Lentiviral Vectors to Study the Differential Function of ERK1 and ERK2 MAP Kinases

Indrigo, Marzia; Papale, Alessandro; Orellana, Daniel; Brambilla, Riccardo

doi:10.1007/978-1-60761-795-2_12

Cited by 22 publications

(20 citation statements)

References 42 publications

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“…In an effort to translate these findings into a clinically relevant therapy, we set out to determine (i) whether a partial inhibition of Ras-GRF1 in the striatum can revert already established dyskinesias and (ii) whether the therapeutic effect also can be seen in an animal model that better recapitulates the human condition. Thus, we generated lentiviral vectors (LV) that express two dominant-negative constructs for Ras-GRF1 under the control of the strong and ubiquitous human phosphoglycerol kinase (PGK) promoter in addition to the GFP marker (28,29). These two constructs were designed using the human sequence of Ras-GRF1 and were tested in the mouse striatum (Fig.…”

Section: Reversion Of Lid In An Nhp Model By a Combined Genetic Inhibmentioning

confidence: 99%

Inhibition of Ras-guanine nucleotide-releasing factor 1 (Ras-GRF1) signaling in the striatum reverts motor symptoms associated with l -dopa–induced dyskinesia

Fasano

Bézard

D'Antoni

et al. 2010

Proc. Natl. Acad. Sci. U.S.A.

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141

154

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L-dopa-induced dyskinesia (LID) is a common debilitating complication of dopamine replacement therapy in Parkinson's disease. Recent evidence suggests that LID may be linked causally to a hyperactivation of the Ras-ERK signaling cascade in the basal ganglia. We set out to determine whether specific targeting of Ras-guanine nucleotide-releasing factor 1 (Ras-GRF1), a brain-specific activator of the Ras-ERK pathway, may provide a therapy for LID. On the rodent abnormal involuntary movements scale, Ras-GRF1-deficient mice were significantly resistant to the development of dyskinesia during chronic L-dopa treatment. Furthermore, in a nonhuman primate model of LID, lentiviral vectors expressing dominant negative forms of Ras-GRF1 caused a dramatic reversion of dyskinesia severity leaving intact the therapeutic effect of Ldopa. These data reveal the central role of Ras-GRF1 in governing striatal adaptations to dopamine replacement therapy and validate a viable treatment for LID based on intracellular signaling modulation. P arkinson's disease (PD) is a neurodegenerative disorder characterized by a loss of dopaminergic neurons in the substantia nigra pars compacta (SNc) causing dopamine depletion in the striatum, the main input nucleus of the basal ganglia. Dopamine replacement therapy with L-dopa remains the most effective treatment for PD, but its use is associated with motor fluctuations and abnormal involuntary movements (AIMs), termed "L-dopa-induced dyskinesia" (LID), which are doselimiting and potentially disabling (1-3).A key objective for the future treatment of PD is to avoid dyskinesia altogether, but doing so will require an understanding of the molecular mechanisms that are involved. LID is attributed to a sequence of events, largely occurring in the striatum, that include pulsatile stimulation of dopamine D1 receptors, downstream changes in proteins and genes, dendritic alterations, and functional abnormalities in nondopaminergic transmitter systems, all of which concur to modify neuronal firing patterns in the basal ganglia-thalamocortical networks (1-4). Once symptoms have appeared, LID can be triggered easily by a single dose of L-dopa even after several weeks of treatment washout (4). Regulation of striatal gene expression is the likely mechanism underlying neuronal plasticity in LID. The ERK signaling cascade is a key regulator of striatal plasticity and an interesting candidate for drug targeting (5-8). Stimulation of dopamine and glutamate receptors on striatal neurons can switch on the small GTPases of the Ras family, which in turn activate the Raf/Mek/ Erk protein kinase cascade (5-8). Sustained activation of these biochemical pathways leads to synaptic rearrangements requiring de novo gene expression and protein synthesis. Importantly, in neurotoxic models of PD, such as the unilaterally 6-hydroxydopamine (6-OHDA)-lesioned rodent and the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated nonhuman primate (NHP), a supersensitivity of striatal D1 receptors leads to ERK hyperactivation in ...

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Section: Reversion Of Lid In An Nhp Model By a Combined Genetic Inhibmentioning

confidence: 99%

Inhibition of Ras-guanine nucleotide-releasing factor 1 (Ras-GRF1) signaling in the striatum reverts motor symptoms associated with l -dopa–induced dyskinesia

Fasano

Bézard

D'Antoni

et al. 2010

Proc. Natl. Acad. Sci. U.S.A.

Self Cite

141

154

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“…This is due to a de-repression of ERK2, the main MAP kinase. In normal conditions, ERK1 acts as a built-in partial agonist, keeping ERK2 activity tightly regulated (Vantaggiato et al, 2006; Indrigo et al, 2010). Despite this general effect of the biochemistry of ERK2, ERK1 mutant animals do not show any sign of cognitive deterioration as one would have predicted from the early studies on the NF1 KO mice.…”

Section: The Ras–erk Paradox: Balancing Synaptic Excitation and Inhibmentioning

confidence: 99%

Ras?ERK Signaling in Behavior: Old Questions and New Perspectives

Fasano¹,

Brambilla²

2011

Front. Behav. Neurosci.

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The role of Ras–ERK signaling in behavioral plasticity is well established. Inhibition studies using the blood–brain barrier permeable drug SL327 have conclusively demonstrated that this neuronal cell signaling cascade is a crucial component of the synaptic machinery implicated in the formation of various forms of long-term memory, from spatial learning to fear and operant conditioning. However, abnormal Ras–ERK signaling has also been linked to a number of neuropsychiatric conditions, including mental retardation syndromes (“RASopathies”), drug addiction, and l-DOPA induced dyskinesia (LID). The work recently done on these brain disorders has pointed to previously underappreciated roles of Ras–ERK in specific subsets of neurons, like GABAergic interneurons of the hippocampus or the cortex, as well as in the medium spiny neurons of the striatum. Here we will highlight the open questions related to Ras–ERK signaling in these behavioral manifestations and propose crucial experiments for the future.

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“…Generation and production of Lentiviral (LV) constructs for RNAi of ERK1 and ERK2 (both in shRNA and mir RNA configuration) as well as LV overexpressing ERK1, ERK2 and their swapped counterparts have been previously described 8,54 . Briefly, third generation non-replicative LVs were modifications of the originally described backbone 55 .…”

Section: Methodsmentioning

confidence: 99%

“…MEF cultures were prepared from wild type E13.5 embryos obtained from C57Bl/6 mice as previously described 54 . Then MEF were fixed 3 or 5 days after LV infection in PFA 4% for 25 minutes at RT, rinsed in PBS 1X and then stored in 70% ethanol at −20°C.…”

Section: Methodsmentioning

confidence: 99%

Modulation of ERK1/MAPK3 potentiates ERK nuclear signalling, facilitates neuronal cell survival and improves memory in mouse models of neurodegenerative disorders

Indrigo

Morella

Orellana

et al. 2018

Preprint

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Cell signalling mechanisms are central to neuronal activity and their dysregulation may lead to neurodegenerative processes and associated cognitive decline. So far, a major effort has been directed toward the dissection of disease specific pathways with the still unmet promise to develop precision medicine strategies. With a different approach, here we show that a selective genetic potentiation of neuronal ERK signalling prevents cell death in vitro and in vivo in the mouse brain while ERK attenuation does the opposite. This neuroprotective effect can also be induced pharmacologically by a cell permeable peptide mimicking the loss of ERK1 MAP kinase, leading to a selective enhancement of ERK2 mediated nuclear cell signalling. The drug treatment prevents neurodegeneration in mouse models of Huntington’s (HD), Alzheimer’s (AD), and Parkinson’s disease (PD). Importantly, the selective potentiation of ERK2 signalling facilitates both structural and synaptic plasticity, enhances cognition in healthy mice and rescues mild cognitive impairments in both models of AD and HD. Altogether, our observation truly represents a remarkable example of a shared molecular mechanism across multiple neurodegenerative disorders and a potentially valuable therapeutic target for neuro-enhancement.

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Lentiviral Vectors to Study the Differential Function of ERK1 and ERK2 MAP Kinases

Cited by 22 publications

References 42 publications

Inhibition of Ras-guanine nucleotide-releasing factor 1 (Ras-GRF1) signaling in the striatum reverts motor symptoms associated with l -dopa–induced dyskinesia

Inhibition of Ras-guanine nucleotide-releasing factor 1 (Ras-GRF1) signaling in the striatum reverts motor symptoms associated with l -dopa–induced dyskinesia

Ras?ERK Signaling in Behavior: Old Questions and New Perspectives

Modulation of ERK1/MAPK3 potentiates ERK nuclear signalling, facilitates neuronal cell survival and improves memory in mouse models of neurodegenerative disorders

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