Title: To define the molecular mechanisms that determine cellular and organismal longevity
Funder: Biotechnology and Biological Sciences Research Council

Abstract: Axonal longevity. The NAD+ synthetic enzyme Nmnat2 is an essential axonal protein(38) (MC Lab) Nmnat2 must be delivered by axonal transport to sites up to a metre from the neuronal cell body for the cell to avoid entering a novel cell death pathway. This concept is the basis of our proposal that Nmnat2 is a crucial cell survival factor in neurones and we propose that this delivery, like that of other axonal transport cargoes, declines with age. Nmnat2 has a short half-life (<1hr). As axonal transport requires up to two days to deliver proteins to the ends of our longest axons this means processes regulating Nmnat2 transport and stability are paramount to axon survival. The consequences of losing Nmnat2 are a decline in its product, NAD+, and a rise in the substrate, NMN+. Pharmacological and genetic evidence point to a key role for NMN+ accumulation as a toxic intermediate triggering axonal degeneration. In addition to our studies on WldS, an on-going international collaboration recently led us to a new axon protective mutation (“Wld2”). Deletion of MyD88-5, an adapter protein in TLR signalling, preserves injured axons in Drosophila for the entire adult lifespan and extends axon survival in mice by tenfold (Osterloh et al, Science, revised). NAD+ as a cell survival signal. NAD+ is vital in all cell types as a redox cofactor, for Ca2+ signaling (cADPR, NAADP+), transcription and DNA repair (PARPs and sirtuins). Nampt, the rate-limiting enzyme on the NAD+ salvage pathway, promotes stress resistance and extends cellular lifespan(128). Our recent data (SC lab) suggests a novel role for Nampt in repressing apoptosis. Nampt inhibition causes cell cycle arrest, increases the expression of the pro-apoptotic proteins, including Bim, and induces cell death; these effects are rescued by nicotinic acid or NAD+ indicating that they reflect NAD+ depletion. We have shown that Bim promotes cell death arising from growth factor withdrawal with the major Bim splice variant BimEL being regulated by the ERK1/2 pathway. However, these new results potentially define a completely novel mode of regulation in which all Bim splice variants are strongly induced despite activation of ERK1/2 and PKB, two pathways that normally repress Bim. In addition, we find that Nampt inhibition results in the post-translational modification of Nampt; we speculate that this is a novel feedback regulatory event that represents the cell’s attempts to rescue NAD+ synthesis and may be relevant to declining NAD signalling with age. The role of DYRK1b in oxidative defences and cell senescence. Reactive oxygen species (ROS) elicit DNA, lipid and protein damage and ER stress that can drive apoptosis, suppress cell proliferation/growth or cause senescence. ROS damage is a common causal event in cellular and organismal models of ageing and anti-oxidant interventions extend lifespan and healthspan in model organisms Indeed, mutations that activate the anti-oxidant transcription factor Nrf2 in Drosophila increase oxidative stress resistance and extend lifespan(129). Nrf2 drives the expression of anti-oxidant/de-toxification genes including Heme Oxygenase-1 (HMOX1). In new work we have found that the inducible protein kinase DYRK1b, which controls cell cycle progression, also promotes the expression of HMOX1 and 3 other Nrf2 target genes (SC lab). We propose that DYRK1b is a novel regulator of the Nrf2 pathway and acts to protect cells against ROS. MAPK/SAPK signalling pathways in stress responses and stress resistance. ERK1/2, JNK and p38 are activated by ER stressors and oxidants but their role in stress and stress resistance is complex and context dependent. For example, we find that ERK1/2 signalling protects against ER stress-induced death (SC lab) and it can also promote Nrf2-dependent gene expression(129); conversely, ERK1/2 activation can induce ROS production, cell cycle arrest and senescence(130) and is reduced in long-lived mice strains. Similarly, JNK can promote cell death but also increases Drosophila lifespan. Some of these conflicting studies reflect cellular context and that fact that these pathways do not act in isolation but alongside multiple concident pathways in signalling systems We will examine this in Obj 5 (Modelling signalling networks controlling cellular longevity, SC with M. Narita, Cambridge & J. Saez-Rodriguez, EBI). However, we do need to resolve these paradoxes and define the role these pathways play in stress resistance and cell longevity.


Research activity codes: 1.1 NORMAL BIOLOGICAL DEVELOPMENT AND FUNCTIONING
Health categories: GENERIC HEALTH RELEVANCE