Armen Parsyan

MicroRNA inhibition of translation initiation in vitro by targeting the cap-binding complex eIF4F

MicroRNAs (miRNAs) play an important role in gene regulatory networks in animals. Yet, the mechanistic details of their function in translation inhibition or messenger RNA (mRNA) destabilization remain controversial. To directly examine the earliest events in this process, we have developed an in vitro translation system using mouse Krebs-2 ascites cell–free extract that exhibits an authentic miRNA response. We show here that translation initiation, specifically the 5′ cap recognition process, is repressed by endogenous let-7 miRNAs within the first 15 minutes of mRNA exposure to the extract when no destabilization of the transcript is observed. Our results indicate that inhibition of translation initiation is the earliest molecular event effected by miRNAs. Other mechanisms, such as mRNA degradation, may subsequently consolidate mRNA silencing.

Control of Cell Survival and Proliferation by Mammalian Eukaryotic Initiation Factor 4B

ABSTRACT Translation initiation plays an important role in cell growth, proliferation, and survival. The translation initiation factor eIF4B (eukaryotic initiation factor 4B) stimulates the RNA helicase activity of eIF4A in unwinding secondary structures in the 5′ untranslated region (5′UTR) of the mRNA in vitro. Here, we studied the effects of eIF4B depletion in cells using RNA interference (RNAi). In agreement with the role of eIF4B in translation initiation, its depletion resulted in inhibition of this step. Selective reduction of translation was observed for mRNAs harboring strong to moderate secondary structures in their 5′UTRs. These mRNAs encode proteins, which function in cell proliferation (Cdc25C, c-myc, and ODC [ornithine decarboxylase]) and survival (Bcl-2 and XIAP [X-linked inhibitor of apoptosis]). Furthermore, eIF4B silencing led to decreased proliferation rates, promoted caspase-dependent apoptosis, and further sensitized cells to camptothecin-induced cell death. These results demonstrate that eIF4B is required for cell proliferation and survival by regulating the translation of proliferative and prosurvival mRNAs.

The helicase protein DHX29 promotes translation initiation, cell proliferation, and tumorigenesis

Translational control plays an important role in cell growth and tumorigenesis. Cap-dependent translation initiation of mammalian mRNAs with structured 5′UTRs requires the DExH-box protein, DHX29, in vitro. Here we show that DHX29 is important for translation in vivo. Down-regulation of DHX29 leads to impaired translation, resulting in disassembly of polysomes and accumulation of mRNA-free 80S monomers. DHX29 depletion also impedes cancer cell growth in culture and in xenografts. Thus, DHX29 is a bona fide translation initiation factor that potentially can be exploited as a target to inhibit cancer cell growth.

eIF4B controls survival and proliferation and is regulated by proto-oncogenic signaling pathways

Messenger RNA translation, or protein synthesis, is a fundamental biological process affecting cell growth, survival and proliferation. Initiation is the rate limiting and hence the most regulated step of translation. In eukaryotes, translation initiation is facilitated by multiple protein factors collectively called eIFs (for eukaryotic translation initiation factors). The complex consisting of the eIF4 group factors including the mRNA cap-binding eIF4E protein, large scaffolding protein eIF4G and RNA helicase eIF4A is assisted by the eIF4B co-factor to unwind local secondary structures and create a ribosome landing pad on mRNA. Recruitment of the ribosome and augmentation in the mRNA scanning process culminates in the positioning of the ribosome over the start codon. Deregulated translational control is believed to play an important role in oncogenic transformation. Indeed, many eIFs are bona fide proto-oncogenes. In many types of human cancers, eIFs are either overexpressed or ectopically activated by Ras-MAPK and PI3K-mTOR signaling cascades, resulting in increased survival and accelerated proliferation. In this review we will analyze the bulk of data describing eIF4B and its role in cell survival and proliferation. Recent studies have shown that eIF4B is phosphorylated and activated by Ras-MAPK and PI3K-mTOR signaling cascades. In addition, eIF4B regulates translation of proliferative and pro-survival mRNAs. Moreover, eIF4B depletion in cancer cells attenuates proliferation, sensitizes them to genotoxic stress driven apoptosis. Taken together, these findings identify eIF4B as a potential target for development of anti-cancer therapies.

Control of Synaptic Plasticity and Memory via Suppression of Poly(A)-Binding Protein

Control of protein synthesis is critical for synaptic plasticity and memory formation. However, the molecular mechanisms linking neuronal activity to activation of mRNA translation are not fully understood. Here, we report that the translational repressor poly(A)-binding protein (PABP)-interacting protein 2A (PAIP2A), an inhibitor of PABP, is rapidly proteolyzed by calpains in stimulated neurons and following training for contextual memory. Paip2a knockout mice exhibit a lowered threshold for the induction of sustained long-term potentiation and an enhancement of long-term memory after weak training. Translation of CaMKIIα mRNA is enhanced in Paip2a⁻/⁻ slices upon tetanic stimulation and in the hippocampus of Paip2a⁻/⁻ mice following contextual fear learning. We demonstrate that activity-dependent degradation of PAIP2A relieves translational inhibition of memory-related genes through PABP reactivation and conclude that PAIP2A is a pivotal translational regulator of synaptic plasticity and memory.

Selective Regulation of GluA Subunit Synthesis and AMPA Receptor-Mediated Synaptic Function and Plasticity by the Translation Repressor 4E-BP2 in Hippocampal Pyramidal Cells

The eukaryotic initiation factor 4E-binding protein-2 (4E-BP2) is a repressor of cap-dependent mRNA translation and a major downstream effector of the mammalian target of rapamycin (mTOR) implicated in hippocampal long-term synaptic plasticity and memory. Yet, synaptic mechanisms regulated by 4E-BP2 translational repression remain unknown. Combining knock-out mice, whole-cell recordings, spine analysis, and translation profiling, we found that 4E-BP2 deletion selectively upregulated synthesis of glutamate receptor subunits GluA1 and GluA2, facilitating AMPA receptor (AMPAR)-mediated synaptic transmission and affecting translation-dependent chemically induced late long-term potentiation (cL-LTP). In 4E-BP2 knock-out (4E-BP2−/−) mice, evoked and miniature EPSCs were increased, an effect mimicked by short-hairpin RNA knockdown of 4E-BP2 in wild-type mice, indicating that 4E-BP2 level regulates basal transmission at mature hippocampal AMPAR-containing synapses. Remarkably, in 4E-BP2−/− mice, the AMPA to NMDA receptor (NMDAR) EPSC ratio was increased, without affecting NMDAR-mediated EPSCs. The enhanced AMPAR function concurred with increased spine density and decreased length resulting from greater proportion of regular spines and less filopodia in 4E-BP2−/− mice. Polysome profiling revealed that translation of GluA1 and GluA2 subunits, but not GluN1 or GluN2A/B, was selectively increased in 4E-BP2−/− hippocampi, consistent with unaltered I–V relation of EPSCs mediated by GluA1/GluA2 heteromers. Finally, translation-dependent cL-LTP of unitary EPSCs was also affected in 4E-BP2−/− mice, lowering induction threshold and removing mTOR signaling requirement while impairing induction by normal stimulation. Thus, translational control through 4E-BP2 represents a unique mechanism for selective regulation of AMPAR synthesis, synaptic function, and long-term plasticity, important for hippocampal-dependent memory processes.

On the diversification of the translation apparatus across eukaryotes

Diversity is one of the most remarkable features of living organisms. Current assessments of eukaryote biodiversity reaches 1.5 million species, but the true figure could be several times that number. Diversity is ingrained in all stages and echelons of life, namely, the occupancy of ecological niches, behavioral patterns, body plans and organismal complexity, as well as metabolic needs and genetics. In this review, we will discuss that diversity also exists in a key biochemical process, translation, across eukaryotes. Translation is a fundamental process for all forms of life, and the basic components and mechanisms of translation in eukaryotes have been largely established upon the study of traditional, so-called model organisms. By using modern genome-wide, high-throughput technologies, recent studies of many nonmodel eukaryotes have unveiled a surprising diversity in the configuration of the translation apparatus across eukaryotes, showing that this apparatus is far from being evolutionarily static. For some of the components of this machinery, functional differences between different species have also been found. The recent research reviewed in this article highlights the molecular and functional diversification the translational machinery has undergone during eukaryotic evolution. A better understanding of all aspects of organismal diversity is key to a more profound knowledge of life.

Influence of preoperative magnetic resonance imaging on the surgical management of breast cancer patients.

BACKGROUND Magnetic resonance imaging (MRI) is gaining popularity in the preoperative management of breast cancer patients. However, the role of this modality remains controversial. We aimed to study the impact of preoperative MRI (pMRI) on the surgical management of breast cancer patients. METHODS This retrospective study included 766 subjects with breast cancer treated operatively at the specialized academic center. RESULTS Between those who underwent pMRI (MRI group, n = 307) and those who did not (no-MRI group, n = 458), there were no significant differences (P = .254) in the proportions of either total mastectomies (20.5% vs 17.2%, respectively) or segmental mastectomies (79.5% vs 82.8%). Patients in the MRI group were significantly more likely (P = .002) to undergo contralateral surgery (11.7% vs 5.5%). Similar results were obtained in multivariate analysis adjusting for age, with the proportions of contralateral breast operations significantly higher in the MRI group (Odds Ratio = 2.25, P = .007). pMRI had no significant effect (P = .54) on the proportion of total re-excisions (7.5% vs 8.7%) or the type of re-excision (total vs segmental mastectomy) between the groups. CONCLUSIONS pMRI does not have a significant impact on the type of operative intervention on the ipsilateral breast but is associated with an increase in contralateral operations. Similarly, pMRI does not change the proportion of re-excisions or the type of the re-excision performed. This study demonstrates that pMRI has little impact on the surgical management of breast cancer, and its value as a routine adjunct in the preoperative work-up of recently diagnosed breast cancer patients needs to be re-examined.

Translation initiation in colorectal cancer

Colorectal cancers (CRC) are one of the most common causes of morbidity and mortality in high-income countries. Targeted screening programs have resulted in early treatment and a substantial decrease in mortality. However, treatment strategies for CRC still require improvement. Understanding the etiology and pathogenesis of CRC would provide tools for improving treatment of patients with this disease. It is only recently that deregulation of the protein synthesis apparatus has begun to gain attention as a major player in cancer development and progression. Among the numerous steps of protein synthesis, deregulation of the process of translation initiation appears to play a key role in cancer growth and proliferation. This manuscript discusses a fascinating and rapidly growing field exploring translation initiation as a fundamental component in CRC development and progression and summarizing CRC treatment perspectives based on agents targeting translation initiation.

Cancers of the Urinary System

The role of translation and its regulation in cancers of the urinary system is poorly studied and understood, and is limited to investigations of the activity of eIF4E and its binding proteins (4E-BPs) in bladder cancer and, to a lesser extent, RCCs. The lack of studies on this topic is surprising given the established role of the mTOR signaling pathway, which converges on the translation machinery, in the pathobiology and therapeutic targeting in RCC. Nevertheless, the available body of evidence suggests involvement of eIF4E overexpression and activation via the mTOR/4E-BPs module in the biology of cancers of the urinary system. This chapter discusses the current state of our knowledge on alterations of the translation machinery in these malignancies.