Model of a human hepatitis B disease virus capsid. (Credit: theasis)
Hepatitis B virus (HBV) affects 300 million people worldwide and is a major driver of liver diseases. But while decades of research have led to vaccines, antivirals, and a better understanding of the virus, how this pathogen establishes persistent infection in cells at the molecular level remains poorly understood.
Now, a collaboration between The Rockefeller University, Memorial Sloan Kettering Cancer Center (MSK), and Weill Cornell Medicine has uncovered a key mechanism that allows HBV to infect liver cells. Their research, published in Cell, suggests that the virus hijacks host chromatin structures to activate its own genes. The findings solve a long-standing mystery about the basic biology of the virus while also pointing to an anticancer drug candidate that, at relatively low doses, may disrupt HBV's ability to establish long-lasting infection in liver cells.
"We were delighted to discover not only important details about the mechanisms of HBV gene regulation, but also a promising path toward a new therapeutic tool against HBV," says Viviana I. Risca, head of the Laboratory of Genome Architecture and Dynamics at Rockefeller.
A chicken-egg problem
Hepatitis B infection is a biological paradox. Studies have shown that host cells will shut down viral gene expression unless the host's defenses can be counteracted by the viral protein HBx. Yet that protein cannot be produced without viral gene expression. It's a chicken-and-egg problem-HBx protects the very process that makes it-indicating that the timing of the earliest molecular events in HBV infection is critical to determining a winner in the battle between HBx and its host. The precise choreography of these steps is a puzzle urgently in need of a solution; the gene that encodes HBx to initiate infection is also an oncogene, known to help transform chronic HBV infections into deadly liver cancers.
To unveil the mysteries of HBx, first author Nicholas Prescott, a graduate student in the Tri-Institutional Program in Chemical Biology, and MSK chemical biologist Yael David started by generating a model for HBV infection in vitro, with the help of Robert Schwartz at Weill Cornell, whose lab contributed its biological and clinical expertise in the virus, along with human liver cell models. Their platform was uniquely well suited for interrogating the earliest events in HBV infection and gene expression. It revealed that the HBV X gene, which produces HBx protein, is particularly sensitive to the formation of nucleosomes-the basic units of chromatin, made of histone proteins, which package and organize DNA to regulate gene activity. HBV's resulting "mini chromosome" promotes HBV X gene transcription.
This provided a mechanism for how HBV solves the chicken-and-egg problem, because the X gene is the earliest to be expressed, hobbling the host cell's defenses before it can catch up to the virus.
To ensure that their platform reflected the realities of human infection, the researchers turned to Risca, an expert in mapping how DNA interacts with chromatin to regulate gene expression. Using high-resolution nucleosome mapping, the Risca lab analyzed how HBV's DNA interacts with host histones to form chromatin structures. Their work confirmed that the mini-chromosome closely resembled how HBV's genome organizes in infected liver cells.
Risca and colleagues were surprised to discover that the way the HBx gene is packed inside the virus determines whether it gets activated. "Conventional wisdom says that packaging a gene's DNA into nucleosomes would block or slow down the cell's ability to read out that gene to make functional proteins, like X," Risca says. "But in complex organisms like humans and in the viruses that infect us, gene regulation is not always so straightforward. We found that to be the case for the HBV gene encoding protein X-the presence of nucleosomes on the viral genome is necessary for the transcription of RNA that gives rise to functional protein X."
New treatments on the horizon
The findings shed new light on the basic biology of a virus that infects millions, and also may lead to sorely needed therapies. Because while HBV is treatable, it is hardly beaten. Antivirals only stop the virus from making new copies of itself, rather than clearing its DNA from the cells entirely; vaccines prevent the disease, but maintaining immunity requires frequent booster shots.
With this in mind, the team tested five small molecules known to disrupt formation of the chromatin structures that the virus needs to begin expressing HBx. One of these compounds, an anticancer drug candidate, blocked the production of HBx protein in liver cells. The drug was effective at relatively small doses, which appeared to stymie the virus while sparing human cells. If proven safe and effective, this small molecule could also treat other chromatinized DNA viruses, such as herpesviruses, papillomaviruses, and adenoviruses, according to the study.
But first, the team will need to study this drug candidate in animal models. If successful, such trials could mark a turning point in HBV treatment-targeting the virus at its root rather than merely suppressing its replication.
The researchers noted that these breakthroughs wouldn't have been possible without the close collaboration between the three institutions, which brought together the necessary expertise and technological resources-from MSK's atomic force microscopy imaging at its Molecular Cytology Core to the Genomics Resource Center and High-Performance Computing Cluster at The Rockefeller University. "The HBV genome is a fascinating microcosm of gene regulation," Risca says. "Studying it tells us not only about the virus, but also about how our own gene regulation works. Curiosity about these basic questions motivated us to collaborate."
"This is a great example of how investment in 'basic science' and investigation of fundamental biological questions can open the door to medical advances," says Prescott, who is now a postdoctoral fellow in Rockefeller's Laboratory of Chromosome and Cell Biology headed by Hironori Funabiki. "Never in a million years did I expect to lead a project that identified such a strong candidate for drug development for a global scourge like hepatitis B."
