Genome-wide analysis of regulatory G-quadruplexes affecting gene e-pression in human cytomegalovirus
- Ravichandran S, Kim YE, Bansal V, Ghosh A, Hur J, Subramani VK, Pradhan S, Lee MK, Kim KK, Ahn JH.
- PLoS Pathog. 2018 Sep 28;14(9):e1007334. doi: 10.1371/journal.ppat.1007334.
The discovery of the double-helix structure of DNA in 1953 heralded a new era of molecular biology. It was later also discovered that DNA can adopt a myriad of alternative conformations such as Z-DNA, cruciform, H-DNA, and G-quadruplexes. It is interesting that while DNA is expected to be double stranded, why does it adopt these conformations? Many researchers have been trying to unravel the mysteries behind these alternate DNA conformations.
Recently, a lot of focus has been placed on G-quadruplexes (GQs), which are formed from repetitive guanine tracts. They are shown to be more stable than double-stranded DNA (dsDNA) and play major roles in cancer and other neurodegenerative diseases such as FTD (Frontotemporal Dementia) and ALS (Amyotrophic Lateral Sclerosis). While the implications of these structures have been elucidated in humans, the impact on prokaryotes such as bacteria and viruses are only recently being explored.
The recent outbreaks of Ebola, SARS, and Human Papilloma Virus (HPV) have resulted in an urgent search for new methods to combat viruses. Therefore, we hypothesized that since the GQ motifs show regulatory activities in both humans and bacteria, they must play critical roles in viruses too. We chose to study the genome wide effect of GQ formation in the regulatory regions of Human Cytomegalovirus (HCMV) because of its ability to be life - threatening in immunocompromised individuals and babies.
Bioinformatics analysis indicated that GQ motifs were enriched in the HCMV genome. We chose to study the effects of GG structures on gene regulation, especially in those genes linked to virulence. Unlike previous studies, our study aimed to provide a holistic view of gene regulation by GQ structures. We found that stable GQ structures could form within the promoter regions of HCMV. In vitro reporter assays showed that the formation of GQ structures could affect gene expression in a context-dependent manner, where additional factors such as proteins that bind to GQ structures could mediate gene regulation. We also confirmed gene regulation in vivo by creating a mutant virus that showed increased gene expression when the GQ structure was abolished. We also found that GQ-stabilizing ligand NMM-lX can attenuate gene expression and affect virus replication at sub-cytotoxic concentrations.
Our study provides a platform to show how GQ mediated gene regulation could be effective in suppressing the expression of virulence genes. Additionally, thereby providing a new avenue for GQ-binding ligands to be used as antiviral therapy. In our future studies, we aim to extend our research to other viruses and evaluate other G4-binding ligands for antiviral activities. We are also attempting to explore the underlying molecular mechanisms that control the dynamic stabilization and destabilization of G-quadruplexes and other secondary structures in viral genomes.
The Structural Biology Lab headed by Prof. Kyeong Kyu Kim is a multidisciplinary laboratory with focus on DNA and protein structure and function. Our ongoing research projects include some proteins involved in protein quality control, signal transduction and pathogenesis, and non-canonical DNA structures such as G-quadruplexes, Z-DNA, and i-motifs. These topics have been received extensive attention as the major questions of current biology. In addition to the major research areas, we have established collaborative relationship with some of leading research groups around the world.