Generation of a family-specific virus through repeated human passage

Generation of a family-specific virus through repeated human passage

Hayden A M Liston¹, Lydia E Makaroff¹ and Adrian Liston ^1,2*
1 Sleepytown University, Brussels 1060, Belgium
² VIB, Leuven 3000, Belgium
*send correspondance to adrian.liston@gmail.com

Nature Junior 8(2) 103-7 

Background. Effective control over viral infection relies on the host carrying appropriate HLA alleles for viral antigen presentation. The explosive expansion of viruses like small-pox into previously isolated human populations demonstrates the potential for certain viral strains to have a disproportionate effect on particular racial groups. As yet, however, a virus with pathogenic potential restricted to the family level has not been identified. Objective. To generate a family-specific virus in an experimental setting, in order to test the feasibility of this occurrence in nature. Methods. A common cold virus was repeatedly passaged between two related individuals for six months. Mechanisms of transmission included frequent kisses, the placement of hands and feet into the mouth and in one instance direct vomiting into the mouth. Results. A single viral strain was propagated with the capacity to chronically infect both members of this family, while having seemingly non-pathological consequences upon exposure to unrelated individuals. The pathogenic loci are predicted to be a dominant HLA carried by both family members, as the experimental inoculation of a third individual, related to one family member but not the other, did not result in pathology. Conclusions. Generation of a family-specific virus is feasible through repeated experimental transfer between family members. A natural situation analogous to the experimental set-up used here would be the transmission that can occur between parents and young children with low levels of personal hygiene. The dominant activity of the HLA cluster in this infection suggests the generation of a regulatory T cell population which inhibits effective immunity against the family-specific virus.

Key Words: virus, horizontal transfer, HLA, human genetics, regulatory T cell.

Advice on applying for an ERC Start Grant (part 3)

I was asked to give some advice on ERC Start Grant applicants, as a current grant holder. As this has come up several times I thought I would write a series of blog posts covering my hints and tips. Partly, this advice is specific to the ERC grant system, although most points are valid across any grant. In a previous posts I gave advice about the written application - Part B1 and Part B2. In this final post I will deal the interview portion of the grant.

The Interview

The interview is not simply an oral version of your written application. There is a panel of around 15 panel members, each of these panel members will be experts on maybe 5 applications and more-or-less bystanders on the other 15 applications.

• Experts. Your chance to impress the experts was your written application, and if you made it to the interview stage than you already succeeded here. The experts are familiar with your work from reading your 30 page dossier; they do not expect to learn anything new from the talk. Instead they will be waiting for the question time to hit you with any issues they have.
• Additional panel members. These are people who are within your general area of research, but outside your specific discipline. They only glossed over your proposal, if they looked at it at all. Design your talk as if they haven’t read your application and focus on importance and strategy. Don’t get bogged down in experimental details and don’t think they really care too much about your discipline – explain to them the advantage in the knowledge that you propose to generate. Focus on the importance and novelty, and why your approach will succeed while others have failed.

Question Time

The questions you get asked will vary based on your project and your application. Have you been wildly ambitious? Expect to get a lot of questions on feasibility. Have you stuck very close by your existing research? Expect to get questions about competitiveness. The experts should ask most of the questions, any technological or methodological concerns they have will be raised here. Generally these will be along the lines of “X is risky, what will you do if it doesn’t work?” or “this is a highly competitive field, how will you compete?” If there is enough time you may get some standard questions from chair or other panel members, such as questions about your long-term career plan and so forth. A few general points apply across the different questions you will get:

• Listen politely to the full question, never assume where it is going or interrupt to answer
• Your tone and attitude matter as much as your words – a grant application is a sales pitch!
• Being right is less important than having a clear articulate message and sounding competent. Even if the expert is wrong there is little benefit in arguing – it certainly comes off badly to the rest of the panel. That said, you can still disagree – “based on my experience the approach is feasible, but in case we do hit a roadblock there is an alternative strategy that we can take...” is completely reasonable response.
• Don’t waffle. It wastes time and it makes it look like you have not thought about the question before. A clear and concise answer reassures the entire panel that you are aware of the issue and have already got a strategy in place. You don’t need an answer for everything, but you need to look like you are capable with dealing with anything.
• Sometimes this involves thinking quickly on your feet and bluffing

On the Day

• Talk clearly and smoothly
• Do not waste time
• Know what you are going to say
• Make every sentence count
• Look at the panel
• Be calm and confident
• Exude gravitas
• Be polite rather than adversarial

On the day of the interview you will arrive at the ERC building, show your passport and be given a visitors badge to enter. You then need to go and upload your talk and deliver ~15 copies of a printed version of your talk before being shown to the waiting room. The room will be full of the other candidates that are being interviewed that day and the wait can be several hours. When your interview is approaching you will be shown up to second waiting room where you will be alone, at this point there is only 10 minutes or so. You will then be led into the interview room. There will be no introductions of the panel members, your talk will already be on the screen and you will be expected to essentially go straight into your presentation.  

Behind the scenes of a panel discussion

In a typical panel, such as the ERC, only a fraction of the applications are read by each panel member. All the panel members are active scientists and all want to support good science. Typically, when going into a panel meeting, each member has a handful of application that they are really keen to push forward – and invariably there is not enough money available to cover all of these applications. In the discussion the experts will take up 90% of the time talking about each grant, but the decision making is split evenly between the panel members. It is not unusual to see an expert trying to convince the rest of the panel that their favourite project is more deserving than your favourite project. In the ERC you have a unique chance to help out the experts on your side, by pitching your talk to the non-experts. If it is dry and technical they will basically ignore it. As an immunologist who regularly sits on an immunology-biochemistry panel I almost fall asleep when there is an application by a structural biologist to find the structure of protein X. So if you are a structural biologist don’t waste your time describing purification strategies to the experts who already read your application – instead use this opportunity to tell the non-structural biologists why this gene is important and what you will be able to do with the structural information (eg, the role of the gene in disease, solid examples of how structural knowledge can be used for rational drug design – perhaps you have a collaboration with chemists?).

Click here for a download of my full set of ERC Start Grant hints and tips.

Advice on applying for an ERC Start Grant (part 2)

I was asked to give some advice on ERC Start Grant applicants, as a current grant holder. As this has come up several times I thought I would write a series of blog posts covering my hints and tips. Partly, this advice is specific to the ERC grant system, although most points are valid across any grant. In a previous post I gave advice for Part B1, in this second post I will deal the written application Part B2.

ERC Start Grant - Part B2

• As a rough length guide, think of ~4 pages for state-of-the-art and objectives, ~2 pages for progress beyond state-of-the-art, ~8 pages for methodology, ~1 page for budget. Adapt to your particular project.
•  You need to be ambitious. Prove that you are thinking as future PI, not as a post-doc. This is not a conservative FWO or IWT grant application, where they pick solid projects. The ERC sees itself more like a MacArthur or Howard Hughes “genius award”, to fund the best and brightest. You can definitely go too far (for example, see the reviewers’ comments that I got from my application), but the panel is generally much more forgiving on over-ambition than under-ambition. The criticism I had on feasibility and over-ambition would have been fatal in an FWO application, but at the ERC the project was approved.

“This is a ground breaking project that interconnects genetic studies, cohort studies and biological studies… It is an extremely ambitious proposal with important and broad objectives and diverse perspectives.”

“Some of the research directions could be difficult to accomplish during the project time, in particular some of the objectives of RT3. Perhaps, the PI should have planned them more realistically.”

“The proposal goes beyond the current state of the art, but its major problem is the over-ambition.”

“The proposed research involves an innovative and ambitious study design, but the risk is justified by the potential impact in the field.”

•  Refer to your unique edge on this project. Is this a direct continuation of your post-doc work? If so, describe how this builds off some technique or tool that you pioneered, giving you an edge over the competition (and either here or in Part B1 make it very clear that you will not be competing with your former PI). Is this a meld of the skills you picked up in your different training periods? Then work in references to strategies you have used in the past. Is this possible due to a unique combination of institute resources or collaborations? Then work in the network you created. Be relatively subtle, the place for direct marketing of your work is Part B1, but references like “using the strategy that I previously designed for gene Y (Jones, Science 2010)” show that you are highly capable of getting this to work.
• The application needs to have an accurate assessment of risk – do you have a back-up plan in case that approach doesn’t work? Why is it that you have a shot of getting this to work while no one else does? (if it is due to your training or past successes, this should be the focus on Part B1). It is not enough to have a grand idea; you need to show that you will have a decent change at success.
• You need to show a future career path. The ERC is not just funding a project, it is funding the start of a new elite laboratory. You need to have tangible outcomes during the 5 year period, but there should also be a sense of how you will build on this after the grant has finished.
• Ethical issues need to show that you have a realistic idea of what is involved, but you do not need to have approval at the time of application (you will need to before you get money from the ERC, however). If it just involves mice a simple referral to an animal ethics committee should be sufficient, if it involves humans or primates you need to demonstrate that you have sufficient knowledge of the ethical and legal framework to make your project practical. 
  

More hints and tips - the interview.

Advice on applying for an ERC Start Grant (part 1)

I was asked to give some advice on ERC Start Grant applicants, as a current grant holder. As this has come up several times I thought I would write a series of blog posts covering my hints and tips. Partly, this advice is specific to the ERC grant system, although most points are valid across any grant. In this first post I will deal the written application Part B1.

ERC Start Grant - Part B1

• Write every part of B1 in the context of the project that you are going to propose - fully utilise every section to sell your application
  
• In your CV you are selling yourself, not describing yourself. Identify your relative strengths and make them stand out. Perhaps you have lots of middle authorships in great journals – then put the journal impact factors in bold, so a quick scan of the page will highlight the great journals rather than your position on them. Perhaps you haven’t published in the top journals, but your work has gathered a disproportionate number of citations – then don’t put the journal impact factor in bold, instead put your individual number of citations in bold.
• Most importantly, when you are presenting your “scientific or scholarly contributions to the field” this is not a generic description. Use this to show how you are uniquely suited to run the project that you have proposed. For example, if you are proposing a project that melds skills you learned from your PhD and your post-doc, place special emphasis on these skills. Your career descriptions should be interwoven with the perspective of where you are going.
• Do not use the extended synopsis in Part B1 to simply summarise the project of Part B2. Use it to discuss the novelty of the approach or the concept. You do not know which part a reviewer will read first, so each document needs to be able to stand alone. Part B2 has a key function in showing that the outcome of your work will be important

Key tip: write about your career projection in the same way you write a scientific paper. You wouldn’t write “we investigated gene X, because of the twelve candidate genes the lab next door had a knockout of this one available”. Instead you would write up results that placed intent and direction in your activity, justifying gene X as your primary focus for a reason. Likewise, don’t describe your career trajectory as it actually occurred, “I did a PhD in metabolism, then my partner moved to Leuven so I looked for a post-doc and got offered one in dendritic cell biology”, rewrite it with intent and direction – “I have had a long-term interest on the impact of metabolism on the innate immune response, so in order to gain skills in both disciplines I first pursued a PhD in biochemistry and afterwards moved to a dendritic cell laboratory. Now I am able to utilise my training in both disciplines, with my independent laboratory focused on the effect of metabolic processes on monocyte activity.”

More hints and tips - Part B2 and the interview.

Scientific thought for the day

Richard Dawkins: "The power of a scientific theory may be measured as a ratio: the number of facts that it explains divided by the number of assumptions it needs to postulate in order to do the explaining."

Autoimmune Genetics Laboratory in the news

In De Staandard, 26th July

KUL en VIB boeken doorbraak in strijd tegen immuunziekten

De K.U.Leuven en het VIB (Vlaams Instituut voor Biotechnologie) hebben een stap voorwaarts gezet in de strijd tegen immuunziekten.

De onderzoekers ontdekten een nieuw type cellen dat kan helpen om het evenwicht te bewaren tussen een overactief en een onvoldoende actief immuunsysteem. Dat laten beide instellingen dinsdag weten in een mededeling.

Talloze mensen lijden aan een ziekte van het afweer- of het immuunsysteem. Als het systeem overactief is, kan dat leiden tot allergieën en auto-immuunziekten zoals Systemische lupus erythematosus (afgekort SLE), een aandoening waarbij het afweersysteem zich op overdreven wijze tegen het eigen lichaam richt. Maar is het afweersysteem onvoldoende actief, dan treden infecties of tumoren op. Het juiste evenwicht vinden, is dus essentieel.

Adrian Liston, van het VIB en de K.U.Leuven, is een nieuw type cellen op het spoor die helpen om dat evenwicht te bewaren. De ’folliculaire regulatorische T-cellen’ (Tfrs) zetten een rem op de groei van afweercellen die antistoffen aanmaken. Verder onderzoek zal het uiteindelijke belang van de Tfr-cellen moeten uitwijzen.

In Het Nieuwsblad, 26th July

KUL en VIB boeken doorbraak in strijd tegen immuunziekten

De K.U.Leuven en het VIB (Vlaams Instituut voor Biotechnologie) hebben een stap voorwaarts gezet in de strijd tegen immuunziekten.

De onderzoekers ontdekten een nieuw type cellen dat kan helpen om het evenwicht te bewaren tussen een overactief en een onvoldoende actief immuunsysteem. Dat laten beide instellingen dinsdag weten in een mededeling.

Talloze mensen lijden aan een ziekte van het afweer- of het immuunsysteem. Als het systeem overactief is, kan dat leiden tot allergieën en auto-immuunziekten zoals Systemische lupus erythematosus (afgekort SLE), een aandoening waarbij het afweersysteem zich op overdreven wijze tegen het eigen lichaam richt. Maar is het afweersysteem onvoldoende actief, dan treden infecties of tumoren op. Het juiste evenwicht vinden, is dus essentieel.

Adrian Liston, van het VIB en de K.U.Leuven, is een nieuw type cellen op het spoor die helpen om dat evenwicht te bewaren. De ’folliculaire regulatorische T-cellen’ (Tfrs) zetten een rem op de groei van afweercellen die antistoffen aanmaken. Verder onderzoek zal het uiteindelijke belang van de Tfr-cellen moeten uitwijzen.

Balance of the immune system determined by newly discovered T cells

From the VIB Press release: A newly discovered cell type helps to determine the balance of the immune system. The cells are derived from FoxP3(+) regulatory T cells, which recently have been demonstrated to suppress disease during transplantation. Nature Medicine and Blood, two high-ranking scientific journals, are publishing findings by the Autoimmune Genetics Laboratory about this discovery.

 

Innumerable people suffer from immune system disorders. If the immune system is overactive, it can result in allergies and autoimmune diseases such as Systemic lupus erythematosus (SLE). If the immune system is not active enough, infections or tumors occur. It is therefore essential to maintain the right balance.

 

The Autoimmune Genetics Laboratory is investigating a new type of cell that helps to maintain this balance. These ‘follicular regulatory T cells’ (Tfrs) suppress the process by which antibodies are produced during infections and SLE. The Tfrs themselves are daughter cells of FoxP3(+) T cells, key regulators of the immune system.

 

From lupus to cancer

In order to prevent disease the immune system needs to be in balance. If the system is too active, it produces antibodies against harmless substances, as is the case with allergies. An overactive immune system can even attack the body’s own tissues, causing autoimmune diseases such as lupus, rheumatism and diabetes. However, an insufficiently active immune system opens the way for rampant infections and tumors.

 

A complex network of regulatory cells is active to maintain this balance in our bodies. This must ensure that the immune system is sufficiently active and selective so that pathogenic intruders are recognized and eliminated in good time, but at the same time that it does not get out of control. This network of regulatory and activating cells is a long way from being fully unraveled. Dozens of researchers around the world are working to map this network.

 

This Tfr research is a joint collaboration between researchers from VIB-K.U.Leuven, the Australian National University (Aus) and the University of Cambridge (U.K.).

 

Publications

Linterman et al, Foxp3(+) follicular regulatory T cells control the germinal center response, Nat Med, 2011, doi:10.1038/nm.2425

 

Tian et al, Foxp3+ regulatory T cells exert asymmetric control over murine helper responses by inducing Th2 cell apoptosis, Blood, 2011, doi:10.1182/blood-2011-04-346056

FWO post-doctoral fellowship awarded to Bénédicte Cauwe

This week it was announced that Dr Bénédicte Cauwe won an FWO post-doctoral fellowship to perform research in the Autoimmune Genetics Laboratory. Dr Cauwe recently finished her PhD in the laboratory of Professor Ghislain Opdenakker at the Rega Institute and will continue her research on systemic lupus erythematosus at the Autoimmune Genetics Laboratory.

Academic independence

What is academic independence?

In the mind of many a post-doc it is quite simple, it is the freedom that you gain when you step up from being a post-doc to becoming a faculty member. As a post-doc, your principle investigator has the final say over your research program, while as a faculty member you are the principle investigator.

It seems straight-forward, but in practice the distinction can be quite blurred. As a senior PhD student in the Goodnow laboratory I effectively had academic independence. My principle investigator had funding and placed trust in me so that I could run my research more or less independently. Hopefully the PhD students in our laboratory feel the same way. Could I have done any hair-brained project I wanted to? Certainly not, it had to be within reason, but the research interests I had were aligned with that of my mentor, so in effect I had the independence to pursue the research that I wanted to pursue.

This is not qualitatively different from the academic independence I have now as a faculty member. Yes, I can chose the research program that I want to pursue, but again the within reason proviso applies. I no longer have a faculty member above me, acting as the final arbiter, but there are still limitations. The most obvious limitation is the grant review process. If I want to do an experiment I require funding, which necessitates my research aims being in line with the granting body and being approved by a panel of experts. Then of course, as junior faculty, I will have a jury over-looking my renewal. These juries invariably have something to say about the direction of your science - your research interests are too broad/too narrow, you are spending too much/ too little time on collaborative ventures, etc. In the modern "big science" era, your colleagues and collaborators form another restraint - you may need to negotiate for time on certain equipment or access to particular samples.

Some of these restraints may be reduced over time, but unless you are a Nobel Prize winner with guaranteed block funding for life there will always be some limitations to academic independence. Perhaps the biggest difference in the academic freedom between a post-doc and faculty member is the diffusion and immediacy of responsibility. As a post-doctoral fellow, the limitations on your research are concentrated in a single person who can have immediate impact - a particular line of research can be shut down today with a single decision. As a faculty member, by contrast, the limitations on your research are delayed and the decision-making capacity is diluted out into a plethora of juries. If one grant foundation chooses not to support your work, another (with a distinct jury) may, and often there are avenues for pursuing research for some months or even years without direct funding.

So rather than the qualitative leap in academic independence that a faculty position represents to some, perhaps it is more accurate to think of a gradual shift in responsibility. Someone moving from a post-doctoral position in a restrictive laboratory to a well-funded start-up faculty position will feel an enormous leap in academic freedom. But for others, being a senior post-doc in a rich laboratory supervised by a figure of benign neglect, the entry into a world of constant grant review may even result in a loss of freedom to pursue your research interest.

IRO fellowship won by Dina Danso-Abeam

Today it was announced that Ms Dina Danso-Abeam in the Autoimmune Genetics Laboratory was awarded an IRO fellowship to perform research towards her PhD.