Understanding the Molecular Basis of Avian Influenza Host Shift

A recent study by the Cusack group clarifies how a crucial enzyme of the avian influenza virus can change, enabling the virus to spread to mammals.

The impact of seasonal flu epidemics brought on by human influenza viruses A and B has been significantly reduced in recent years through vaccination, surveillance, and public health initiatives. However, there is a serious risk to the public's health from a potential outbreak of avian influenza A, also referred to as "bird flu," in mammals, including humans.

The Cusack group at EMBL Grenoble investigates how influenza viruses replicate. A recent study conducted by this group clarified the various mutations that the avian influenza virus can undergo to replicate in mammalian cells.

Certain strains of avian influenza can result in fatalities and serious illness. Fortunately, avian influenza rarely spreads from birds to other species due to the substantial biological differences between birds and mammals. The avian influenza virus must mutate twice to infect mammals: once to enter the cell and once to multiply inside it. It also needs to be able to spread among people to start an epidemic or pandemic.

On the other hand, it is becoming more frequent for wild and domestic mammals to become occasionally infected with bird flu. The sudden infection of dairy cows in the USA by the avian H5N1 strain, which runs the risk of becoming endemic in cattle, is especially concerning.

This could make it easier for the organism to adapt to humans. In fact, there have been a few documented cases of transmission that have only produced mild symptoms thus far.

The enzyme polymerase, which directs the virus's replication inside host cells, is at the center of this process. This adaptable protein can reorganize itself to suit its various roles during infection. These include replication, which involves creating duplicates of the viral RNA to encase new viruses, and transcription, which involves converting the viral RNA into messenger RNA to produce viral proteins.

Due to the involvement of two viral polymerases and the host cell protein ANP32, viral replication is difficult to study. These three proteins work together to form the replication complex, a molecular apparatus that replicates. ANP32 is referred to as a "chaperone," which denotes that it stabilizes specific cellular proteins.

Its long, acidic tail is one important structure that allows it to do this. Although ANP32's role in influenza virus replication was identified in 2015, it remained largely unclear.

The study, published in the journal Nature Communications, demonstrates that ANP32 serves as a link between the replicase and encapsidase viral polymerases. The names refer to the two different conformations that the polymerases adopted to carry out their two distinct tasks: replicating the viral RNA (replicase) and encasing the copy with the assistance of ANP32 to form a protective coating (encapsidase).

ANP32 stabilizes the replication complex through its tail, enabling the complex to form inside the host cell. While the protein's core is largely the same, the ANP32 tail varies between birds and mammals. This biological difference explains why the avian influenza virus does not replicate as readily in mammals as it does in humans.

The key difference between avian and human ANP32 is a 33-amino-acid insertion in the avian tail, and the polymerase has to adapt to this difference. For the avian-adapted polymerase to replicate in human cells, it must acquire certain mutations to be able to use human ANP32.”

Benoît Arragain, Postdoctoral Fellow and Study First Author, EMBL Grenoble

Arragain and colleagues obtained the structure of the replicase and encapsidase conformations of a human-adapted avian influenza polymerase (from strain H7N9) while it was interacting with human ANP32 to understand this process better.

The precise information provided by this structure explains which amino acids are required to form the replication complex and what mutations might make it possible for the avian influenza polymerase to adapt to mammalian cells.

Arragain conducted in vitro experiments at EMBL Grenoble, utilizing the cryo-electron microscopy platform provided by the Partnership for Structural Biology, the ISBG biophysical platform, and the Eukaryotic Expression Facility to achieve these results.

We also collaborated with the Naffakh group at the Institut Pasteur, who carried out cellular experiments. In addition, we obtained the structure of the human type B influenza replication complex, which is similar to that of influenza A. The cellular experiments confirmed our structural data.”

Benoît Arragain, Postdoctoral Fellow and Study First Author, EMBL Grenoble

Using these new insights into the influenza replication complex, we can study polymerase mutations in similar strains of the avian influenza virus. Thus, the structure derived from the H7N9 strain can be modified to fit other strains, like H5N1.

The threat of a new pandemic caused by highly pathogenic, human-adapted avian influenza strains with a high mortality rate needs to be taken seriously. One of the key responses to this threat includes monitoring mutations in the virus in the field. Knowing this structure allows us to interpret these mutations and assess if a strain is on the path of adaptation to infect and transmit between mammals.”

Stephen Cusack, Senior Scientist and Study Lead, EMBL Grenoble

Cusack has been studying influenza viruses for 30 years.

These findings also benefit the long-term view of anti-influenza drug development since no medications currently on the market specifically target the replication complex.

“But it’s just the beginning. What we want to do next is to understand how the replication complex works dynamically, in other words, to know in more detail how it actively performs replication,” said Stephen Cusack.

The group has successfully carried out similar studies on the role of influenza polymerase in the viral transcription process.