Ferrero, R. (Roberto)

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    Fine tuning of the unfolded protein response by ISRIB improves neuronal survival in a model of amyotrophic lateral sclerosis
    (2020) Ferrero, R. (Roberto); Toledo, E. (Estefanía); Aragón, T. (Tomás); Baltanas, A. (Ana); Bugallo, R. (Ricardo); Marlin, E. (Elías); Arrasate, M. (Montserrat); Larrea, L. (Laura); Vinueza-Gavilanes, R. (Rodrigo)
    Loss of protein folding homeostasis features many of the most prevalent neurodegenerative disorders. As coping mechanism to folding stress within the endoplasmic reticulum (ER), the unfolded protein response (UPR) comprises a set of signaling mechanisms that initiate a gene expression program to restore proteostasis, or when stress is chronic or overwhelming promote neuronal death. This fate-defining capacity of the UPR has been proposed to play a key role in amyotrophic lateral sclerosis (ALS). However, the several genetic or pharmacological attempts to explore the therapeutic potential of UPR modulation have produced conflicting observations. In order to establish the precise relationship between UPR signaling and neuronal death in ALS, we have developed a neuronal model where the toxicity of a familial ALS-causing allele (mutant G93A SOD1) and UPR activation can be longitudinally monitored in single neurons over the process of neurodegeneration by automated microscopy. Using fluorescent UPR reporters we established the temporal and causal relationship between UPR and neuronal death by Cox regression models. Pharmacological inhibition of discrete UPR processes allowed us to establish the contribution of PERK (PKR-like ER kinase) and IRE1 (inositol-requiring enzyme-1) mechanisms to neuronal fate. Importantly, inhibition of PERK signaling with its downstream inhibitor ISRIB, but not with the direct PERK kinase inhibitor GSK2606414, significantly enhanced the survival of G93A SOD1-expressing neurons. Characterization of the inhibitory properties of both drugs under ER stress revealed that in neurons (but not in glial cells) ISRIB overruled only part of the translational program imposed by PERK, relieving the general inhibition of translation, but maintaining the privileged translation of ATF4 (activating transcription factor 4) messenger RNA. Surprisingly, the fine-tuning of the PERK output in G93A SOD1-expressing neurons led to a reduction of IRE1-dependent signaling. Together, our findings identify ISRIB-mediated translational reprogramming as a new potential ALS therapy.
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    TMEM173 alternative spliced isoforms modulate viral replication through STING pathway
    (American Association of Immunologists, 2018) Rodríguez-García, E. (Estefanía); Ferrero, R. (Roberto); González-Aseguinolaza, G. (Gloria); Ríus-Rocabert, S. (Sergio); Olagüe, C. (Cristina); Larrea, E. (Esther); Nistal-Villan, E. (Estanislao); Prieto, J. (Jesús); Fortes, P. (Puri); Llorens, C. (Carles)
    The innate immune system provides a primary line of defense against pathogens. Stimulator of IFN genes (STING), encoded by the TMEM173 gene, is a critical protein involved in IFN-b induction in response to infection by different pathogens. In this study, we describe the expression of three different alternative-spliced human (h) TMEM173 mRNAs producing STING truncated isoforms 1, 2, and 3 in addition to the full-length wild-type (wt) hSTING. All of the truncated isoforms lack exon 7 and share the N-terminal transmembrane region with wt hSTING. Overexpression of the three STING truncated isoforms failed to induce IFN-b, and they acted as selective pathway inhibitors of wt hSTING even in combination with upstream inducer cyclic-di-GMP-AMP synthase. Truncated isoforms alter the stability of wt hSTING, reducing protein t1/2 to some extent by the induction of proteasome-dependent degradation. Knocking down expression of truncated isoforms increased production of IFN-b by THP1 monocytes in response to intracellular cytosolic DNA or HSV-1 infection. At early stages of infection, viruses like HSV-1 or vesicular stomatitis virus reduced the ratio of full-length wt hSTING/truncated STING isoforms, suggesting the skewing of alternative splicing of STING toward truncated forms as a tactic to evade antiviral responses. Finally, in silico analysis revealed that the human intron–exon gene architecture of TMEM173 (splice sites included) is preserved in other mammal species, predominantly primates, stressing the relevance of alternative splicing in regulating STING antiviral biology.
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    The exposure to uteroplacental insufficiency is associated with activation of unfolded protein response in postnatal life
    (Plos One, 2018) Ferrero, R. (Roberto); Germani, D. (Daniela); Stefanis, C. (Cristiano); Cianfarani, S. (Stefano); Aragón-Amonárriz, T. (Tomás); Deodati, A. (Annalisa); Puglianiello, A. (Antonella); Nobili, V. (Valerio); Argemí, J. (Josepmaria); Alisi, A. (Anna)
    Early life events are associated with the susceptibility to chronic diseases in adult life. Perturbations of endoplasmic reticulum (ER) homeostasis activate the unfolded protein response (UPR), which contributes to the development of metabolic alterations. Our aim was to evaluate liver UPR in an animal model of intrauterine growth restriction (IUGR).
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    Pharmacological inhibition of the integrated stress response accelerates disease progression in an amyotrophic lateral sclerosis mouse model
    (Wiley, 2023) Ferrero, R. (Roberto); Artieda, J. (Julio); Nicolas, M.J. (María Jesús); Pineda-Lucena, A. (Antonio); Valencia, M. (Miguel); Peregrín, N. (Nuria); Aragón, T. (Tomás); Marlin, E. (Elías); Arrasate, M. (Montserrat); Vinueza-Gavilanes, R. (Rodrigo)
    Background and purpose: The integrated stress response (ISR) regulates translation in response to diverse stresses. ISR activation has been documented in amyotrophic lateral sclerosis (ALS) patients and ALS experimental models. In experimental models, both ISR stimulation and inhibition prevented ALS neurodegeneration; however, which mode of ISR regulation would work in patients is still debated. We previously demonstrated that the ISR modulator ISRIB (Integrated Stress Response InhiBitor, an eIF2B activator) enhances survival of neurons expressing the ALS neurotoxic allele SOD1 G93A. Here, we tested the effect of two ISRIB-like eIF2B activators (2BAct and PRXS571) in the disease progression of transgenic SOD1G93A mice. Experimental approach: After biochemical characterization in primary neurons, SOD1G93A mice were treated with 2BAct and PRXS571. Muscle denervation of vulnerable motor units was monitored with a longitudinal electromyographic test. We used a clinical score to document disease onset and progression; force loss was determined with the hanging wire motor test. Motor neuronal survival was assessed by immunohistochemistry. Key results: In primary neurons, 2BAct and PRXS571 relieve the ISR-imposed translational inhibition while maintaining high ATF4 levels. Electromyographic recordings evidenced an earlier and more dramatic muscle denervation in treated SOD1G93A mice that correlated with a decrease in motor neuron survival. Both compounds anticipated disease onset and shortened survival time. Conclusion and implications: 2BAct and PRXS571 anticipate disease onset, aggravating muscle denervation and motor neuronal death of SOD1G93A mice. This study reveals that the ISR works as a neuroprotective pathway in ALS motor neurons and reveals the toxicity that eIF2B activators may display in ALS patients.