1001 Genomes

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A resource that we are increasingly using in our research is what’s known as the ‘1001 Genomes‘.

This is where the genome sequences for over 1000 different Arabidopsis plants are available for anyone to play with.

Just like you and me, we are all slightly different. Different hair colour, different height, different shoe size. The same is true for plants. The sequences for the 1001 genomes  helps us work out how Arabidopsis has evolved, and might therefore help us understand how, for example, climate change will affect plants (and ultimately our food crops). For our research, we’re interested in how plants perceive and respond to temperature. How do plants survive and adapt to very different temperature environments?  We can use the 1001 Genomes resource to help us address this question.

The flagship paper that describes this invaluable resource is a paper published by the 1001 Genomes Consortium in the journal Cell in 2016.

The paper shows that Arabidopsis ‘relict’ plants – something akin to the plant’s Founding Fathers – were prominent in the Iberian (Spain and Portugal) Peninsula – and seemed to hang about on the periphery of the last ice age (around 12,000 years ago), whereupon there was an expansion, or ‘sweep’ into more Northerly latitudes. What changes to the genome helped Arabidopsis survive in different habitats in their sweep North?

As pointed out in their Conclusion the authors state that “temperature and precipitation vary greatly across the species’ range and between groups and one would expect differences in physiological and developmental responses of Spanish and Swedish accessions”.

Why is all this important? Well, its been demonstrated that rice grain yield declines by 10% for each 1°C increase in growing-season minimum (i.e. night-time) temperature. One approach might be to therefore grow crops at higher latitudes, but by doing this our crops will need to adapt to different day-lengths. A higher latitude results in greater seasonality i.e. larger differences in day-length and temperature at higher latitudes compared to regions nearer the equator.

Why don’t we work directly with rice, wheat, barley, potato etc? Why do we work with Arabidopsis, which is a weed after all. Well the genome size of our staple crop plants are much bigger and more complex and so obtaining 1001 potato genomes would be a truly mammoth (and expensive) task. We also have a wealth of genetic resources available for Arabidopsis,  And for me – never known for my horticultural skills – Arabidopsis is easy to grow (its a weed after all), and it lives and dies quickly (around 5 weeks) meaning that there can be a quick turn around of experiments.

There are some good web-resources that allows us to play around with the 1001+ sequences from all of these different natural variants. One that I find particularly useful is the SALK 1001 genomes browser, where you can plot all the single base pair changes across a gene region for as many of the 1001 genomes you can fit on your web-browser – see an example below.

Makes you wonder, looking at all of these small changes in DNA sequence – what do they mean for the plant, and how best do we test what these changes make?

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12h L:D

tex toc coldLL gating cartoon

Today is the vernal equinox, and traditionally marks the beginning of Spring. It’s also when  daytime and night-time are of approximately equal duration.

This has resonance with our (artificial) experimental set-up of day:night (dark:light) for our plants growing in environmentally controlled cabinets. When we present our circadian data we would typically denote ‘the day’ on our slides or papers as white and black bars denoting day and night, respectively.

I suppose today is the day we should be doing our experiments in Nature instead of the growth cabinets…

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abundance of transcript=amount of protein?

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Thought that this was an interesting paper, published recently in PNAS.

The paper shows that alternative splicing produces different transcript isoforms for the 5’UTR region of the human gene encoding α-1-antitrypsin called SERPINA1, such that splicing of 5’UTR modulates the inclusion of long upstream ORFs (uORFs). What’s new with all this I hear you say. Well, the authors go on to show that while SERPINA1 transcripts produce the same protein isoform, they do so with different translation efficiencies. Differences in uORF content and 5’UTR secondary structure combine to differentiate the translational efficiencies of SERPINA1 transcripts.

α-1-antitrypsin is of interest because deficiencies in this protein are associated with chronic obstructive pulmonary disease (COPD), liver disease, and asthma. This work points to the possibility that genetic alterations in noncoding gene regions, such as the 5’UTR region, could result in α-1-antitrypsin deficiency.

The work also reinforces the idea that the amount of protein produced from a gene is not a simple function of the abundance of the transcript.

The reference is: Proc Natl Acad Sci U S A. 2017 Nov 21;114(47):E10244-E10253. doi: 10.1073/pnas.1706539114. Epub 2017 Nov 6.

The image used is their Figure 3. SHAPE-MaP structure probing data for SERPINA1 transcripts.

We are Detectorists

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Loved the whimsical comedy of the ‘Detectorists‘ series with the excellent MacKenzie Crook (as Andy Stone) and Toby Jones (as Lance Stater). It follows Lance and Andy’s lives as their personal situations and ambitions orbit their central obsession with metal detecting.

It made me think of some analogies with Science. Obsession – yeah, tick. Do we ever stop thinking about our research topic? From hours of scanning the landscape only to uncover ‘ring-pull’ duds (yeah tick – see ‘failed experiments, no?). Or is it that we are just centimetres away from that Anglo-Saxon gold hoard.  Maybe we have selected the wrong ‘field’ in the first place. I think that, like Lance and Andy, most of us dream of landing ‘the big one’ –  maybe that discovery that will be transformative? We can but dream?

Especially loved the concept of the “discovery dance” – what kind of celebration would you do when the the game changing discovery comes your way….can you practice it? should you practice it?…or should it come as a natural reaction?

Here’s a link to a brief YouTube clip:

I think we are all Detectorists at heart. Think I’ll get practicing my dance….

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Seed spotting

After overfilling the tubes with the agar/growth media a cross is sliced into the agar with a sterile blade (see clip above).  Doing this his will hopefully help the roots find a way down through the agar/growth media.

Then the Arabidopsis seeds (sterilised in bleach and suspended in sterile water) are pipetted onto the surface of the agar one-by-one (ok, maybe sometimes two-by two) – see clip below.

 

The trays are then sealed with micropore tape and that’s it – now to wait to see if the seeds germinate – should take around a week to see the emergence of the very young shoots.

 

Eat, Sleep, Pipette, repeat

 

Setting up the next experiment. It involves pipetting agar growth media into (literally) 100’s of tubes – see movie clip above. I try to overfill the tubes with the molten agar mixture (this is important for a later step….where I place single Arabidopsis seeds on the agar surface…watch this space).

Science is 99% perspiration, and 1% inspiration….and this is the ‘perspiration’ bit.

 

 

“now acceptable for publication” :-)

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Ahhh….the best 4 words in scientific research? It’s been a long, arduous trip but finally we shall be adding a few dents to our current knowledge of alternative splicing/splicing factors/temperature and the clock….will keep you posted.

Made me think about the time it takes to publish scientific research, and I came across this commentary article in Nature from 2016.

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I think many of us working at the coal-face of research will recognise a lot of what it says, e.g.

Many….feel trapped in a cycle of submission, rejection, review, re-review and re-re-review that seems to eat up months of their lives, interfere with job, grant and tenure applications and slow down the dissemination of results.”

Also talks about “resetting the clock” – not to do with circadian clocks, but related to the time stamp of submission and resubmission(s).

Is it taking longer to publish? One contributor to the article says that the average time for their group of papers took 9 months…[9 months is good, no?]

Anyway, for the time being lets focus on…”now acceptable for publication” 🙂

 

Splicing based body-temperature thermometer

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This work from the Lab of RNA Biochemistry at the Freie University Berlin shows just how sensitive splicing is to small changes in body temperature.

They looked at alternative splicing (AS) of U2af26 across a physiologically relevant temperature range (35-40oC). [U2af26 is a component of the essential splicing factor U2af (U2 auxiliary factor) where it can substitute for U2af35 in heterodimers with U2af65]

The authors show that U2af26 exon 6/7 skipping showed a very nice linear correlation with the temperature (see their figure below), suggesting that AS is able to react in a thermometer like way to read body temperature changes.

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The paper goes on to show an involvement for SR proteins in temperature-regulated U2af26 AS, primarily via modulation of the phosphorylation state of SRs. The authors speculate that there will be a physiological role for temperature-controlled AS in other phenomena, such as hypothermia and fever.

 

 

Drawing Splicing 1

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Back to thinking about how to explain alternative splicing in an easy, graphical or pictorial way.

Here’s an attempt at sketching plant cells under a microscope. Grid like arrangement of cells, with chloroplasts (photosynthesis organelles) as greenish circles, and the cell nucleus as dark circles/blobs.

Not entirely sure where this is going…maybe a cartoon. Still hope to include Pandas somewhere along the line…