In slide at the top of page
Schematic representation of our working model. In budding yeast, intranuclear foci containing high concentrations of Hsf1 are formed in response to acute thermal stress. These represent phase-separated condensates that are linked to the dramatic 3D genome restructuring and transcriptional activity of HSP genes. My goal is to understand the role of Mediator, and in particular its tail subunit, Med15, in driving formation of these transcriptional condensates.
Gurrana Male, PhD
Education:
PhD, Biochemistry, University of Hyderabad, India, 2021
Gurranna Male's Research
The role of Mediator in heat shock-induced 3D genome restructuring in Saccharomyces cerevisiae
Chromatin architecture critically impacts nuclear processes that occur in a precise spatiotemporal manner. Perturbations in chromosomal topology can lead to human disease, including developmental deformities and cancer. The response of budding yeast to acute thermal stress (“heat shock”) is a dynamic example of transcriptional control and 3D genome organization. The evolutionarily conserved DNA-binding protein, Heat Shock Factor 1 (Hsf1), is a sequence-specific activator that orchestrates the heat shock transcriptional response of eukaryotes from yeast to humans. Previous studies in our lab using chromosome conformation capture (3C) and single-cell imaging showed that in response to heat shock, Hsf1-dependent Heat Shock Protein (HSP) genes engage in robust intergenic interactions across and between chromosomes and that these interactions involve both gene regulatory and coding regions. Concurrently, Hsf1 forms intranuclear foci that have properties of phase-separated intranuclear condensates. These foci, which contain the co-activator Mediator and RNA Pol II, appear to underlie the restructuring of the 3D genome that occurs during the acute phase of heat shock (Chowdhary et al, Mol. Cell 2022).
To investigate the role of Mediator in driving the formation of Hsf1-containing condensates and in restructuring the yeast 3D genome, I am focusing on the Tail subunit, Med15, that our previous work showed to be a physical target of Hsf1’s N- and C-terminal activation domains (Kim & Gross, JBC 2013). I am conducting a detailed dissection analysis of the activator binding domains (ABD’s) located in the N-terminal half of the protein. Specifically, I am determining the effect of individual med15 domain deletion mutations on HSP gene expression as well as the effect of such mutations on the recruitment of Hsf1 and RNA Pol II to these genes using precision ChIP assays. My long-term goal is to elucidate the role that Med15 and other subunits of Mediator play in the dramatic 3D repositioning of HSP genes during heat shock.
Med15 has an unusual structure and amino acid composition.
Cartoon depicts the major features of Med15:
- Amino acid coordinates of the four Activator Binding Domains (ABDs).
- The intrinsic disorder profile of Med15 corresponding to residue position.
- Location of each of the 20 amino acids within the protein. Shaded in pink are the Poly Q tracts of Med15.
- Amino acid composition of each ABD and location of the domain deletions that I am analyzing in my project.
