Double Trouble: IFI44 and IFI44L

An important component of host defence against viral infection is cell intrinsic immunity. This type of immunity is mediated at a cellular level rather than requiring recruitment of other cells to restrict the infection. It is characterised by the induction of an anti-viral state, which limits the ability of viruses to enter cells, make copies of themselves within the cell or exit the cell having replicated. The induction of this anti-viral state is triggered by a signalling molecule called interferon. Interferon signalling leads to the expression of a multitude of interferon stimulated genes (ISG). Many of these ISG are uncharacterised in terms of function.

Technological developments over the last twenty years have changed the way that we investigate how cells work. In particular, the use of transcriptomics, where the messenger RNA (mRNA) in a sample is measured. mRNA is important because it is the intermediary between the cell nucleus, where the genetic information is stored and the ribosome, where proteins are made. Transcriptomics gives an overview of what the cell is doing. However, transcriptomics is a broad-brush tool that does not necessarily give the fine detail of what individual genes do in the prevention of infection.

Over the last few years we have undertaken a program of work to understand the role of individual ISG in the control of viral infection. In particular we are interested in respiratory syncytial virus (RSV). RSV infects the lungs of children – all children will be infected with it before the age of 2 years old, most before 6 months of age. Some of these children will get extremely sick with RSV infection and we hypothesized that this is because they fail to control the virus early on during infection. However, prioritising which ISG to investigate was an issue, especially given the large amount of data available. We therefore used a screening process to identify those genes which are more commonly associated with RSV infection (https://doi.org/10.1128/mSystems.00051-16).

This screening process led us to work on a pair of genes called Interferon-induced protein 44 (IFI44) and interferon-induced protein 44-like (IFI44L) which we published in the Journal of Virology (https://jvi.asm.org/content/early/2020/06/26/JVI.00297-20). We confirmed that the genes were induced following RSV infection and then set about exploring whether they played a role in the control of infection. The first question was what would happen in the absence of either gene. Using two different gene-knockout approaches, CRISPR-cas and siRNA, we showed that when you reduce expression of either gene, the virus replicated better. We then did the opposite experiment, increasing the amount of both genes in the cells, this led to decreased viral replication. These initial findings were supported by studies in mice and children. Mice lacking the IFI44 gene were more susceptible to RSV infection and children with lower expression levels of the gene, as determined using transcriptomics on their blood, were more likely to have a more severe infection – though this was a weak association.

The question remains as to how IFI44 and IFI44L prevent viral infection. One of our observations was that altering the levels of the two genes altered the ability of cells themselves to replicate. When there was more IFI44, the cells replicated more slowly, when it was removed they replicated faster. We think that this gives us a clue as to their function – somehow they limit resources that both the cells and the virus need to make more copies of themselves. We are now looking to understand exactly how this happens. What is fascinating is that there are so many different genes involved in the prevention of viral infection and an important question is how do they interact to protect us.

It takes a Village

First published in Times Higher Education

The greater prominence enjoyed by scientists during the Covid-19 pandemic has led to some individuals gaining a high profile – with the attendant praise and demonisation that this can bring. But these public figures are just the visible tip of a huge iceberg of effort taking place to combat the pandemic.

To convert one bright idea into 7.5 billion doses of vaccine will take a huge team of people. This includes not just the lab team developing and test the vaccine, but also the animal care staff enabling the pre-clinical studies, the safety staff maintaining a safe environment to work with a potentially fatal pathogen, the lab managers ensuring that essential reagents are available, the administrators preparing the relevant grant applications, the ethics boards reviewing the trials and the trial managers, doctors, nurses, med-students and volunteers. Not to mention the contracts team negotiating with equipment manufacturers, the accountants moving the money around, the security officers keeping the doors open and the communications experts informing the public of progress.

That is just at one institution. And the work is not performed in isolation: there are external funders, suppliers, manufacturers, regulators, toxicologists, shippers, couriers and warehouse staff, all of whom are vital to the process.

Unlike the standard image of an old white academic, staff in professional roles tend to be more diverse with more women, more BAME and more LGBT. But, in the UK, they will be excluded by the government’s proposed post-EU new immigration rules. This would be deeply counter-productive: if the pool of skilled individuals is reduced, there will be a clear impact on the ability to deliver cutting edge research, particularly in a time of crisis.

Highlighting the role of these critical core staff is vital. They are often under-represented in the media. For example, coverage of the recent UK pension strikes focused on the academics taking part, rather than on all the other higher education staff who shared the picket lines with them.

As well as not accurately reflecting science as a collective endeavour, a focus on individuals can, in fact, be toxic. Much of what is wrong with academia is driven by the narrative that it is a zero-sum game, where only one person can come out on top. This leads to the back-stabbing, bullying and bitchiness that characterises the very worst of our sector.

Now as never before, kindness in the workplace is critical. Developing the vaccine that the world so desperately needs can serve as a demonstration that great things can be done collaboratively rather than competitively, belying the inaccurate depiction of it in some places as a race between different universities. In the UK’s case, the race is supposedly between the University of Oxford and Imperial College London – but the fact that some of the ChAdOx (Oxford vaccine) trials are being performed at Imperial tells a very different story.

Thinking ahead, maybe we can use this time as a trigger to rethink the whole of academia. The first step is acknowledging that it is about more than the academics. It’s been said before, but when you look at the numbers, academia is actually the alternative career for science trainees: most enter other sectors – including academic support roles. All these paths should be supported and celebrated equally.

If none of the above persuades you, then consider this. Representing the team nature of science de-risks the process for the individuals, the institutions and the ideas themselves. People sometimes make mistakes, often unrelated to the science itself, but this can tarnish the idea. In an increasingly combative media space, any perceived fault can be manipulated to damage a broader theme. Demonstrating that science is collective removes one tool from the arsenal of those that seek to discredit ideas that have universal benefit, such as vaccination or combatting climate change.

The Wellcome Trust’s Reimagine Research campaign is currently looking into ways to rebalance the research space. But you don’t need to be a funder to make a difference. We can all play our part to make higher education kinder and more inclusive. Take time to say thanks. Reach out to teams outside your immediate remit. Be public in your praise, raising awareness of the whole team, not just the star signing. Applaud the whole community of effort.

Let me start and make an Oscars style acknowledgement of some of the amazing team at Imperial (and sorry if I missed you) – thank you Kasia, Kat, Kai, K, Krunal, Kostas, Catherine, Tessa, Anna, Hannah, Hadi, Michelle, Glenda, Genevieve, Paul, Paul, Carolyn, Ruth, Jennifer, Tom, Lesley, Sharron, Jesses, Leon, Aaron and Jo –I know that most people reading this will not know who they are, but without them we might never get back to work.

The Coronavirus Diaries

Head on over to Nature to read my diaries under Coronavirus Lockdown:

Week 1: Who I am https://www.nature.com/articles/d41586-020-01076-3

Week 2: On being a scientist without a lab https://www.nature.com/articles/d41586-020-01138-6

Week 3: On home schooling https://www.nature.com/articles/d41586-020-01293-w

Week 4: On getting distracted https://www.nature.com/articles/d41586-020-01323-7

Week 5: Things I don't miss https://www.nature.com/articles/d41586-020-01395-5

Week 6: People I do miss https://www.nature.com/articles/d41586-020-01476-5

Week 7: Getting ideas https://www.nature.com/articles/d41586-020-01555-7

Week 8: Getting rejected https://www.nature.com/articles/d41586-020-01624-x

Week 9: Taking a break not a holiday: https://www.nature.com/articles/d41586-020-01696-9

Week 10: Back to the lab again: https://www.nature.com/articles/d41586-020-01782-y

Week 11: Learning from my son: https://www.nature.com/articles/d41586-020-01867-8

Week 12: Unsocial Media: https://www.nature.com/articles/d41586-020-01948-8

Let it grow (Frozen Parody)

The DNA glows orange on the gel tonight

Not a fingerprint to be seen

A kingdom of eukaryotes

And I’m looking for the gene

The centrifuge is howling because it's unbalanced inside

Couldn't get the genes in, heaven knows I've tried

Won't let my genes in, won’t work for me

Be the good clone you always have to be

Anneal, don't heal, won't bloody grow

And they still don’t grow

Let it A, let it G
Can't clone shit anymore
Let it T, let it C
Turn away and slam the incubat-or
I don't care what they're going to say
Let the Taq rage on
The heat never bothered it anyway

It's funny how Magnesium makes every band less small
And the PCR that once controlled me can't get to me at all
It's time to see what I can do
To insert that gene and break on through
No blue, all white, great clones for me
I'm free

Let it A, let it G
I am one with the DNA
Let it T, let it C
You'll never see me cry
Here I stand and here I stay
Let the project rage on

My power flurries through the lab and all around
My science is spiraling from my pipette to my hand
And one thought crystallizes like an icy blast
I'm never going back, the past is in the past

Let it A, let it G
And I’ll pipette like I’m a boss

Let it T, let it C
That perfect gene is cloned
Here I stand in the light of day
Bring the western on,

Molecular biology never bothered me anyway.

How does flu affect poo?

There are lots of motivations to do science – to solve a grand global challenge, to answer a fundamental question about how the world works, because it’s a job and you’ve got to do something before pub opening times. For me, it is about solving puzzles – piecing together a coherent story from a jumble of observations.

In our most recently published paper, we got to do exactly that, and best of all, we got to draw it all on a white board, like in CSI.

Solving science problems, one whiteboard at a time

We started with an observation (from a previously published paper). When mice get viral lung infections, the bacteria in their guts (the gut microbiome) changes. This was quite surprising. So in the follow up we wanted to understand why, but also to understand if the changes in the microbiome affected anything.

But first of all, why we did care at all. As a brief reminder, it is entirely normal to have gut bacteria, in fact various estimates put the number of cells of gut bacteria as higher than the cells of person surrounding them. The gut bacteria do a whole range of important things, from breaking down your food for you to regulating your immune system.

Back to the story. One other thing we did know was that when infected with a virus, mice lose weight. In fact, relative to body size they lose quite a lot of weight, up to 25%, which they can put back on very quickly. This was our starting point and we speculated it could be due to a number of reasons, including the increased effort of breathing. Helen (the student working on the project) made the very simple step of measuring the amount of food eaten. She found that over the course of the infection, the mice stopped eating and it was this that caused the weight loss. When we mimicked this reduction in eating by reducing the amount of food the mice had per day, we saw an identical change in the gut microbiome. So problem one solved.

However, this then led to the next question – why do the mice stop eating? We drew on our previous studies, in which we had showed that the immune response to infection was associated with weight loss after infection. Specifically we were interested in an immune cell called the CD8 T cell and a signalling molecule called Tumour Necrosis Factor (TNF), which the immune system releases to activate other cells. We used antibodies (molecules that are recognise other proteins very specifically) to block CD8 T cells or TNF to see if the immune response was responsible for weight loss. To cut the story short, T cells were important, TNF not so much. It still begs the question, why does the immune system cause weight loss. At this point the simple answer is, we don’t know.

We then switched our attention to the downstream effects. One thing we wanted to see was if the change in the gut bacteria, changed susceptibility to other gut infections. In our hands, they didn’t – when we infected mice with a lung virus, there was no effect on a subsequent gut infection. However, we did observe one difference after infection. There is a complex network of chemicals in your guts, called the metabolome. After infection, these biochemicals in the guts changed. What was intriguing was that some of the chemicals that increased have previously been described as anti-inflammatory. One crazy speculation from this is that by stopping eating the mice actually get better, but we have no proof that this is true and it might just be a correlation.

So what did we learn? When mice are infected with a respiratory virus (in their lungs) their gut microbes change. The gut microbes change because they lose weight. They lose weight because they stop eating. And they stop eating because of something to do with the immune system. So problem solved, sort of.

Coronavirus Q and A

What is the ‘Wuhan coronavirus (2019-nCoV)’ and what do we know about it so far?

Dr John Tregoning from Imperial’s Department of Infectious Disease spoke to the School of Public Health’s Prof Steven Riley about the ‘Wuhan coronavirus’ outbreak that recently began in China.

·       Who has been working on the outbreak epidemiology at Imperial College London?

SR: I work as part of the MRC Centre for Global Infectious Disease  Analysis and the Abdul Latif Jameel Institute for Disease and Emergency Analytics centre with Professor Neil Ferguson, Dr Natsuko Imai, Dr Ilaria Dorigatti and Dr Anne Cori.

·       So what is the ‘Wuhan coronavirus'?

It is a viral infection that was first discovered in the Chinese city of Wuhan in 2019 that has been associated with a number of cases of pneumonia – an infection of the tissue in the lungs. You might see it being called ‘2019-nCoV’, which stand for novel (or new) coronavirus. More information has been provided by the World Health Organisation.

·       What is a coronavirus?

JT: Viruses are infectious organisms that rely upon the cells in our bodies to replicate. A virus needs to enter our cells and hijack them to make copies of itself. They enter our cells by sticking to the outside of the cell, using viral proteins to recognise proteins made by the human cells.

Coronavirus are respiratory viruses, which means they are viruses that infect the nose and the lungs. They are a large family of different viruses causing a range of different illnesses from colds to more severe diseases. The coronavirus family is known to be potentially zoonotic, so able to jump between different species. They are from a broader group of viruses called RNA viruses, which means their genetic material is carried on RNA molecules, not DNA molecules, which is important when it comes to thinking about how they can mutate.

·       What are the symptoms?

SR: Since nCoV is a respiratory virus, it will cause symptoms ranging from a cold (blocked nose) and a cough, to chest infection and pneumonia. Fever (a temperature over 38°C or 100.4°F) has been commonly observed with nCoV infections. As the virus was first identified in a cluster of pneumonia patients, we can probably assume that it can cause pneumonia in the more severe cases.

·       Is it anything like SARS, MERS or Ebola?

SR: The novel coronavirus (nCoV) associated with this outbreak, is somewhat similar to SARS (Severe acute respiratory syndrome) which emerged in 2003 and MERS (Middle east respiratory syndrome) which emerged in 2012. Both of these infections, SARS and MERS, are caused by coronaviruses and are respiratory infections.

It is nothing like Ebola, which is caused by a different virus type altogether (Filoviridae). Ebola spreads from person to person by contact with bodily fluids from infected individuals.

·       How does nCoV spread?

SR: We don’t fully know. However, we can compare it to other respiratory viruses. Most of which are spread by respiratory droplets, for example from a sneeze to a hand to a surface, which is then picked up by a new person who then touches their face. Sometimes the virus can be airborne and then inhaled, but it’s more difficult to measure how much transmission happens this way.

·       How was it discovered?

SR: There was a cluster of pneumonia cases in Wuhan, China. Genetic material was isolated from these patients and this coronavirus was found. The team that found it have rapidly shared this information with the global research community, which has enabled research towards new vaccines and diagnostics.

·       How do you test patients for a new virus like this?

SR: Following the identification of the virus causing the infection, we can use pre-existing technology called PCR to test patients. PCR (polymerase chain reaction) is a highly specific test that recognises a genetic sequence and amplifies it so that its presence can be detected.

·       Where did it come from – did it ‘jump’ from reptiles or bats?

SR: The first cases have been closely tied to a specific market in Wuhan. These no reliable evidence for it coming from snakes. The closest known virus is found in bats, but we don’t know for sure if this is where it came from.

·       And how did it jump to humans?

JT: Viruses, and in particular RNA viruses, mutate over time. This is because when they use your cells to make copies of themselves they have poor proofreading, so each copy is not quite the same as the original. Most of these changes will make the virus less infectious, but some might enable them to infect a slightly different type of cell or species.

SR: For other zoonotic viruses (viruses which spread between animals and humans), the viruses normally move from a species with which we are closely physically associated, so for example influenza can move from pigs and chickens to people. The huge number of animals raised for meat and their close physical proximity to people can make these jumps more likely.

·       How is it spreading now?

SR: One of the tools we use as epidemiologists is the R0 value. This is the number of new individuals infected by the first infected person. So for example an R0 of 2 means that each infected person infects 2 more people: in an unbroken transmission chain these 2 people would then infect 2 more each – so 2 becomes 4, which becomes 8 and so on. We then want to apply behavioural and treatment approaches to reduce the R0 to less than 1 so the infection contracts. We do not know the R0 value for nCoV yet and it is too early to be confident about what it is.

·       Is the virus mutating?

SR: It is very hard to tell. Viruses mutate all the time as part of their replication process. It’s very difficult to detect significant mutations that really change the behaviour of a virus.

·       Is this virus any more or less dangerous than seasonal flu?

SR: We don’t know. We are concerned that it is more dangerous than the 2009 strain of influenza, which was milder than other influenza pandemics. One way to think about this is the difference between case fatality rate and infection fatality rate. Very roughly speaking case fatality rate (CFR) is the number of deaths per confirmed cases of the virus (so in this example people who have gone to a doctor or a hospital and had a confirmed diagnosis of the virus). Infection fatality rate (IFR) is the number of deaths of all the people who have been infected.

·       Why is the difference between IFR and CFR important?

JT: If the majority of people who get infected with the virus do not have severe enough illness to need to go to hospital or the Doctor, then the case fatality rate will be higher than the infection fatality rate and the disease will be less serious than it appears. Essentially clinical cases, those that require medical assistance is the tip of an iceberg and the question is how big is the underlying iceberg? If most people infected don’t develop any symptoms, then it is a mild disease, if most people infected need hospitalisation, it is a cause for concern.

·       Why don’t our existing vaccines or antiviral drugs work?

JT: Vaccines work by training your immune system to recognise specific features (called antigens) of the virus it is protecting you against. Since this is a new virus, which looks different to other viruses and has different antigens, current vaccines cannot provide protection.

·       Why don’t we have a new vaccine?

JT speaking to Professor Robin Shattock (Imperial College London). Vaccines take time to manufacture, even with our best new platforms, any new vaccine takes at least 3 months to manufacture enough material to test in people for safety. It is then a big step from there to manufacture enough doses of vaccine to cover the world population.

Likewise, antiviral drugs target key components of the viral replication. They have been developed for other families of viruses and so are not necessarily specific enough to inhibit the coronaviruses.

·       Is there anything people can do to reduce their risk: for example wearing facemasks or washing hands?

JT: Masks are important in clinical settings when properly used, however they have little value for the general public. Handwashing and reducing contact from hands to face can be helpful, as this helps stop the spread of the virus through respiratory droplets from coughs and sneezes.

·       Will the quarantine in Wuhan work?

SR: It is an unprecedented step, so we have no evidence either way. But it is a strong statement from the government and this will increase the awareness of the infection and therefore reduce spread.