Nitric Oxide (NO)-mediated Toxicity in Neurodegeneration
Programme Leader: Joern Steinert
Summary of Research Interests
NO signalling has been implicated in several neurodegenerative diseases such as Alzheimer’s (AD), Huntington’s (HD) and Parkinson’s disease (PD), but its exact contribution to neuronal death remains elusive due to the great complexity of downstream nitrergic activities. Elevated Nitric Oxide (NO) levels, as seen in many diseased states, can lead to the formation of cytotoxic peroxynitrite (ONOO-) which in turn can directly modulate a wide range of protein functions via nitration of tyrosine residues (3-Nitrotyrosination/Tyr-NO). Furthermore, at higher concentrations, toxic NO signalling can alter the functioning of proteins in a process known as S-nitrosylation (S-NO). Both signalling pathways have been widely reported, rendering it of similar functional importance to phosphorylation and ubiquitination processes. The key aim of this project is the identification of substrates subjected to this regulation by nitrergic signalling pathways. To date, little is known as to what extent NO-mediated post-translational modifications contribute to or exacerbate disease development and which key modifications cause dysfunctional neuronal signalling. The identification of novel nitrergic signalling pathways in neurodegeneration and correlation with early functional changes before disease onset will allow a better understanding of cytotoxic NO signalling related to disease progression.
Our group investigates cell signalling pathways involved in Nitric Oxide (NO)-induced neurotoxicity and neuroinflammation with the aim to identify putative targets for therapeutic intervention(s).
Key Objectives are
i) to investigate the contribution of cytotoxic NO/ONOO- to synaptic communication and transmitter release using a multi-systemic model approach (mouse, cell culture and Drosophila).
ii) to identify nitrergic mechanisms by which synaptic target proteins are regulated leading to cytotoxicity.
iii) to investigate post-translational control by NO associated with neurodegenerative signalling.
NO signalling pathways
NO is a highly diffusible molecule with a short half-life and is involved in many physiological and pathological processes. In the brain, it modulates neuronal excitability, neurotransmitter release, long-term potentiation and neurovascular coupling. Nanomolar concentrations of NO are enzymatically generated by endothelial (e)NOS and neuronal (n)NOS, whereas the inducible (i)NOS can produce micromolar levels in response to pro-inflammatory stimuli.
NO potentially acts via multiple downstream signalling mechanisms, depending on the concentration, with low levels being neuroprotective and mediate physiological signalling (e.g. neurotransmission or vasodilatation), whereas higher concentrations mediate immune/inflammatory actions and are neurotoxic. After its generation by one of the three isoenzymes (nNOS, eNOS, iNOS) NO is preserved in its molecular structure by a group of spontaneously decomposing endogenous NO donors. The chemistry of NO involves inter-related redox forms (NO., NO+, NO-) with different chemical reactivities towards distinct target groups, thus explaining the great variety of biological actions.
- Robinson SW, Bourgognon J, Breda C, Campesan S, Dinsdale D, Morone N, Mistry R, Smith TM, Guerra-Martin M, Challiss RAJ, Giorgini F & Steinert JR. Nitric oxide-mediated post-translational modifications control neurotransmitter release by modulating complexin farnesylation and enhancing its clamping ability. Plos Biol, in revision.
- Robinson SW, Gutierrez-Olmo M, Martin M, Smith TM, Morone N & Steinert JR (2017). Endogenous nitric oxide synthase activity regulates synaptic transmitter release. Opera Med Physiol, 3 (2): 31-38.
- Bradley SA, Steinert JR (2016). Nitric oxide mediated post-translational modifications: impacts at the synapse. Oxidative Medicine and Cellular Longevity 2016;2016:5681036.
- Bradley SA, Steinert JR (2015). Characterisation and comparison of temporal release profiles of nitric oxide generating donors. J Neurosci Methods 245:116-124
- Peretti D, Bastide A, Radford H, Verity N, Molloy C, Martin MG, Moreno JA, Steinert JR, Smith T, Dinsdale D, Willis AE, Mallucci GR (2015). RBM3 mediates structural plasticity and protective effects of cooling in neurodegeneration. Nature 518(7538):236-9
- Robinson SW, Nugent ML, Dinsdale D & Steinert JR (2014). Prion protein facilitates synaptic vesicle release by enhancing release probability. Hum Mol Genet 23(17):4581-96.
- Tozer AJB, Forsythe ID & Steinert JR (2012). Nitric oxide signalling augments neuronal voltage-gated L-type (CaV1) and P/Q-type (CaV2.1) in the mouse Medial Nucleus of the Trapezoid Body. PLoS One 7(2):e32256.
- Steinert JR, Campesan S, Richards P, Kyriacou CP, Forsythe ID & Giorgini F (2012). Rab11 modulates synaptic dysfunction and behavioural deficits in a Drosophila model of Huntington’s disease. Hum Mol Genet 1(21): 2912-22
- Steinert JR, Robinson SW, Tong H, Haustein MD, Kopp-Scheinpflug C & Forsythe ID (2011). Nitric oxide is an activity-dependent regulator of target neuron intrinsic excitability. Neuron 71, 291-305.
- Steinert JR, Chernova T & Forsythe ID (2010). Nitric Oxide in brain function, dysfunction and dementia. Neuroscientist 16, 435-452.
- Steinert JR, Chernova T & Forsythe ID (2009). Nitric Oxide can alter brain function. Adv Clin Neurosci Rehabil 9, 10-12.
- Steinert JR, Kopp-Scheinpflug C, Baker C, Challiss RAJ, Mistry R, Haustein MD, Griffin SJ, Tong H, Graham BP & Forsythe ID (2008). Nitric oxide is a volume transmitter regulating neuronal excitability in the auditory pathway. Neuron 60, 642-656 *
- Steinert JR, Kuromi H, Hellwig A, Knirr M, Wyatt AW, Kidokoro Y, Schuster CM (2006). Experience-dependent formation and recruitment of large vesicles from reserve pool. Neuron 50, 723-733
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Dr Flaviano Giorgini (University of Leicester)
Dr Ezio Rosato (University of Leicester)
Prof Ian Forsythe (University of Leicester)
Prof Linda Partridge (Max Planck Institute for Biology of Ageing)
Dr Guy Bewick (The University of Aberdeen)
Whole-cell voltage and current clamp recordings in acute brain slices and cultured neurons (Double Patchstar System, Scientifica)
Drosophila CNS patch clamp recordings
Intracellular recordings at Drosophila larvae NMJs (DCC, dSEVC, TEVC)
Calcium imaging using CCD
FM dye imaging
Multiphoton confocal microscopy (LSM 510 with Mai Tai Deep See)
Nitric oxide detection using fluorescent dyes/electrodes (WPI)
Immunocytochemistry, Western Blotting, PCR
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