Glass Lab Research

OVERVIEW

Even though the DNA in all of our cells is nearly the same, it is epigenetic factors, including modifications to DNA, RNA, and chromatin that control the gene expression patterns in different cells, under various environmental conditions, and at specific times (1).  Post-translational modifications on histone tails function as a very complex code for controlling gene expression.  Deciphering this code is at least as important as the genetic code for understanding its role in stem cell programming, maternal effects during fetal development, and potential reversibility in cancer therapies.  However, it is also orders of magnitude more complex (2).  We study the molecular mechanisms driving the recognition of histone modifications, and how combinations of these chemical signals regulate protein interactions on the nucleus in both normal and disease states.

 Major research efforts in the lab include:

  • To uncover how multiple chromatin reader domains cooperatively regulate the activity of epigenetic regulatory complexes.
  • To reveal how cross-talk between epigenetic marks modulates the function of histone binding proteins and chromatin remodeling enzymes.
  • To identify and functionally characterize how acetyllysine signaling and recognition contributes to the pathogenesis of parasitic infections.
  • To correlate how altered epigenetic modifications influence the progression of cancer, heart, and infectious disease.

BACKGROUND

The human genome is compacted into chromatin, allowing nearly three meters of DNA to fit into the small volume of the nucleus.  Chromatin is composed of DNA and proteins, and this DNA-protein complex is the template for a number of essential cell processes including transcription and replication.  Understanding the role of chromatin's higher order structure in transcriptional control is important as loss of this regulation underlies many disease processes.

The basic structural unit of chromatin is the nucleosome.  Nucleosomes are comprised of 147 base pairs of DNA wrapped around a core histone octamer.  The histone octamer contains two molecules of each histone:  H2A, H2B, H3, and H4.  Each of these core histones contains two separate functional domains - a globular domain, which interacts with the DNA and other histones and a flexible tail domain that protrudes from the nucleosome.  The tail domains can be modified by the reversible addition of chemical groups such as acetyl-, methyl-, and phospho- groups.

Modifications of the histone tail have been shown to be important in altering chromatin structure, facilitating access for DNA-binding transcription factors, but they also act as markers allowing non-histone proteins to interact with the chromatin.  The "Histone Code Hypothesis" suggests that histone tail modifications constitute an epigenetic (beyond genes) code, which is read by other proteins.  It postulates that these proteins and protein complexes are able to recognize/read distinct tail modifications, just like a language or code.  This consequently triggers downstream events resulting in unique and specific biological outcomes such as cell death; cell cycle regulation; and the transcription, repair, or replication of DNA.

RECENT WORK

Epingenetic signaling by histone post-translational modifications

We are investigating the structure and function of chromatin binding domains, including the bromodomain, which interact specifically with acetylated histones.  There are about 60 human bromodomain-containing proteins, and these nuclear proteins have a wide variety of biological activities (3).  Bromodomains bind to specific acetylation marks on histone tail and tether associated proteins and enzymatic complexes to histones to regulate chromatin structure and gene expression (4).  However, how these protein modulates differentiate between multiple actylated lysine residues, that are often found alongside other post-translation modifications, to read the histone code is unkown. We recently esablished the molelcular basis of histone acetyllysine recognition by the ATAD2 and ATAD2B bromodomains and discovered that these bromodomains prefer to interact with histones H4, H2A, and the H2A.X histone variant when they contain multiple acetylated lysine residues.  The structural and mechanistic details of histone recognition by bromodomains is crucial for the development of new therapeutic interventions and molecular tools to study a variety of diseases.  Our research has fundamentally advanced our understanding of how bromodomains recognize and select for acetyllysine marks.

 

Figure 1 - Multiple modifications on individual histone tails...

 

Bromodomain-containing proteins in cardiovascular and infectious diseases

Plasmodium falciparum is a unicellular protozoan parasite that causes malaria infections in humans.  Malaria is a significant global health problem that affected 247 million people in 2021, resulting in approximately 619,000 deaths (5).  Unfortunately, this disease disproportionally affects infants and children, and the cases have been expanding since 2016 due to climate changes and drug resistance (5).  In humans, P. falciparum first replicates in the liver cells, and as the disease progresses, it moves into the red blood cells (RBCs) (6).  The symptoms of malaria are associated with repeated rounds of parasite infection, and invasion into the red blood cells (7).  At the red blood cell stage of infection, P. Falciparum consumes the RBCs hemoglobin, preventing it from carrying oxygen to the heart which results in anemic heart failure (8).  In addition, parasitized RBCs stick to the wall of blood vessels in the heart and brain to evade the immune system, which often leads to inflammation and causes blood vessel blockage in these vital organs.  These infection-related complications are directly associated with the invasion of RBCs by the parasite.  The P. falciparum Bromodomain Protein 1 (PfBDP1) is a multi-domain nuclear protein that contains a unique combination of ankyrin repeats (ANK) followed by a bromodomain (BRD) (Fig. 2A).  In-vivo studies have demonstrated that association of PfBDP1 with acetylated chromatin at the promoters of invasion genes promotes their expression, and knockdown of PfBPD1 strongly reduces expression, and blocks the invasion of RBCs (Fig. 2B) (9,10).  The chromatin binding activity of PfBDP1 is thought to play an essential role in the red blood cell invasion process by controlling the expression of genes that are required for cell entry.  Thus, it is imperative to understand the essential factors involved in the P. falciparum RBC invasion process to develop therapeutic interventions.

Figure 2 - Role of PfBDP1 in regulating the expression of invasion related genes

The focus of my research is aimed at determining the structures of chromatin binding domains, including the bromodomain and PHD finger, in complex with the histone tail to elucidate how histone tail modifications are recognized.  This research will aid in the deeper understanding of how chromatin remodeling complexes are targeted to the chromatin and regulate gene expression.  A greater understanding of how these molecular signaling pathways function and are regulated will provide insights into how they can be therapeutically manipulated, and may help identify new diagnostic markers and targets to prevent and treat disease.

 

 

Dr. Karen Glass

E-mail: Karen.Glass@med.uvm.edu

Phone: (802)656-5760
Office Location: Firestone 362

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REFERENCES:

1. Cloud J. (2010).  Why Your DNA Isn't Your Destiny.  Time Magazine 172. (2010)

2. Aebersold R, et al.  How many human proteoforms are there?  Nat Chem Biol 14, 206-214. (2018)

3. Filippakopoulos P, et al.  Histone recognition and large-scale structural analysis of the human bromodomain family.  Cell 149, 214-231 (2012)

4. Lubula MY, et al.  Structural insighes into recognition of acetylated histone ligands by the BRPF1 bromodomain.  FEBS Lett (2014)

5. WHO, G.M.P. World malaria report 2022. (ed. Organization, W.H.) 293 (https://www.who.int/publications/i/item/9789240064898, 2022) 

6. Vaughan, AM & Kappe, S.H.I. Malaria Parasite Liver Infection and Exoerythrocytic Biology.  Cold Spring Harb Perspect Med 7 (2017)

7. Mohandas, N & An, X. Malaria and human red blood cells.  Med Microbiol Immunol 201, 593-8 (2012)

8. Weatherall, DJ, et al. Malaria and the red cell.  Hematology Am Soc Hematol Educ Program, 35-57 (2002)

9. Josling GA, et al.  A Plasmodium Falciparum Bromodomain Protein Regulates Invasion Gene Expression.  Cell Host Microbe 17, 741-51 (2015)

10. Quinn, JE, et al.  The Putative Bromodomain Protein PfBDP7 of the Human Malaria Parasite Plasmodium Falciparum Cooperates with PfBDP1 in the Silencing of Variant Surface Antigen Expression.  Front Cell Dev Biol 10, 816558 (2022)