CRISPR Digest #2

Since the CRISPR review mentioned in the last post (Hochstrasser & Doudna (2014)) was published, dealing mainly with the nitty-gritty biochemistry (including crystallography) of the three different CRISPR-Cas systems, another new review has been released. This time co-authored by the two heroes – or should I rather say heroines? – of the CRISPR craze (Doudna & Charpentier (2014)), it is relatively short, easy to read and focusses primarily on the applications of the type II CRISPR-Cas9 system.

An interesting point they make, which I hadn’t thought of before, is that CRISPR offers an excellent opportunity for studying the RNA interference (RNAi) pathway itself. Since the CRISPR system is not naturally present in eukaryotic cells and once expressed from an exogenous source is completely autonomous, it does not compete with other cellular pathways. Therefore CRISPR can be used to target/knock-out components of the RNAi pathway, such as Dicer or Argonaute proteins; this would overcome the difficulty of interpreting data obtained from knocking down expression of these proteins using short interfering RNAs (siRNAs), which themselves rely on the expression of Dicer and Argonaute.

They summarise the overarching ideas and projects involving CRISPR in this diagram (copied directly from the paper):

Doudna & Charpentier (2014) Fig 6

Most of these future applications, such as RNA targeting and gene therapy, were already discussed previously, but the review also includes some even more recent advances and forays into other areas of medicine and agriculture.

For example, it may become possible to use the CRISPR-Cas9 system in its original capacity as an anti-viral defence mechanism to remove proviruses (viruses whose genomes have integrated into the host DNA), such as HIV-1, from infected cells (Hu et al. (2014)).

Probably just as important as the therapeutic/translational applications, but somewhat neglected by me, are the applications of CRISPR in plants and fungi. Several recent papers have shown that genome-editing using CRISPR is possible not only in the plant model organism Arabidopsis thaliana, but also in crop plants such as rice and wheat (e.g. Zhang et al. (2014)). These technological advances will probably speed up a lot of research that aims to improve crop yields and increase resistances to adverse weather conditions as well as infectious diseases.

References:

Doudna JA, Charpentier E (2014) The new frontier of genome engineering with CRISPR-Cas9. Science 346

Hochstrasser ML, Doudna JA (2014) Cutting it close: CRISPR-associated endoribonuclease structure and function. Trends in Biochemical Sciences

Hu WH, Kaminski R, Yang F, Zhang YG, Cosentino L, Li F, Luo BA, Alvarez-Carbonell D, Garcia-Mesa Y, Karn J, Mo XM, Khalili K (2014) RNA-directed gene editing specifically eradicates latent and prevents new HIV-1 infection. Proceedings of the National Academy of Sciences of the United States of America 111: 11461-11466

Zhang H, Zhang JS, Wei PL, Zhang BT, Gou F, Feng ZY, Mao YF, Yang L, Zhang H, Xu NF, Zhu JK (2014) The CRISPR/Cas9 system produces specific and homozygous targeted gene editing in rice in one generation. Plant Biotechnology Journal 12: 797-807

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Interview Season

Apart from the fact that today is “Bridgemas” (i.e. Cambridge Christmas) Boxing Day and that therefore the actual festive season is about to begin, it is also the start of PhD interview season. Next week two other people from the Cambridge biochemistry course and I will head to London to be interviewed (and interrogated about a particular paper that we will have read in advance) at the MRC Clinical Sciences Centre. So anyone reading this who has been through the PhD interview process themselves: I’m sure we would appreciate some advice. For example, were there any questions that completely caught you by surprise? Maybe in hindsight you realised that was an obvious thing to discuss, but it just hadn’t crossed your mind while you were preparing for the interview? What’s the most useful thing you can ask the current students and post-docs in a lab of interest? Any help is welcome!

Lastly, on an unrelated note but regarding one of my favourite subjects, CRISPR: a new review by one of the pioneering CRISPR researchers, Jennifer Doudna, was released only a few days ago (Hochstrasser & Doudna (2014)) and gives a great overview of the three different CRISPR/Cas systems. I like it especially because it focusses mainly on the underlying biochemistry (including Cas protein structures), which is becoming more and more overlooked as many people have started to use CRISPR for genome-engineering on a daily basis.

Reference:

Hochstrasser ML, Doudna JA (2014) Cutting it close: CRISPR-associated endoribonuclease structure and function. Trends in Biochemical Sciences.

Worm vs worms

Two days ago I found myself in a similar situation to that which triggered the beginning of this blog: a friend of mine at university has nominated me to complete the worm vs. worms challenge. I attempted to take a picture of myself cloaked/draped in a duvet, but I failed quite miserably (who knew a selfie would be impossible when covered from head to foot in artificial down?) The challenge was started at the first Polygeia conference, which is documented here:

Polygeia is a “Cambridge-based organisation designed to empower students to research and write policy on global health issues”. The worm vs. worms challenge is meant to spark a wave of donations to the Schistosomiasis Control Initiative (SCI; this video is quite graphic at some points, so you might want to sit down before you watch it):

Schistosomiasis (also known as bilharzia) is the cause of death of approximately 280,000 people every year. Compared to amyotrophic lateral sclerosis (ALS), for example, which affects one to two people per 100,000, schistosomiasis and other related parasite infections are extremely prevalent. The morbidity associated with schistosomiasis is even greater: an estimated 240 million people are infected worldwide. [These data were retrieved from here.]

Compared to the ALS ice-bucket challenge, the SCI makes it a lot more transparent what the donated money is used for (and has the great advantage of not wasting about ten litres of clean water). For example, their website outlines the steps they take to deliver praziquantel and albendazole to children in sub-Saharan Africa.

The drugs, praziquantel (which is not approved for use in humans in the UK) and albendazole, are themselves cheap. Additionally, the WHO provides praziquantel “free of charge to high-disease burden countries in sub-Saharan Africa, through a donation from Merck Serono”. Therefore the main role of SCI is to facilitate the efficient delivery of the drugs to the “right” places.

The SCI was established in 2002 by the Bill and Melinda Gates Foundation’s Global Health Programme and is affiliated with Imperial College London. To me personally, the affiliation with a reputable university increases my trust in the initiative (but maybe this is a biased and misled viewpoint).

Lastly, the treatment and reduction of helminth diseases is bound to have large global social/political/economic implications, but my knowledge of possible outcomes and effects is minimal and other people certainly have more informed opinions about this. What is clear, however, is that whereas to us here (in the the “developed” world) ALS is a much scarier disease than schistosomiasis, to people infected with helminths ALS is completely irrelevant because they will probably die before the onset of symptoms of the neurodegenerative disorder. Yes, my friends, my family and relatives and I are more likely to get ALS than schistosomiasis, but that is an insufficient reason not to donate to SCI.

This is where you can donate if you would like to: Schistosomiasis Control Initiative at Imperial College London.

A Short (Literary) Digression

Instead of bombarding you with too much text I thought I would share this YouTube channel with you, which is run by the journal Cell and features the authors of some of the papers published there as they explain their own research in a few minutes. Admittedly, some videos are more accessible than others, but browsing through is certainly a valid form of procrastination.

On the more literary front, I received a complimentary copy of this book in the post last week:

CareerOpBioSci_f

In fourteen chapters it outlines various career paths that biomedical scientists might take after doing a PhD and post-doc positions, ranging from science policy and patent law to science writing and publishing. Probably the three main take-home messages are that a) it is important to network (whatever that really means), b) one has to be somewhat daring in who one is willing to contact about potential jobs, and c) serendipity is usually involved in the process somewhere.

Moving from scientific self-help book to non-fiction/biography: I finally managed to finish Georgina Ferry’s biography of Max Perutz:

MaxPerutz_f

Perutz was a crystallographer and molecular biologist. Originally from Vienna, he went to Cambridge to do his PhD, where, apart from being deported to Canada during World War II as an “enemy alien”, he stayed until his death. Together with John Kendrew he won the Nobel prize in Chemistry in 1962 for “studies of the structures of globular proteins”. Undoubtedly a great researcher, and possibly an even greater mentor who founded and guided the prestigious Laboratory of Molecular Biology, he was certainly also a slightly difficult person to deal with. His constant psychosomatic illnesses, although certainly real ailments, drove some people up the wall. I wonder whether “great” and famous people become difficult as a result of their greatness and fame, or whether they have to be difficult in the first place to achieve greatness? If it’s the former then I’m not sure I want to become a great researcher, and if it’s the latter then by being quite “normal” (as I would like to believe) it seems unlikely that I ever will be a great researcher.

Although this is now moving from non-fiction to fiction, Candide by Voltaire is still in the realm of philosophy. If you are feeling strong and robust then I recommend this short 18th century novel for a rainy afternoon. But be prepared to be faced with the (irrefutable?) truth that we do not live in the best of all possible worlds, even if, like in this picture, it sometimes might seem that we do.

image1If, on the other hand, you are not feeling up to Voltaire, then why not go see Interstellar? It was valuable in that it taught me some poetry by Dylan Thomas:

Do not go gentle into that good night,
Old age should burn and rave at close of day;
Rage, rage against the dying of the light.

Though wise men at their end know dark is right,
Because their words had forked no lightning they
Do not go gentle into that good night.

Good men, the last wave by, crying how bright
Their frail deeds might have danced in a green bay,
Rage, rage against the dying of the light.

Wild men who caught and sang the sun in flight,
And learn, too late, they grieved it on its way,
Do not go gentle into that good night.

Grave men, near death, who see with blinding sight
Blind eyes could blaze like meteors and be gay,
Rage, rage against the dying of the light.

And you, my father, there on the sad height,
Curse, bless, me now with your fierce tears, I pray.
Do not go gentle into that good night.
Rage, rage against the dying of the light.

Ageing Research – A promise to “cure” all communicable diseases that plague our society?

Possibly one of the most up-and-coming yet controversial areas in biomedical science is the ever-growing field of ageing research. Last week Guy Brown gave introductory lectures on this topic, which were interesting and thought-provoking, especially because I previously had not given ageing much thought (neither as a thing that would affect me personally, nor as a topic of experimentation). His opening slide included this cartoon, which I copied from here:

Aging Researchers

As with most controversial topics, the controversy starts with the definition: currently there is no all-encompassing, universally accepted definition of ageing. However, most people would agree that ageing is a process that involves “progressive physical deterioration over time” (according to Prof. Brown). Furthermore, it is generally accepted that ageing is a major risk factor for a multitude of communicable late-onset diseases such as cancer, diabetes, cardiovascular disease and neurodegenerative diseases.

Therefore, if it were possible to reduce ageing – by which I mean to increase the human health span as opposed to the average/maximum human life span – then could we be preventing these diseases? In other words, should more (financial) efforts be put towards ageing research instead of cancer research, for example? A short and fairly straightforward review (Guarente (2014)) from a proponent of ageing research gives a good, if slightly biased, introduction to the area.

Generally speaking, I would argue that there are two approaches one can take when researching the process of ageing. On the one hand, one can ask why animals (but also single-celled organisms) age, and on the other hand, one can ask how animals age.

As to the why there are a lot of theories, but the main arguments include the following (and it is worth bearing in mind that ageing does not generally occur in “the wild”, since most animals die from extrinsic causes (e.g. predation or infection) before they age):

  • Mutation accumulation theory (Medawar, 1952): as we age we accumulate damage in our cells, and as long as this happens after reproduction (i.e. after selection pressure has been removed to keep our DNA healthy) then this should not adversely affect our offspring’s fitness.
  • Antagonistic pleiotropy theory (Williams, 1957): this theory posits that some genes, which are beneficial early during life and may increase the chances of having healthy offspring, can have detrimental effects later in life.
  • Disposable soma theory (Kirkwood, 1977): after reproduction, which is costly to the individual and uses up a lot resources, an organism is less able to repair damage.

Although these theories may seem logical to a certain extent, it can be easy to start invoking teleological arguments: maybe we age (and die) in order to make space for our children? But how would that information be encoded in our genetic material if ageing (by definition, I would say) occurs after reproduction? Would Darwin be happy with this?

I dare say he might prefer to know how we age. Again, there are a slew of theories regarding the mechanism of (cellular) ageing. Lopez-Otin et al. (2013) attempted to summarise these into nine hallmarks of ageing (in analogy to Hanahan & Weinberg (2011), who have famously done this for cancer). The mechanisms proposed include the accumulation of damage to both DNA (e.g. in the form of mutations) and proteins (e.g. they might start to form toxic aggregates), the exhaustion of stem cell self-renewal capabilities, a deregulation of nutrient sensing and cellular metabolism, as well as chronic inflammation.

You might accuse me of having a pessimistic outlook on life, but from the little I have read about the process and mechanisms of ageing and the still developing area of ageing research, it seems unlikely that vastly different diseases such as cancer and diabetes can easily be tackled from this one approach. Until now (but of course who knows what the future will bring) no treatments have unequivocally been shown to increase human health span. Possibly the most promising to date is a calorie restriction regime. To me personally, however, that seems quite unappealing – do I want to live healthily but hungrily to the age of 100, or satiated but with greater probability of dying by the age of 80?

References:

Guarente L (2014) Aging Research – Where Do We Stand and Where Are We Going? Cell 159: 15-19

Hanahan D, Weinberg RA (2011) Hallmarks of Cancer: The Next Generation. Cell 144: 646-674

Lopez-Otin C, Blasco MA, Partridge L, Serrano M, Kroemer G (2013) The Hallmarks of Aging. Cell 153: 1194-1217