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Tolwinski, N. S.

Publications and source records attributed to Tolwinski, N. S..

4 recordsLinked to original sources

Slowing ageing using drug synergy in C. elegans

Pharmacological interventions that target human ageing would extend individual healthspan and result in dramatic economic benefits to rapidly ageing societies worldwide. For such interventions to be contemplated they need to comprise drugs that are efficacious when given to adults and for which extensive human safety data are available. Here we show that dramatic lifespan extension can be achieved in C.elegans by targeting multiple, evolutionarily conserved ageing pathways using drugs that are already in human use. By targeting multiple synergistic ageing pathways, we are able to slow ageing rate, double lifespan and improves healthspan while minimize developmental and fitness trade-offs. Moreover, we established that there is no synergistic benefit in a daf-2 or daf-7 background, implying the involvement of the TGF{beta} and IGF pathways in this synergy. Employing lipidomics and transcriptomics analysis we found lipid metabolism to be affected resulting in increased monounsaturated fatty acids (MUFA) and decrease membrane peroxidation index. Our best drug combination showed a conserved lifespan extension in fruit flies. To the best of our knowledge, this is the largest lifespan effect ever reported for any adult-onset drug treatment in C. elegans. This drug-repurposing approach, using drugs already approved for humans to target multiple conserved aging pathways simultaneously, could lead to interventions that prevent age-related diseases and overall frailty in a rapidly ageing population.

biochemistry

An Embryonic System To Assess Wnt Transcriptional Targets

During animal development, complex signals determine and organize a vast number of tissues using a very small number of signal transduction pathways. These developmental signaling pathways determine cell fates through a coordinated transcriptional response that remains poorly understood. The Wnt pathway is involved in a variety of these cellular functions, and its signals are transmitted in part through a {beta}-catenin/TCF transcriptional complex. Here we report an in vivo Drosophila assay that can be used to distinguish between activation, de-repression and repression of transcriptional responses, separating upstream and downstream pathway activation and canonical/non-canonical Wnt signals in embryos. We find specific sets of genes downstream of both {beta}-catenin and TCF with an additional group of genes regulated by Wnt, while the non-canonical Wnt4 regulates a separate cohort of genes. We correlate transcriptional changes with phenotypic outcomes of cell differentiation and embryo size, showing our model can be used to characterize developmental signaling compartmentalization in vivo.

developmental biology

Coupling optogenetics and light-sheet microscopy to study signal transduction in vivo

Optogenetics allows precise, fast and reversible intervention in biological processes. Light-sheet microscopy allows observation of the full course of embryonic development from egg to larva. Bringing the two approaches together allows unparalleled precision into the temporal regulation of signaling pathways and cellular processes in vivo. To develop this method, we investigated the regulation of canonical Wnt signaling during anterior-posterior patterning of the Drosophila embryonic epidermis. Cryptochrome 2 (CRY2) from Arabidopsis Thaliana was fused to mCherry fluorescent protein and Drosophila {beta}-catenin to form an easy to visualize optogenetic switch. Blue light illumination caused oligomerization of the fusion protein and inhibited downstream Wnt signaling in vitro and in vivo. Temporal inactivation of {beta}-catenin confirmed that Wnt signaling is required not only for Drosophila pattern formation, but also for maintenance later in development. We anticipate that this method will be easily extendable to other developmental signaling pathways and many other experimental systems.

developmental biology

Decoding temporal interpretation of the morphogen Bicoid in the early Drosophila embryo

Morphogen gradients provide essential spatial information during development. Not only the local concentration but also duration of morphogen exposure is critical for correct cell fate decisions. Yet, how and when cells temporally integrate signals from a morphogen remains unclear. Here, we use optogenetic manipulation to switch off Bicoid-dependent transcription in the early Drosophila embryo with high temporal resolution, allowing time-specific and reversible manipulation of morphogen signalling. We find that Bicoid transcriptional activity is dispensable for embryonic viability in the first hour after fertilization, but persistently required throughout the rest of the blastoderm stage. Short interruptions of Bicoid activity alter the most anterior cell fate decisions, while prolonged inactivation expands patterning defects from anterior to posterior. Such anterior susceptibility correlates with high reliance of anterior gap gene expression on Bicoid. Therefore, cell fates exposed to higher Bicoid concentration require input for longer duration, demonstrating a previously unknown aspect of morphogen decoding.

developmental biology