dependent on RP mRNA translational efficiency, which changes according to physiological circumstances. Despite many years of investigation, the trans‐acting factors and the signaling pathways involved in this regulation are still elusive. At the same time, however, new techniques and classic approaches have opened up new perspectives as regards RP regulation and function. In fact, the proteasome seems to play a crucial and unpredicted role in regulating the availability of RPs for subunit assembly. In addition, the study of human ribosomal pathologies and animal models for these diseases has revealed that perturbation in the synthesis and/or function of an RP activates a p53‐dependent stress response. Surprisingly, the effect of the ribosomal stress is more dramatic in specific physiological processes: hemopoiesis in humans, and pigmentation in mice. Moreover, alteration of each RP impacts differently on the development of an organism.

TNF receptor-associated protein 1 (TRAP1), the main mitochondrial member of the heat shock protein (HSP) 90 family, is induced in most tumor types and is involved in the regulation of proteostasis in the mitochondria of tumor cells through the control of folding and stability of selective proteins, such as Cyclophilin D and Sorcin. Notably, we have recently demonstrated that TRAP1 also interacts with the regulatory protein particle TBP7 in the endoplasmic reticulum (ER), where it is involved in a further extra-mitochondrial quality control of nuclear-encoded mitochondrial proteins through the regulation of their ubiquitination/degradation. Here we show that TRAP1 is involved in the translational control of cancer cells through an attenuation of global protein synthesis, as evidenced by an inverse correlation between TRAP1 expression and ubiquitination/degradation of nascent stress-protective client proteins. This study demonstrates for the first time that TRAP1 is associated with ribosomes and with several translation factors in colon carcinoma cells and, remarkably, is found co-upregulated with some components of the translational apparatus (eIF4A, eIF4E, eEF1A and eEF1G) in human colorectal cancers, with potential new opportunities for therapeutic intervention in humans. Moreover, TRAP1 regulates the rate of protein synthesis through the eIF2α pathway either under basal conditions or under stress, favoring the activation of GCN2 and PERK kinases, with consequent phosphorylation of eIF2α and attenuation of cap-dependent translation. This enhances the synthesis of selective stress-responsive proteins, such as the transcription factor ATF4 and its downstream effectors BiP/Grp78, and the cystine antiporter system xCT, thereby providing protection against ER stress, oxidative damage and nutrient deprivation. Accordingly, TRAP1 silencing sensitizes cells to apoptosis induced by novel antitumoral drugs that inhibit cap-dependent translation, such as ribavirin or 4EGI-1, and reduces the ability of cells to migrate through the pores of transwell filters. These new findings target the TRAP1 network in the development of novel anti-cancer strategies.

RPS19 has been identified as the first gene associated with Diamond–Blackfan anemia (DBA), a rare congenital hypoplastic anemia that includes variable physical malformations. It is mutated in ∼25% of the patients although doubts remain as to whether DBA clinical phenotype depends on the ribosomal function of RPS19 or on an extra-ribosomal role or on both. RPS19 mRNAs with mutations that introduce premature stop codons or eliminate it are rapidly turned over by the surveillance mechanisms possibly causing a decrease in the RPS19 protein level. A decrease in RPS19 level has been shown to cause a defect in the maturation of 18S ribosomal RNA. Less clear is the effect of missense mutations in RPS19. With the aim of analyzing the functional features of mutated RPS19, we prepared cDNA constructs expressing RPS19 containing 11 missense mutations and a trinucleotide insertion found in DBA patients. After transfection, we analyzed the following properties of the mutated proteins: (i) protein stability, (ii) subcellular localization and (iii) assembly into ribosomes. Our results indicate that some RPS19 mutations alter the capacity of the protein to localize in nucleolar structure and these mutated RPS19 are very unstable. Moreover, none of the mutated RPS19 analyzed in this study, including those proteins that appear localized into the nucleolus, is able to be assembled into mature ribosome.

The synthesis of adequate amounts of ribosomes is an essential task for the cell. It is therefore not surprising that regulatory circuits exist to organize the synthesis of ribosomal components. It has been shown that defect in ribosome biogenesis (ribosomal stress) induces apoptosis or cell cycle arrest through activation of the tumor suppressor p53. This mechanism is thought to be implicated in the pathophysiology of a group of genetic diseases such as Diamond Blackfan Anemia which are called ribosomopathies. We have identified an additional response to ribosomal stress that includes the activation of eukaryotic translation elongation factor 2 kinase with a consequent inhibition of translation elongation. This leads to a translational reprogramming in the cell that involves the structurally defined group of messengers called terminal oligopyrimidine (TOP) mRNAs which encode ribosomal proteins and translation factors. In fact, while general protein synthesis is decreased by the impairment of elongation, TOP mRNAs are recruited on polysomes causing a relative increase in the synthesis of TOP mRNA-encoded proteins compared to other proteins. Therefore, in response to ribosomal stress, there is a change in the translation pattern of the cell which may help restore a sufficient level of ribosomes.

We have analysed the effect of transcription inhibitors on the polysomal localization of 5′ terminal oligopyrimidine (TOP‐) mRNAs. It is known that, in vertebrates, the translation of this group of mRNAs is regulated according to the growth status of the cell. Mitogenic stimulation of quiescent cells induces a rapid recruitment of TOP mRNAs from translationally inactive light messenger ribonucleoprotein particles to polysomes. It was found that administration of transcription inhibitors to resting cells causes a similar collective translational activation of TOP mRNAs, without affecting global translation. A number of transcription inhibitors were tested in amphibian and mammalian cultured cells. Actinomycin D (act D), cordycepin, and 5,6‐dichloro‐1‐β‐d‐ribofuranosylbenzimidazole caused a similar activation whereas α‐amanitin or low doses of act D did not induce the translational response. Concentrations of act D sufficient to induce TOP mRNA translation also induce 40S ribosomal protein S6 kinases 1 (S6K1) activation. Moreover at these concentrations of act D increased phosphorylation of 4E‐BP1 was also observed, indicating the involvement of FRAP/mTOR. Consistent with this observation, pretreatment of resting cells with rapamycin suppresses the activation of TOP mRNA translation induced by act D. These results indicate that the effect of act D on translation is mediated by the S6Ks through FRAP/mTOR.