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Metabolic and Genetic Regulation of Ageing

Periodic Reporting for period 3 - MetAGEn (Metabolic and Genetic Regulation of Ageing)

Reporting period: 2018-08-01 to 2020-01-31

Ageing is a complex physiological process that affects almost all species, including humans. Despite its importance for all of us, the biology of ageing is insufficiently understood. To uncover the molecular underpinnings of ageing, I am performing an interdisciplinary research program that will identify and investigate metabolic and genetic regulators of ageing. Progressive loss of cellular homeostasis causes ageing and an age-associated decline in protein quality control has been implicated in numerous diseases, including neurodegeneration. This link between ageing and disease demonstrates the importance of basic research into the biology of ageing. On the one hand, this research offers new insights into fundamental biological processes. On the other hand, a deep understanding of biological underpinnings of the ageing process will open up new ways to address underlying causes for many diseases that affect Western societies.
Seeking for ways to improve protein quality, I have identified a novel longevity pathway in Caenorhabditis elegans. In a forward genetic screen, I found a link between metabolites in the hexosamine pathway and cellular protein quality control. Hexosamine pathway activation extends C. elegans lifespan, suggesting modulation of ageing by endogenous molecules.
In a first step, I am exploring the mechanism by which hexosamine metabolites improve protein quality control in mammals, using cultured mammalian cells and a mouse model for neurodegeneration. Preliminary data show that hexosamine pathway metabolites enhance proteolytic capacity in cells and reduce protein aggregation, suggesting conservation. Second, I am investigating molecular mechanisms that activate the hexosamine pathway to promote protein homeostasis and counter ageing. Third, I am performing a direct forward genetic screen for modulators of ageing in C. elegans. For the first time, mutagenesis and next generation sequencing can be paired in forward genetic screens to interrogate the whole genome for lifespan-extending mutations in a truly unbiased manner. This innovative approach has the potential to reveal novel modulators of the ageing process.
Taken together, this work aims to understand molecular mechanisms that maintain cellular homeostasis to slow the ageing process, and to develop a new technology to identify yet unknown genetic modulators of ageing.
I. Role of mammalian GFAT-1 in protein quality control. Using cell biological and biochemical assays, I am investigating if the HP can regulate protein homeostasis in mammals. Within the first reporting period we could show that indeed the toxic aggregation of human disease-associated proteins is reduced upon HP upregulation. This is dependent upon endogenous degradative processes. I will also use a mouse model of neurodegeneration to test if GFAT-1 activation might ameliorate an age-associated disease. We have generated the mouse model and are currently crossing triple-transgenic mice in order to investigate the role of neuron-specific GFAT-1 activation in a mouse model for a neurodegenerative disease.

II. Molecular characterization of GFAT-1. I will investigate the biochemical mechanism of GFAT-1 gof through enzyme kinetic assays and an unbiased search for interaction partners. We have now successfully purified recombinant human GFAT-1 and have established a number of activity assays. This work has revealed the mechanism for the GFAT-1 gain-of-function.

III. Unbiased direct forward genetic screen for longevity genes in C. elegans. I will perform the first forward genetic screen that uses longevity as a primary readout. We have now generated more than 100 long-lived C. elegans mutants and have completed next-generation sequencing for all of these within reporting period 1. Analysis of this rich and unique data set has revealed a number of candidates for novel longevity genes are we are in the process of confirming these using the CRISPR/Cas9 technology. Next, we will prioritize on the most interesting mutants and will devise a publication strategy.
I. Role of mammalian GFAT-1 in protein quality control. GFAT-1 is a key regulator of sugar metabolism and thus an interesting target for drug development with the purpose of ameliorating age-related disease. In C. elegans, its gain-of-function extends health- and lifespan. In Aim 1 (WP1), we are first asking if, as in the nematode, mammalian GFAT-1 might carry similar potential, particularly regarding neurodegenerative processes. Our cell-based data show that GFAT-1’s function is indeed conserved, not only in its role in controlling the HP, but also as a key regulator of protein homeostasis. This insight provides a novel modulator of processes relevant for healthy ageing.

II. Molecular characterization of GFAT-1. This work has identified the molecular mechanism of GFAT-1 gain-of-function. We are now using this knowledge to perform high throughput small molecule screens in a collaboration outside the MetAGEn project. This work will reveal potential candidate compounds that might improve protein quality control in vivo.

III. Unbiased direct forward genetic screen for longevity genes in C. elegans. Identification of novel longevity loci is critical to complete the picture of the genomic landscape of ageing. Previous technologies were unable to resolve genetic alterations at the level of individual amino acids or gain-of-function mutations in a systematic and unbiased way. Our collection of long-lived strains provides new knowledge about the role of various known longevity pathways, and also reveals new unexpected longevity genes. At this point, translational aspects of this work are unclear, but in the second reporting period, we will analyze a number of the most interesting mutants in greater detail and this will include research into their conservation to human biology.