Cell type-specific modulation of healthspan by Forkhead family transcription factors in the nervous system

Reduced activity of insulin/insulin-like growth factor signaling (IIS) increases healthy lifespan among diverse animal species. Downstream of IIS, multiple evolutionarily conserved transcription factors (TFs) are required; however, distinct TFs are likely responsible for these effects in different tissues. Here we have asked which TFs can extend healthy lifespan within distinct cell types of the adult nervous system in Drosophila. Starting from published single-cell transcriptomic data, we report that forkhead (FKH) is endogenously expressed in neurons, whereas forkhead-box-O (FOXO) is expressed in glial cells. Accordingly, we find that neuronal FKH and glial FOXO exert independent prolongevity effects. We have further explored the role of neuronal FKH in a model of Alzheimer’s disease-associated neuronal dysfunction, where we find that increased neuronal FKH preserves behavioral function and reduces ubiquitinated protein aggregation. Finally, using transcriptomic profiling, we identify Atg17, a member of the Atg1 autophagy initiation family, as one FKH-dependent target whose neuronal overexpression is sufficient to extend healthy lifespan. Taken together, our results underscore the importance of cell type-specific mapping of TF activity to preserve healthy function with age

Activating transcription factor 4-dependent lactate dehydrogenase activation as a protective response to amyloid beta toxicity

Accumulation of amyloid beta peptides is thought to initiate the pathogenesis of Alzheimer’s disease. However, the precise mechanisms mediating their neurotoxicity are unclear. Our microarray analyses show that, in Drosophila models of amyloid beta 42 toxicity, genes involved in the unfolded protein response and metabolic processes are upregulated in brain. Comparison with the brain transcriptome of early-stage Alzheimer’s patients revealed a common transcriptional signature, but with generally opposing directions of gene expression changes between flies and humans. Among these differentially regulated genes, lactate dehydrogenase (Ldh) was up-regulated by the greatest degree in amyloid beta 42 flies and the human orthologs (LDHA and LDHB) were down-regulated in patients. Functional analyses revealed that either over-expression or inhibition of Ldh by RNA interference (RNAi) slightly exacerbated climbing defects in both healthy and amyloid beta 42-induced Drosophila. This suggests that metabolic responses to lactate dehydrogenase must be finely-tuned, and that its observed upregulation following amyloid beta 42 production could potentially represent a compensatory protection to maintain pathway homeostasis in this model, with further manipulation leading to detrimental effects. The increased Ldh expression in amyloid beta 42 flies was regulated partially by unfolded protein response signalling, as ATF4 RNAi diminished the transcriptional response and enhanced amyloid beta 42-induced climbing phenotypes. Further functional studies are required to determine whether Ldh upregulation provides compensatory neuroprotection against amyloid beta 42-induced loss of activating transcription factor 4 activity and endoplasmatic reticulum stress.Our study thus reveals dysregulation of lactate dehydrogenase signalling in Drosophila models and patients with Alzheimer’s disease, which may lead to a detrimental loss of metabolic homeostasis. Importantly, we observed that down-regulation of ATF4-dependent endoplasmic reticulum-stress signalling in this context appears to prevent Ldh compensation and to exacerbate amyloid beta 42-dependent neuronal toxicity. Our findings therefore suggest caution in the use of therapeutic strategies focused on down-regulation of this pathway for treatment of Alzheimer’s disease, since its natural response to the toxic peptide may induce beneficial neuroprotective effects

The neuronal receptor tyrosine kinase Alk is a target for longevity

Inhibition of signalling through several receptor tyrosine kinases (RTKs), including the insulin-like growth factor receptor and its orthologues, extends healthy lifespan in organisms from diverse evolutionary taxa. This raises the possibility that other RTKs, including those already well studied for their roles in cancer and developmental biology, could be promising targets for extending healthy lifespan. Here, we focus on anaplastic lymphoma kinase (Alk), an RTK with established roles in nervous system development and in multiple cancers, but whose effects on aging remain unclear. We find that several means of reducing Alk signalling, including mutation of its ligand jelly belly (jeb), RNAi knock-down of Alk, or expression of dominant-negative Alk in adult neurons, can extend healthy lifespan in female, but not male, Drosophila. Moreover, reduced Alk signalling preserves neuromuscular function with age, promotes resistance to starvation and xenobiotic stress, and improves night sleep consolidation. We find further that inhibition of Alk signalling in adult neurons modulates the expression of several insulin-like peptides, providing a potential mechanistic link between neuronal Alk signalling and organism-wide insulin-like signalling. Finally, we show that TAE-684, a small molecule inhibitor of Alk, can extend healthy lifespan in Drosophila, suggesting that the repurposing of Alk inhibitors may be a promising direction for strategies to promote healthy aging

Evidence for involvement of the alcohol consumption WDPCP gene in lipid metabolism, and liver cirrhosis.

Acknowledgements: R.P. was supported by Rutherford Fund fellowship from the Medical Research Council (MR/R026505/1 and MR/R026505/2). B.A., X.J., and F.O. were supported by Rutherford Fund from Medical Research Council MR/R026505/2. R.M. was funded by the President’s PhD Scholarship from Imperial College London. PE is Director of the MRC Centre for Environment and Health and acknowledges support from the Medical Research Council (MR/S019669/1). PE also acknowledges support from the UK Dementia Research Institute, Imperial College London (UKDRI-5001), Health Data Research UK London (HDRUK-1004231) and the British Heart Foundation Imperial College London Centre for Research Excellence (BHF-RE/18/4/34215). The Airwave Health Monitoring Study was funded by the UK Home Office (780- TETRA, 2003-2018) and is currently funded by the MRC and ESRC (MR/R023484/1) with additional support from the NIHR Imperial College Biomedical Research Centre in collaboration with Imperial College NHS Healthcare Trust. R.C.P is supported by the UK Dementia Research Institute (UKDRI-5001), which receives its funding from UK DRI Ltd, funded by the UK Medical Research Council, Alzheimer’s Society and Alzheimer’s Research UK. Work in LMM’s laboratory is supported by the UK Medical Research Council, intramural project MC_UU_00025/3 (RG94521). The views expressed are those of the authors and not necessarily those of the sponsors. We thank Prof. Ulrike Heberlein, (Janelia Research Campus, Virginia, USA) for generously providing us the hppy17-51 fly lines. This research was funded, in whole or in part, by the Medical Research Council (MR/R026505/1 and MR/R026505/2). A CC BY or equivalent licence is applied to the Author Accepted Manuscript (AAM) arising from this submission, in accordance with the grant’s open access conditions.Biological pathways between alcohol consumption and alcohol liver disease (ALD) are not fully understood. We selected genes with known effect on (1) alcohol consumption, (2) liver function, and (3) gene expression. Expression of the orthologs of these genes in Caenorhabditis elegans and Drosophila melanogaster was suppressed using mutations and/or RNA interference (RNAi). In humans, association analysis, pathway analysis, and Mendelian randomization analysis were performed to identify metabolic changes due to alcohol consumption. In C. elegans, we found a reduction in locomotion rate after exposure to ethanol for RNAi knockdown of ACTR1B and MAPT. In Drosophila, we observed (1) a change in sedative effect of ethanol for RNAi knockdown of WDPCP, TENM2, GPN1, ARPC1B, and SCN8A, (2) a reduction in ethanol consumption for RNAi knockdown of TENM2, (3) a reduction in triradylglycerols (TAG) levels for RNAi knockdown of WDPCP, TENM2, and GPN1. In human, we observed (1) a link between alcohol consumption and several metabolites including TAG, (2) an enrichment of the candidate (alcohol-associated) metabolites within the linoleic acid (LNA) and alpha-linolenic acid (ALA) metabolism pathways, (3) a causal link between gene expression of WDPCP to liver fibrosis and liver cirrhosis. Our results imply that WDPCP might be involved in ALD

The neuronal receptor tyrosine kinase Alk is a target for longevity

Inhibition of signalling through several receptor tyrosine kinases (RTKs), including the insulin-like growth factor receptor and its orthologues, extends healthy lifespan in organisms from diverse evolutionary taxa. This raises the possibility that other RTKs, including those already well studied for their roles in cancer and developmental biology, could be promising targets for extending healthy lifespan. Here, we focus on anaplastic lymphoma kinase (Alk), an RTK with established roles in nervous system development and in multiple cancers, but whose effects on aging remain unclear. We find that several means of reducing Alk signalling, including mutation of its ligand jelly belly (jeb), RNAi knock-down of Alk, or expression of dominant-negative Alk in adult neurons, can extend healthy lifespan in female, but not male, Drosophila. Moreover, reduced Alk signalling preserves neuromuscular function with age, promotes resistance to starvation and xenobiotic stress, and improves night sleep consolidation. We find further that inhibition of Alk signalling in adult neurons modulates the expression of several insulin-like peptides, providing a potential mechanistic link between neuronal Alk signalling and organism-wide insulin-like signalling. Finally, we show that TAE-684, a small molecule inhibitor of Alk, can extend healthy lifespan in Drosophila, suggesting that the repurposing of Alk inhibitors may be a promising direction for strategies to promote healthy aging

A monocarboxylate transporter rescues frontotemporal dementia and Alzheimer's disease models.

Brains are highly metabolically active organs, consuming 20% of a person's energy at resting state. A decline in glucose metabolism is a common feature across a number of neurodegenerative diseases. Another common feature is the progressive accumulation of insoluble protein deposits, it's unclear if the two are linked. Glucose metabolism in the brain is highly coupled between neurons and glia, with glucose taken up by glia and metabolised to lactate, which is then shuttled via transporters to neurons, where it is converted back to pyruvate and fed into the TCA cycle for ATP production. Monocarboxylates are also involved in signalling, and play broad ranging roles in brain homeostasis and metabolic reprogramming. However, the role of monocarboxylates in dementia has not been tested. Here, we find that increasing pyruvate import in Drosophila neurons by over-expression of the transporter bumpel, leads to a rescue of lifespan and behavioural phenotypes in fly models of both frontotemporal dementia and Alzheimer's disease. The rescue is linked to a clearance of late stage autolysosomes, leading to degradation of toxic peptides associated with disease. We propose upregulation of pyruvate import into neurons as potentially a broad-scope therapeutic approach to increase neuronal autophagy, which could be beneficial for multiple dementias

Over-expression of bumpel reduces DPR levels.

(A) Survival curves of flies over-expressing bumpel (+RU) and their controls (-RU) during normal ageing. p = 0.06 by log rank (B) GR levels in fly heads over-expressing C9 alone or with bumpel after 7 days of induction, measured by ELISA. p = 0.0005 by unpaired t-test (C) GP levels in fly heads over-expressing C9 alone or with bumpel after 7 days of induction, measured by western blot. p = 0.0005 by unpaired t-test (D) mCD8GFP levels in fly heads over-expressing C9+mCD8GFP or with bumpel after 7 days of induction, measured by western blot. p = 0.1723 by unpaired t-test. (E) Survival curves of flies expressing 36(GR) repeats with (pink) and without (black) bumpel. p = 1.5E-44 by log rank test (F) Survival curves of flies expressing 36(PR) repeats with (pink) and without (black) bumpel. p = 2.1E-74 by log rank test (G) Survival curves of flies expressing 36(GA) repeats with (green) and without (black) bumpel. p = 3.2E-9 by log rank test. Genotypes: (A) elavGS/UAS-bumpel (B, C) UAS-36 (G4C2), elavGS, UAS-36 (G4C2), elavGS/UAS-bumpel (D) UAS-36 (G4C2)/UAS-mCD8GFP, elavGS, UAS-36 (G4C2)/UAS-mCD8GFP, elavGS/UAS-bumpel (E) UAS-36GR, elavGS, UAS-36 GR, elavGS/UAS-bumpel (F) UAS-36PR, elavGS, UAS-36PR, elavGS/UAS-bumpel. (G) UAS36GA, elavGS, UAS36GA, elavGS/UAS-bumpel.</p

Log fold change of TFEB targets genes in RNA seq of GR100 expressing flies vs control.

Log fold change of TFEB targets genes in RNA seq of GR100 expressing flies vs control.</p

Bumpel leads to pyruvate import.

(A) and (B) Lactate and pyruvate transport were measured using the lactate sensor Laconic (A) or the pyruvate sensor Pyronic (B). (A) Salivary glands from third instar larvae expressing Laconic and bumpel or its control were isolated and exposed to 1 mM lactate. Traces show the mean fluorescence ratio of mTFP/Venus obtained from 6 independent experiments (6 animals, total of 71 cells in control animals and 77 cells in animals over-expressing bumpel). Slope was calculated from the first 2 min of recording in the presence of lactate. p = 0.026 by Mann Whitney test. Area under the curve was calculated from the fluorescence obtained during the 5 min of lactate exposure. p = 0.004 by unpaired t-test. (B) Salivary glands expressing Pyronic and bumpel or its control were isolated and exposed to 1 mM pyruvate. Traces show the mean fluorescence ratio of mTFP/Venus obtained from 8 independent experiments (69 cells in control animals and 68 cells overexpressing bumpel). Slope were calculated from the first 2 min of recordings in the presence of pyruvate. p = 0.026 by Mann Whitney test. Area under the curve was calculated from the fluorescence obtained in 5 min of pyruvate exposure. p = 0.0346 by unpaired t-test. (C) Western blot of GP levels, relative to actin, in flies injected with 300 mM ethyl pyruvate and 2 M sodium pyruvate, or PBS as a control, and allowed to recover for 24 hours. Two different experimental replicates were combined in the statistical analysis (marked by different symbols), and then compared by t-test, *p = 0.0360 (D) GP levels in C9 and C9 bumpel fly heads with or without downregulation of Ldh by RNAi (E) GP levels in C9 and C9 bumpel fly heads with or without downregulation of Pdha by RNAi. GP levels in both D and E were compared by one way ANOVA followed by Šídák’s multiple comparisons test. All westerns samples shown for each panel were run on the same gel. Gentoypes: (A) UAS-36 (G4C2), actinGal4/UAS-Laconic (B) UAS-36 (G4C2), actinGal4/UAS-Pyronic (C) UAS-36 (G4C2), elavGS, (D) UAS-36 (G4C2), elavGS, UAS-36 (G4C2)/LdhRNAi, elavGS, UAS-36 (G4C2), elavGS/UAS-bumpel, UAS-36 (G4C2)/LdhRNAi, elavGS/UAS-bumpel, (E) UAS-36 (G4C2), elavGS, UAS-36 (G4C2)/Pdha RNAi, elavGS, UAS-36 (G4C2), elavGS/UAS-bumpel, UAS-36 (G4C2)/PdhaRNAi, elavGS/UAS-bumpel.</p