Mechanism of heterochromatin regulation by STAT Heterochromatin is a tightly packed form of DNA important for chromosomal compaction and transcriptional silencing as well as for genome stability, animal longevity, and tumor suppression. How heterochromatin dynamics, i.e., its establishment, maintenance, and loss, is controlled remains incompletely understood. We have previously demonstrated a physiological role of uSTAT in heterochromatin maintenance, but the mechanism remains unclear. The overall goal of this project is to investigate, at the molecular level, the role of STAT in establishing and maintaining heterochromatin, and to understand how STAT phosphorylation can serve as a molecular switch converting gene silencing to active transcription. We have previously shown by immunostaining that a fraction of STAT not phosphorylated at the critical tyrosine around amino acid 700 (termed uSTAT) is localized in the nucleus in association with Heterochromatin Protein 1 (HP1). We have shown genetically that STAT is essential for heterochromatin maintenance, and that STAT activation (by phosphorylation on this tyrosine) is associated with heterochromatin disruption. We have further shown that human uSTAT5A is capable of promoting heterochromatin formation and suppressing tumor growth. In new preliminary studies using chromatin immunoprecipitation followed by deep sequencing (ChIP-seq), we have found that the majority of chromatin-bound Drosophila STAT is localized in heterochromatin, and that loss of STAT leads to a global decrease in heterochromatin, which is marked by trimethylated histone 3 at lys9 (H3K9me3). We have further found that, when forced to bind to euchromatin, uSTAT can repress nearby genes in an HP1-dependent manner. These results suggest that uSTAT may play an important role not only in maintaining but also in initiating heterochromatin formation. HP1 and H3K9me3 are hallmarks of heterochromatin, with HP1 being the central component. It has been shown that recruiting HP1 to euchromatin is sufficient for heterochromatin formation. However, HP1 does not bind DNA directly and has only weak affinity for H3K9me3, and thus may require DNA-binding protein factors for its initial recruitment to heterochromatic loci, or for strengthening its binding to H3K9me3. We hypothesize that uSTAT plays roles in both establishment and maintenance of heterochromatin due to the molecular confirmation of uSTAT dimers. We will use a combination of genomic, genetic, and biochemical tools to investigate the initial events leading to heterochromatin establishment, the respective roles of STAT and HP1 in heterochromatin maintenance, the requirement for cis-element in uSTAT binding and HP1 recruitment, and the properties of uSTAT and pSTAT dimers in DNA and HP1 binding. Understanding the molecular mechanism of noncanonical STAT function in heterochromatin regulation should shed light on not only the basic cell biological process of heterochromatin dynamics but also heterochromatic gene silencing relevant to tumor suppression and epigenetic human diseases.

Public Health Relevance

Heterochromatin is important for multiple cellular processes including chromosomal compaction, genome integrity, transcriptional gene silencing, and tumor suppression. While our previous genetic research has shown a requirement for the JAK/STAT pathway in maintaining heterochromatin, the molecular mechanisms remain unclear. The overall goal of this project is to elucidate the molecular mechanism by which unphosphorylated STAT plays an essential role in heterochromatin establishment and maintenance.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM131044-03
Application #
10003343
Study Section
Nuclear and Cytoplasmic Structure/Function and Dynamics Study Section (NCSD)
Program Officer
Carter, Anthony D
Project Start
2018-09-20
Project End
2022-08-31
Budget Start
2020-09-01
Budget End
2021-08-31
Support Year
3
Fiscal Year
2020
Total Cost
Indirect Cost
Name
University of California, San Diego
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
804355790
City
La Jolla
State
CA
Country
United States
Zip Code
92093