The Denzel lab is interested in understanding the process of ageing through basic research. Ageing leads to a decline in general homeostasis and is characterized by a loss of stress resistance and a dysregulation of protein quality control.

We address this problem though two interconnected approaches: genetics and metabolism.

Genetics: we perform forward genetic mutagenesis screens that interrogate the genome at amino acid resolution. By doing this, we identify novel modulators of stress resistance and aging.

Metabolism: To maintain homeostasis, organisms constantly turn over the basic building blocks of important biomolecules. The relevant pathways need to be finely tuned and we investigate how these pathways affect protein homeostasis and ageing.

Understanding the metabolic and genetic regulation of ageing requires multiple parallel approaches.

We investigate the metabolic modulation of protein homeostasis, mechanisms that control protein synthesis, and the role of stress resistance in longevity. As these fundamental biological processes are well conserved, we use the nematode C. elegans as a discovery tool and to investigate systemic effects. Further, we use stem cells for genetic screens to investigate the role of metabolism in stem cell fate decisions. Finally, we test findings from basic approaches in mice and in clinical studies with the University Hospital Cologne. 

Regulation of protein quality control by the hexosamine pathway.

The hexosamine pathway (HP) produces UDP-N-acetylglucosamine (UDP-GlcNAc), a precursor for glycosylation reactions. Gain-of-function mutations in the HP’s key enzyme glutamine-fructose 6-phosphate aminotransferase (GFAT-1) elevate cellular UDP-GlcNAc levels and improve protein quality control in C. elegans, resulting in lifespan extension (Denzel et al., 2014). The activating GFAT-1 mutations are single amino acid substitutions at distinct positions. This provides an opportunity to elucidate structure-function relationships in this essential enzyme. GFAT-1 is highly conserved and thus we focus on the human enzyme to investigate the gain-of-function mechanism through activity assays and crystallography in collaboration with Prof. U. Baumann (University of Cologne). This work has revealed detailed insights into the allosteric mechanism regulating GFAT-1 activity. Together with the Lead Discovery Centre in Dortmund, we are leveraging this knowledge to perform high throughput screens for GFAT-1 activators.

We further work on functional aspects of the HP in mammalian protein homeostasis. Finally, we perform new forward genetic screens to identify novel regulators of the HP.

This work is funded by the Federal Ministry of Education and Research (BMBF) and by the European Research Council (ERC Starting Grant).

Forward mutagenesis screen identifies novel mechanisms of lifespan extension in C. elegans.

Longevity is a difficult phenotype to follow in large-scale genetic studies and thus whole genome chemical mutagenesis screens for longevity were not performed in a large manner in the past. RNA interference was used instead despite its limitations to knockdown and non-essential genes. To close this gap, we have performed an unbiased screen for longevity using mutagenesis in C. elegans. We have generated 100 independent long-lived strains and performed whole genome sequencing on all of them. Naturally, we identify new alleles of known longevity genes, but we do also identify novel and unexpected longevity genes. We use CRISPR/Cas9 to confirm novel longevity loci.

Although the screen was done in a fully unbiased fashion, we find many genes that control protein homeostasis. This demonstrates its key relevance in extending lifespan. We are currently developing in-depth analyses on the new long-lived mutants.

This work is funded by the European Research Council (ERC Starting Grant).

Use of haploid stem cells for mutagenesis screens

In diploid cells a recessive mutation will hardly result in a phenotype as long as the wild type copy of the gene is present. Thus, genetic screens are particularly effective in organisms that can be maintained in the haploid state, or which generate homozygous mutant offspring. Only in this way, recessive mutations lead to an observable phenotype. To perform recessive genetics with chemical mutagens we have established a screening platform for resistance screens in haploid mouse embryonic stem cells. These enable detailed structure-function analyses and map interactions at amino acid resolution (Horn et al., 2018 and 2019). To investigate modulators of protein homeostasis, we have performed a saturation screen for the proteasome inhibitor (PI) bortezomib and have identified resistance mutation in its target PSMB5. This approach predicts resistance mutations that occur in treated multiple myeloma patients. This work opens the opportunity for predicting chemoresistance in patients and, more importantly, will help to stratify patients to enable individualized treatments, which is currently impossible in multiple myeloma. A prospective clinical study to test this hypothesis is under preparation.

A further application of this work is in target deconvolution of drug candidate compounds. Based on this potential, we have built a start-up company (Acus Laboratories GmbH) to further develop and utilize this platform.

This work was funded by the European Research Council (ERC Proof-of-Concept Grant).

Metabolic regulation of epidermal stem cell fate

Protein synthesis plays a key role in stem cell fate decisions. Low protein synthesis is associated with the stem cell state while differentiating cells induce protein synthesis. A potential causal role of protein synthesis in stem cells is an emerging research field. To understand if this is a regulatory pathway that cells employ, we are investigating the role of upstream metabolic regulators of protein synthesis in hair follicular stem cells.

This work is funded by the German Research Foundation SFB829.

Perspectives

This work has implications for both basic biology and translational research.

Basic biology: Ageing is a phenomenon of (almost) all living organisms, but what determines the rate of ageing remains a mystery. Through our interdisciplinary approaches, we identify new regulators of the ageing process. These insights provide entry points for research on basic biological mechanisms.

Translational research: While general principles of the regulation of ageing are being discovered, there is still a shortage of opportunities to translate this knowledge towards human health. We therefore aim to leverage our insights into the biology of ageing to identify mechanisms extending health- and lifespan.