scholarly journals G-Quadruplex Regulation of VEGFA mRNA Translation by RBM4

2022 ◽  
Vol 23 (2) ◽  
pp. 743
Author(s):  
Kangkang Niu ◽  
Xiaojuan Zhang ◽  
Qisheng Song ◽  
Qili Feng
Keyword(s):  
Mrna Translation ◽  
Vascular Endothelial ◽  
Entry Site ◽  
Independent Manner ◽  
Internal Ribosome Entry ◽  
G Quadruplex ◽  
Expression Activity ◽  
Ires Element ◽  
Endothelial Growth Factor ◽  
Vegfa Gene

In eukaryotes, mRNAs translation is mainly mediated in a cap-dependent or cap-independent manner. The latter is primarily initiated at the internal ribosome entry site (IRES) in the 5′-UTR of mRNAs. It has been reported that the G-quadruplex structure (G4) in the IRES elements could regulate the IRES activity. We previously confirmed RBM4 (also known as LARK) as a G4-binding protein in human. In this study, to investigate whether RBM4 is involved in the regulation of the IRES activity by binding with the G4 structure within the IRES element, the IRES-A element in the 5′-UTR of vascular endothelial growth factor A (VEGFA) was constructed into a dicistronic reporter vector, psiCHECK2, and the effect of RBM4 on the IRES activity was tested in 293T cells. The results showed that the IRES insertion significantly increased the FLuc expression activity, indicating that this G4-containing IRES was active in 293T cells. When the G4 structure in the IRES was disrupted by base mutation, the IRES activity was significantly decreased. The IRES activity was notably increased when the cells were treated with G4 stabilizer PDS. EMSA results showed that RBM4 specifically bound the G4 structure in the IRES element. The knockdown of RBM4 substantially reduced the IRES activity, whereas over-expressing RBM4 increased the IRES activity. Taking all results together, we demonstrated that RBM4 promoted the mRNA translation of VEGFA gene by binding to the G4 structure in the IRES.

scholarly journals The vascular endothelial growth factor mRNA contains an internal ribosome entry site

FEBS Letters ◽  
1998 ◽  
Vol 434 (3) ◽  
pp. 417-420 ◽  
Author(s):  
Diane L Miller ◽  
Justin A Dibbens ◽  
Annette Damert ◽  
Werner Risau ◽  
Mathew A Vadas ◽  
...  
Keyword(s):  
Vascular Endothelial Growth Factor ◽  
Growth Factor ◽  
Internal Ribosome Entry Site ◽  
Vascular Endothelial ◽  
Entry Site ◽  
Internal Ribosome Entry ◽  
Endothelial Growth Factor

scholarly journals Two Independent Internal Ribosome Entry Sites Are Involved in Translation Initiation of Vascular Endothelial Growth Factor mRNA

1998 ◽  
Vol 18 (11) ◽  
pp. 6178-6190 ◽  
Author(s):  
Isabelle Huez ◽  
Laurent Créancier ◽  
Sylvie Audigier ◽  
Marie-Claire Gensac ◽  
Anne-Catherine Prats ◽  
...  
Keyword(s):  
Vascular Endothelial Growth Factor ◽  
Growth Factor ◽  
Translation Initiation ◽  
Mrna Translation ◽  
Vascular Endothelial ◽  
Internal Ribosome Entry ◽  
Internal Ribosome Entry Sites ◽  
Polypyrimidine Tract Binding Protein ◽  
Initiation Of Translation ◽  
Endothelial Growth Factor

ABSTRACT The mRNA of vascular endothelial growth factor (VEGF), the major angiogenic growth factor, contains an unusually long (1,038 nucleotides) and structured 5′ untranslated region (UTR). According to the classical translation initiation model of ribosome scanning, such a 5′ UTR is expected to be a strong translation inhibitor. In vitro and bicistronic strategies were used to show that the VEGF mRNA translation was cap independent and occurred by an internal ribosome entry process. For the first time, we demonstrate that two independent internal ribosome entry sites (IRESs) are present in this 5′ UTR. IRES A is located within the 300 nucleotides upstream from the AUG start codon. RNA secondary structure prediction and site-directed mutagenesis allowed the identification of a 49-nucleotide structural domain (D4) essential to IRES A activity. UV cross-linking experiments revealed that IRES A activity was correlated with binding of a 100-kDa protein to the D4 domain. IRES B is located in the first half of the 5′ UTR. An element between nucleotides 379 and 483 is required for its activity. Immunoprecipitation experiments demonstrated that a main IRES B-bound protein was the polypyrimidine tract binding protein (PTB), a well-known regulator of picornavirus IRESs. However, we showed that binding of the PTB on IRES B does not seem to be correlated with its activity. Evidence is provided of an original cumulative effect of two IRESs, probably controlled by different factors, to promote an efficient initiation of translation at the same AUG codon.

scholarly journals Translation of Vascular Endothelial Growth Factor mRNA by Internal Ribosome Entry: Implications for Translation under Hypoxia

1998 ◽  
Vol 18 (6) ◽  
pp. 3112-3119 ◽  
Author(s):  
Ilan Stein ◽  
Ahuva Itin ◽  
Paz Einat ◽  
Rami Skaliter ◽  
Zehava Grossman ◽  
...  
Keyword(s):  
Vascular Endothelial Growth Factor ◽  
Growth Factor ◽  
Translational Regulation ◽  
Start Codon ◽  
Efficient Production ◽  
Vascular Endothelial ◽  
Entry Site ◽  
Internal Ribosome Entry ◽  
Translational Enhancers ◽  
Endothelial Growth Factor

ABSTRACT Vascular endothelial growth factor (VEGF) is a hypoxia-inducible angiogenic growth factor that promotes compensatory angiogenesis in circumstances of oxygen shortage. The requirement for translational regulation of VEGF is imposed by the cumbersome structure of the 5′ untranslated region (5′UTR), which is incompatible with efficient translation by ribosomal scanning, and by the physiologic requirement for maximal VEGF production under conditions of hypoxia, where overall protein synthesis is compromised. Using bicistronic reporter gene constructs, we show that the 1,014-bp 5′UTR of VEGF contains a functional internal ribosome entry site (IRES). Efficient cap-independent translation is maintained under hypoxia, thereby securing efficient production of VEGF even under unfavorable stress conditions. To identify sequences within the 5′UTR required for maximal IRES activity, deletion mutants were analyzed. Elimination of the majority (851 nucleotides) of internal 5′UTR sequences not only maintained full IRES activity but also generated a significantly more potent IRES. Activity of the 163-bp long “improved” IRES element was abrogated, however, following substitution of a few bases near the 5′ terminus as well as substitutions close to the translation start codon. Both the full-length 5′UTR and its truncated version function as translational enhancers in the context of a monocistronic mRNA.

scholarly journals Constitutive translation of human α-synuclein is mediated by the 5′-untranslated region

Open Biology ◽  
2016 ◽  
Vol 6 (4) ◽  
pp. 160022 ◽  
Author(s):  
Pelagia Koukouraki ◽  
Epaminondas Doxakis
Keyword(s):  
Untranslated Region ◽  
Mrna Translation ◽  
Protein Translation ◽  
Serum Starvation ◽  
Mrna Levels ◽  
Entry Site ◽  
Rna Motifs ◽  
Physiological Mechanisms ◽  
Internal Ribosome Entry ◽  
G Quadruplex

Genetic and biochemical studies have established a central role for α-synuclein (SNCA) accumulation in the pathogenesis of Parkinson's disease. Uncovering and subsequently interfering with physiological mechanisms that control SNCA expression is one approach to limit disease progression. To this end, the long and GC-rich 5′-untranslated region (UTR) of SNCA, which is predicted to fold into stable hairpin and G-quadruplex RNA motifs, was investigated for its role in mRNA translation. Inclusion of SNCA 5′-UTR significantly induced expression of both SNCA and luciferase ORF constructs. This effect was not associated with a change in mRNA levels or differential nucleocytoplasmic shuttling. Further, the presence of the 5′-UTR enhanced SNCA synthesis when cap-dependent translation was attenuated with rapamycin treatment. Analysis using multiple methodologies revealed that the 5′-UTR harbours an internal ribosome entry site (IRES) element that spans most of its nucleotide sequence. Signals such as plasma-membrane depolarization, serum starvation and oxidative stress stimulated SNCA protein translation via its 5′-UTR as well as enhanced its IRES activity. Taken together, these data support the idea that the 5′-UTR is an important positive regulator of SNCA synthesis under diverse physiological and pathological conditions, explaining in part the abundance of SNCA in healthy neurons and its accumulation in degenerative cells.

scholarly journals A Small Ligand That Selectively Binds to the G-quadruplex at the Human Vascular Endothelial Growth Factor Internal Ribosomal Entry Site and Represses the Translation

2021 ◽  
Vol 9 ◽  
Author(s):  
Xiao-Xia Hu ◽  
Sheng-Quan Wang ◽  
Shi-Quan Gan ◽  
Lei Liu ◽  
Ming-Qing Zhong ◽  
...  
Keyword(s):  
Vascular Endothelial Growth Factor ◽  
Growth Factor ◽  
Internal Ribosomal Entry Site ◽  
Luciferase Assay ◽  
Ionization Mass ◽  
Vascular Endothelial ◽  
Entry Site ◽  
G Quadruplex ◽  
Cervical Cancer Cells ◽  
Endothelial Growth Factor

G-quadruplexes are believed to have important biological functions, so many small molecules have been screened or developed for targeting G-quadruplexes. However, it is still a major challenge to find molecules that recognize specific G-quadruplexes. Here, by using a combination of surface plasmon resonance, electrospray ionization mass spectrometry, circular dichroism, Western blot, luciferase assay, and reverse transcriptase stop assay, we observed a small molecule, namely, oxymatrine (OMT) that could selectively bind to the RNA G-quadruplex in 5′-untranslated regions (UTRs) of human vascular endothelial growth factor (hVEGF), but could not bind to other G-quadruplexes. OMT could selectively repress the translation of VEGF in cervical cancer cells. Furthermore, it could recognize VEGF RNA G-quadruplexes in special conformations. The results indicate that OMT may serve as a potentially special tool for studying the VEGF RNA G-quadruplex in cells and as a valuable scaffold for the design of ligands that recognize different G-quadruplexes.

scholarly journals BDNF-induced local translation of GluA1 is regulated by HNRNP A2/B1

2020 ◽  
Vol 6 (47) ◽  
pp. eabd2163
Author(s):  
Youngseob Jung ◽  
Ji-Young Seo ◽  
Hye Guk Ryu ◽  
Do-Yeon Kim ◽  
Kyung-Ha Lee ◽  
...  
Keyword(s):  
Ampa Receptor ◽  
Molecular Mechanisms ◽  
Receptor Expression ◽  
Spine Density ◽  
Entry Site ◽  
Independent Manner ◽  
Internal Ribosome Entry ◽  
Ampa Receptor Subunit ◽  
Internal Initiation ◽  
Underlying Mechanisms

The AMPA receptor subunit GluA1 is essential for induction of synaptic plasticity. While various regulatory mechanisms of AMPA receptor expression have been identified, the underlying mechanisms of GluA1 protein synthesis are not fully understood. In neurons, axonal and dendritic mRNAs have been reported to be translated in a cap-independent manner. However, molecular mechanisms of cap-independent translation of synaptic mRNAs remain largely unknown. Here, we show that GluA1 mRNA contains an internal ribosome entry site (IRES) in the 5′UTR. We also demonstrate that heterogeneous nuclear ribonucleoprotein (hnRNP) A2/B1 interacts with GluA1 mRNA and mediates internal initiation of GluA1. Brain-derived neurotrophic factor (BDNF) stimulation increases IRES-mediated GluA1 translation via up-regulation of HNRNP A2/B1. Moreover, BDNF-induced GluA1 expression and dendritic spine density were significantly decreased in neurons lacking hnRNP A2/B1. Together, our data demonstrate that IRES-mediated translation of GluA1 mRNA is a previously unidentified feature of local expression of the AMPA receptor.

scholarly journals The hnRNA-Binding Proteins hnRNP L and PTB Are Required for Efficient Translation of the Cat-1 Arginine/Lysine Transporter mRNA during Amino Acid Starvation

2009 ◽  
Vol 29 (10) ◽  
pp. 2899-2912 ◽  
Author(s):  
Mithu Majumder ◽  
Ibrahim Yaman ◽  
Francesca Gaccioli ◽  
Vladimir V. Zeenko ◽  
Chuanping Wang ◽  
...  
Keyword(s):  
Protein Synthesis ◽  
Amino Acid ◽  
Binding Protein ◽  
Mrna Translation ◽  
Amino Acid Starvation ◽  
Entry Site ◽  
Internal Ribosome Entry ◽  
Polypyrimidine Tract Binding Protein ◽  
Global Protein Synthesis

ABSTRACT The response to amino acid starvation involves the global decrease of protein synthesis and an increase in the translation of some mRNAs that contain an internal ribosome entry site (IRES). It was previously shown that translation of the mRNA for the arginine/lysine amino acid transporter Cat-1 increases during amino acid starvation via a mechanism that utilizes an IRES in the 5′ untranslated region of the Cat-1 mRNA. It is shown here that polypyrimidine tract binding protein (PTB) and an hnRNA binding protein, heterogeneous nuclear ribonucleoprotein L (hnRNP L), promote the efficient translation of Cat-1 mRNA during amino acid starvation. Association of both proteins with Cat-1 mRNA increased during starvation with kinetics that paralleled that of IRES activation, although the levels and subcellular distribution of the proteins were unchanged. The sequence CUUUCU within the Cat-1 IRES was important for PTB binding and for the induction of translation during amino acid starvation. Binding of hnRNP L to the IRES or the Cat-1 mRNA in vivo was independent of PTB binding but was not sufficient to increase IRES activity or Cat-1 mRNA translation during amino acid starvation. In contrast, binding of PTB to the Cat-1 mRNA in vivo required hnRNP L. A wider role of hnRNP L in mRNA translation was suggested by the decrease of global protein synthesis in cells with reduced hnRNP L levels. It is proposed that PTB and hnRNP L are positive regulators of Cat-1 mRNA translation via the IRES under stress conditions that cause a global decrease of protein synthesis.

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