Helmholtz Zentrum München Deutsches Forschungszentrum für Gesundheit und Umwelt

Project description

Glucocorticoids are widely prescribed anti-inflammatory drugs in the treatment of COVID-19,
asthma, sepsis and others. However, chronic and systemic treatment comes with severe adverse
effects like diabetes type 2, obesity, osteoporosis, muscle atrophy etc. Molecularly,
glucocorticoids diffuse across the cellular membrane and engage with the glucocorticoid
receptor (GR), which translocates into the nucleus. In the nucleus, the GR binds glucocorticoid
response elements and represses as well as activates target gene expression. Interestingly, gene
activation is a hallmark of glucocorticoid-mediated side effects in metabolic tissues, whereas
gene repression is associated with its anti-inflammatory function. Therefore, the Uhlenhaut Lab
is exploring the molecular circuits employed by the GR to understand gene repression versus
activation; a biological conundrum not only restricted to nuclear hormone receptors. Our aim is
to identify new therapeutically targets to improve glucocorticoid therapy and reduce adverse

In recent years, we have studied the molecular mechanisms of gene activation versus repression
in macrophages using next-generation sequencing technologies and were able to describe
factors required for gene activation in a gene-specific manner. We also discovered that the
chromatin states at glucocorticoid binding sites associated with gene repression are highly
diverse, pointing towards gene-specific repression mechanisms employed by the glucocorticoid
receptor. Those observations are in line with the diverse mechanisms described in the literature.

In this project, we will explore the molecular mechanisms employed by GR in the regulation of
inflammatory gene expression. We will use the bacteriophage-derived RNA stem-loops as
transcriptional reporters co-expressed with fluorescently labelled coat proteins (MS2/MCP and
PP7/PCP) in combination with CRISPR/Cas9-mediated gene targeting to monitor transcription
dynamics over time in single cells. In cooperation with the Grima Lab, we will develop and fit
mathematical models of stochastic transcription to live cell-imaging data to quantitatively
understand how glucocorticoids influence transcriptional bursting (frequency and burst size) at
genes with different activation and repression dynamics. In combination with a co-regulator
CRISPR screen, we will identify factors that contribute to inflammatory gene repression by
glucocorticoids globally or gene-specifically. At the end of the project, we will understand how
glucocorticoids affect gene expression of different pro-inflammatory genes and which co-factors
are involved.

Relevant literature

- Greulich F, Bielefeld KA, Scheundel R, Mechtidou A, Strickland B, Uhlenhaut NH, 2021, Research
article, Enhancer RNA Expression in Response to Glucocorticoid Treatment in Murine
Macrophages., Cells;11(1):28

- Greulich F, Wierer M, Mechtidou A, Gonzalez-Garcia O, Uhlenhaut NH, 2021, Research article,
The glucocorticoid receptor recruits the COMPASS complex to regulate inflammatory
transcription at macrophage enhancers., Cell Rep.;34(6):108742

- Strickland BA, Ansari SA, Dantoft W, Uhlenhaut NH., 2022, Review, How to tame your genes:
mechanisms of inflammatory gene repression by glucocorticoids. FEBS Lett. Epub ahead of print

- Cao Z, Grima R., 2020, Analytical distributions for detailed models of stochastic gene expression
in eukaryotic cells. Proceedings of the National Academy of Sciences 117.9: 4682-4692.

- Jiang Q, Fu X, Yan S, Li R, Du W, Cao Z, Qian F and Grima R, 2021. Neural network aided
approximation and parameter inference of non-Markovian models of gene expression. Nature
communications, 12(1), pp.1-12.