No abstract

Plasmodium falciparum requires a two-host system, moving between Anopheles mosquito and humans, to complete its life cycle. To overcome such dynamic growth conditions its histones undergo various post-translational modifications to up-regulate and down-regulate certain genes required for invasion and replication. The P. falciparum genome encodes six bromodomaincontaining proteins, of which Bromodomain Protein 1 (PfBDP1) has been shown to interact with acetylated lysine modifications on histone H3 to regulate the expression of invasion-related genes. Here, we investigated the ability of the PfBDP1 bromodomain to interact with acetyllsyine modifications on additional core and variant histones. A crystal structure of the PfBDP1 bromodomain (PfBDP1-BRD) reveals it contains the conserved bromodomain fold, but our comparative analysis between the PfBDP1-BRD and the 8 human bromodomain families indicates it has a unique binding mechanism. Solution NMR spectroscopy and ITC binding assays carried out with acetylated histone ligands demonstrate that it preferentially recognizes tetra-acetylated histone H4, and we detected weaker interactions with multi-acetylated H2A.Z in addition to the previously reported interactions with acetylated histone H3. Our findings indicate PfBDP1 may play additional roles in the P. falciparum life cycle, and the distinctive features of its bromodomain binding pocket could be leveraged for the development of new therapeutic agents to help overcome the continuously evolving resistance of P. falciparum against currently available drugs.

Bromodomains are evolutionarily conserved protein modules that specifically recognize acetylated lysine residues on histone tails of the nucleosome. During the red‐blood‐cell‐stage of infection with Plasmodium falciparum, the parasite undergoes repeated rounds of replication, egress, and invasion. Erythrocyte invasion involves specific interactions between host cell receptors and parasite ligands and coordinated expression of genes specific to this step of the life cycle. A parasite‐specific protein known as bromodomain protein 1 (PfBDP1) binds to chromatin at the transcriptional start sites of invasion‐related genes and directly controls their expression. Conditional PfBDP1 knockdown causes a dramatic defect in parasite invasion and growth, and results in transcriptional down‐regulation of multiple invasion‐related genes at a time point critical for invasion. Conversely, PfBDP1 overexpression enhances expression of these same invasion‐related genes. PfBDP1 has been shown to interact with acetylated histone H3 and a second bromodomain protein, PfBDP2, suggesting a potential mechanism for gene recognition and control. Since PfBDP1 critically coordinates expression of invasion genes, targeting PfBDP1 could be an invaluable tool in mitigating malaria infections. The domain architecture of PfBDP1 includes a single C‐terminal bromodomain and several ankyrin repeats. PfBDP1 is thought to act by tethering a transcriptional activator complex to acetylated histone H3 to control genes required for parasite invasion. We hypothesized that the bromodomain is crucial for this function of PfBDP1 via recognition of acetylated lysine modifications. We carried out several histone binding assays to identify novel histone interactions the of PfBDP1 bromodomain. We also utilized a combination of structural biology and biophysical approaches to identify amino acid residues critical for ligand coordination. The result of this study will provide molecular details about the recognition of acetylated histone ligands by the PfBDP1 bromodomain and shed light on how this domain recruits the PfBDP1‐PfBDP2 complex to chromatin. Importantly, these results may improve the development of new therapeutics to treat malaria

Epigenetic regulatory mechanisms are important for the growth and survival of eukaryotic organisms. Post‐translational modifications found on histone proteins in the nucleosome function as a dynamic cell signaling mechanism that controls many essential functions such as cell growth, DNA replication, and gene expression. Bromodomains are conserved protein interaction modules that specifically recognize acetylated lysine residues on histones. Among five Plasmodium species, P. falciparum is the major cause of malaria cases worldwide. According to a WHO report in 2019 there were 409,000 deaths reported worldwide and the most affected age group was of children under 5 years of age. Interestingly, the P. falciparum genome encodes eight bromodomain‐containing proteins, and bromodomain protein 1 (PfBDP1) has been shown to play an important role in malaria pathogenesis. The domain architecture of PfBDP1 includes a single C‐terminal bromodomain and several ankyrin repeats. PfBDP1 is thought to act by tethering a transcriptional activator complex to acetylated histone H3 to control genes required for parasite invasion. Additionally, PfBDP1 forms a complex with the bromodomain protein 2 (PfBDP2) to carry out its cellular functions. We hypothesized that the bromodomain is crucial for the function of PfBDP1 via recognition of acetylated lysine modifications. We carried out several histone binding assays to identify novel histone interactions of PfBDP1 bromodomain. We also utilized a combination of structural biology and biophysical approaches to identify amino acid residues critical for ligand coordination. The rise of drug‐resistance towards P. falciparum has created an urgent need to outline the molecular mechanisms driving pathogenesis in hopes of uncovering new potential drug targets. Importantly, the results from this study may improve the development of new therapeutics to treat malaria.