The Glucocorticoid Receptor in Aging and Circadian Endocrinology

Project Information

GRACE

Grant agreement ID: 101086997

DOI

10.3030/101086997

EC signature date 2 March 2023

Start date 1 June 2023

End date 31 May 2028

Funded under

• European Research Council (ERC)

Total cost

€ 1 997 493,75

EU contribution

€ 1 997 493,75

Coordinated by

TECHNISCHE UNIVERSITAET MUENCHEN
Germany

Periodic Reporting for period 1 - GRACE (The Glucocorticoid Receptor in Aging and Circadian Endocrinology)

Reporting period: 2023-06-01 to 2025-11-30

Summary of the context and overall objectives of the project

How Calorie Restriction and Body Rhythms Work Together to Support Health and Longevity
We all know that it’s important how much we eat, but a growing body of research suggests that when we eat may be just as critical for our health. Scientists studying calorie restriction (eating fewer calories without malnutrition) have found that its benefits are most powerful when meals are consumed in sync with the body’s natural daily rhythms.
These rhythms, known as circadian clocks, govern everything from sleep to behavior and hormone levels. One key hormone, cortisol, which regulates stress and metabolism, follows a strong daily cycle. In mice, researchers found that the benefits of calorie restriction were greatest when food was given during the time of day when this hormone naturally peaks.
Digging deeper, my lab and I discovered that under calorie restriction, the body amplifies the daily release of cortisol. Cortisol binds a special protein in liver cells, the glucocorticoid receptor (GR), which switches certain genes on and off. Together with its receptor, the cortisol hormone triggers healthy changes in metabolism and supports cellular repair processes like autophagy, which helps clean up damaged proteins and organelles in cells.
To test the importance of this system, we used genetically modified mice that lacked the GR protein in the liver. These mice didn’t experience the usual health benefits of calorie restriction, showing how essential this hormone-receptor interaction is. We also found that under calorie restriction, a protein called FOXO1 becomes more active, helping to promote healthy gene activity. This suggests that timing and rhythmic patterns are essential: it’s not just about which hormones are present, but when they are active.
Our research explores an exciting new frontier in health science: the interaction between diet, daily biological rhythms, and hormone signaling. It suggests that to get the full benefits of a healthy diet, especially one that reduces calories, we may need to pay closer attention to when we eat, not just to what we eat. In the future, these findings could help shape new dietary guidelines or treatments aimed at improving metabolic health, slowing aging, and preventing diseases, for example by aligning eating patterns with our body’s natural clock

Work performed from the beginning of the project to the end of the period covered by the report and main results achieved so far

Activities Performed and Main Achievements (Technical and Scientific)
The project investigates how glucocorticoid signaling integrates with caloric restriction (CR) and circadian rhythms to regulate liver metabolism at the molecular level. Activities are structured across three primary aims:
Aim 1: Dissecting Glucocorticoid Signaling Under Caloric Restriction
We used liver-specific glucocorticoid receptor (GR) knockout mice and multi-omics approaches (ChIP-seq, RNA-seq) to determine the role of GR in CR-induced gene regulation.
Key finding: GR is essential for activating a de novo, CR-specific rhythmic transcriptional program involving autophagy and metabolic genes.
Artificial corticosterone administration without CR did not replicate this response, instead triggering inflammation and metabolic imbalance.
Motif analysis of GR binding sites revealed diet-dependent co-regulator recruitment (e.g. FOXO1 under CR, STAT5 under corticosterone-only conditions).
Aim 2: Investigating the Role of Rhythmic Hormone Input
A ligand-independent GR mutant (GRΔLBD) was engineered to assess whether rhythmic glucocorticoid input is necessary for GR function.
This modified receptor was delivered in vivo via liver-specific AAV and validated for constitutive nuclear activity.
Complementary studies used corticosterone-releasing pellets to disrupt endogenous GC rhythms.
Preliminary data suggest that constant activation of GR impairs time-specific gene expression and alters metabolic regulation, highlighting the necessity of rhythmic GC signaling for optimal gene control.
Aim 3: Mapping GR-Dependent Enhanceosome Complexes
We used ChIP-MS to identify co-regulators recruited to GR-containing complexes under different physiological conditions.
Genetically modified mice expressing epitope-tagged p300 and CBP were created for high-specificity interaction mapping.
Protocol optimization and antibody validation (via CRISPR knockouts) improved the fidelity of ChIP-MS analyses.
Diurnal variation in cofactor binding (e.g. day vs. night differences in p300/CBP occupancy) was confirmed, supporting the idea of time-encoded enhancer complex dynamics.
Outcomes of the Action
Demonstrated that the beneficial effects of CR depend not only on calorie intake but also on timing relative to glucocorticoid peaks.
Identified GR as a key mediator of CR-induced metabolic gene programs, functioning through co-regulator-specific enhancer complexes.
Showed that rhythmic hormone input is critical for effective GR activity; constant or misaligned signaling disrupts metabolic homeostasis.
Developed validated tools and protocols for ligand-independent GR activation, time-resolved ChIP-MS, and liver-specific transcriptional analysis.
These insights provide a mechanistic foundation for timing-based metabolic therapies, aligning dietary or hormonal interventions with circadian biology to optimize efficacy and reduce side effects

Progress beyond the state of the art and expected potential impact (including the socio-economic impact and the wider societal implications of the project so far)

Several of our findings go significantly beyond the current state of knowledge. Most importantly, we discovered a new transcriptional switch in liver cells involving STAT5 and FOXO1. This switch allows the glucocorticoid receptor (GR) to activate a specific set of rhythmic genes—but only under caloric restriction (CR). This finding shows how signals from the body’s internal clock, diet, and hormones work together to control metabolism. It also shows that the timing and context of GR activation are just as important as hormone levels. Simply raising corticosterone levels could not reproduce the same gene expression seen under CR, highlighting the unique effect of feeding schedule and physiological rhythm.

Another major advance was the creation of a GR mutant (GRΔLBD) that works without needing hormone input. This allowed us to test whether daily hormone rhythms are required for GR to regulate its target genes and co-regulators—an open question in the field.

Lastly, we optimized ChIP-MS to map transcriptional co-regulators in live animals. Using CRISPR-tagged mouse models and knockout controls, we created a reliable method for studying how gene regulatory complexes change with diet and time of day. These combined findings offer both conceptual insights and new tools for studying metabolism, circadian biology, and aging.