Clocked: local SNPs in global pops

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R4RNA arc diagrams (top) for predicted secondary structure comparison of the (upper) G/G/U/G/C and (lower) A/U/G/C/A haplotypes, with (bottom) SNPs aligned along the LHY 5’UTR region (exons; boxes and introns; horizontal lines). [from Fig 1 (b and c) of James, Sullivan and Nimmo, PCE, 2018]
Our study on the correlation between ‘natural variation’ in a clock gene sequence with bioclimatic parameters is out now as OpenAccess in the journal Plant, Cell & Environment.

The paper is called ‘Global spatial analysis of Arabidopsis natural variants implicates 5′UTR splicing of LATE ELONGATED HYPOCOTYL in responses to temperature

The starting point for this work was the idea that the 5’UTR of the core clock gene LATE ELONGATED HYPOCOTYL, also known as LHY, could function as a thermosensor given that we previously saw temperature sensitive alternative splicing of LHY.

We tested our theory using the 1001 genomes resource, a whole-genome sequence database for at least 1001 strains of the reference plant, Arabidopsis thaliana. Arabidopsis is native to Europe, but can now be found in the United States, North Africa and temperate Asia. We examined subtle differences, or polymorphisms, in the DNA sequences of >1001 accessions. These are often referred to as single nucleotide polymorphisms (SNPs). We found that different strains tended to ‘shake out’ as particular ordered assemblies of the SNPs, called haplotypes [for example, in the picture above the G/G/U/G/C haplotype is compared to the A/U/G/C/A haplotype] .

We were interested to see if the distinct haplotypes aligned with particular features of where these plants were growing – maybe the haplotypes grouped according to latitude, longitude, or altitude? Or would they group according to climate, such as temperature? seasonality? or even rainfall? For this we made use of the WorlClim database – a free public resource offering global climate data for ecological modelling.

The key findings were that:

  1. One of the haplotypes has hallmarks of being a signature of ‘relict’ accessions (survivors of the last ice-age and the subsequent expansion of new populations). This version is the most distinct in the respect that, worldwide, the accessions bearing this haplotype are found in regions of low rainfall. They are also associated with the highest elevations with low mean annual temperatures and a wider range of maximum–minimum temperatures
  2. Two of the remaining three haplotypes seem to associate with milder annual mean temperatures and lower altitude and wetter habitats
  3. The fourth haplotype, seems to be a low temperature specialist. This haplotype is commonly found in the mountainous Pyrenees region of northern Spain and is prominent at the limit of Arabidopsis growth in northern Sweden
  4. By measuring the extent of LHY spliced upon cooling in representative strains from two haplotypes we established that haplotype does indeed affect the splicing of LHY transcripts in response to cooling
  5. We propose that the LHY haplotypes possess distinct 5′UTR pre‐mRNA folding thermodynamics and/or specific biological stabilities based around the binding of trans‐acting RNA splicing factors

There is much interest in identifying plant thermometers and how they have evolved to cope with new temperature environments. Our new work shows that subtle differences in the DNA sequence of global populations of Arabidopsis plants influences the scalable splicing sensitivity of the mRNA for this central clock component, thereby finely tuning the clock to specific temperature environments.

We anticipate that these findings will be of interest and relevant to crop breeding programs that aim to produce stable food crops in the face of changing climate. 

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Free the roots

 

Been growin’ the Arabidopsis seedlings in Eppendorf tubes filled with agar media. The young roots do what they do best and grow downwards through the agar media (this is called ‘gravitropism‘). The race is on though….if we don’t help them now they will be trapped in a plastic hell of diminishing nutrients.

That’s when we come to the rescue and clip the bottom of the agar tubes so that the growing roots can escape to freedom. This part of the process of growing Arabidopsis hydroponically is between the two earlier posts ‘Seed spotting‘ + ‘Green shoots emerging‘ and this later one: ‘Looking for roots‘. We use a tube cutter from VWR.

 

Once all the tubes are clipped, a puddle of hydroponic ‘root juice’ is added to the tray. This is double the strength of the hydroponic media used to make the agar in the tubes. We do this to help tease the roots down and out of the tube…it’s their reward for their escape to freedom.

Got a good crop this time…hopefully that’ll mean we can do lots of interesting experiments.

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C’est Chic

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Caught a piece on Nile Roger’s new ‘Chic’ album on GMTV this morning…what a inspiration!

He explained that his new album is called ‘It’s About Time

Wow…talk about great minds thinking alike! Our blog, and research is also ‘about’…’time’.

Only we are all about circadian clocks and how plants tell the time.

Just how do plants anticipate the light at dawn for photosynthesis, and how do they time the breakdown of the starch that they’ve made during the day for surviving the dark of the night? How do they do this?

It’s their circadian clocks – molecular DNA and protein ‘cogs’ that link together to make a molecular timer. They just keep ticking – even during the night – maybe it even allows plants to ‘stay up all night to get lucky!’

Pretty funky, no?

We have circadian clocks too. Nile Rogers is an international jet set superstar. I’m sure he’ll feel his clock ‘squeal’ at times with all the globe trotting and jet lag. That’s when the internal clock becomes de-synchronised with its new environment, and it takes a day or two to readjust. Understanding circadian biology is a really ‘cool’ area of research just now. You only have to look at last years Nobel Prize for Physiology or Medicine – it went to circadian clock research!

All this funk makes me want to get down to our research song! Check it out here:

https://abouttimeresearch.com/2015/05/22/its-nice-2-alt-splice/

Good times………

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Feeling Fly

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This fly paper caught my eye. It examines how Drosophila monitors daily temperature changes via network of circadian clock regulated neurons. It seems the fly continually integrates temperature informations in order to coordinate sleep and activity patterns.

The work shows that nodes within the circadian network are sensitive to brief changes in temperature, and show that particular neurons are inhibited by heating and excited by cooling. It seems also that light and temperature are processed in distinct ways in the clock neutron network.

Interested to see the use of a fluorescent protein tool called CaMPARI that photo-converts from green to red in proportion to Ca2+ levels –  could this be used in plant work? Would require both light (photo-activation) and temperature manipulation…

The kinetics of temperature response was monitored by measuring intracellular Ca2+ concentrations using a calcium sensor called GCaMP6m and showed that particular neurons showed increases in intracellular calcium during cooling and decreases during heating.

The authors state that their findings reveal that the circadian network transduces brief and transient temperature changes and prolonged increases in temperature in distinct ways.

Thermoreceptors are found in structures in the antennae, called the aristae. Each arista contains both cold-sensitive and heat-sensitive cells. From their figure (below), they found that the responses to cooling and heating were attenuated when the aristae was removed.

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This work is interesting to use since we are trying to understand how plants respond to everyday changes in temperature – both short-term (daily fluctuations) and long term (seasonal) changes in temperature.

Beer o’clock

After a long and difficult day at work, some of us like to treat ourselves to a nice beer. “The clock” by Eden Mill brewery has taken the ‘beer o’clock’ expression almost word for word.

We are now wondering, is this alcohol-seeking behaviour at specific times of the day controlled by our circadian clock?

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Photo by Prof John Brown.

Note: we must also remind you that chronic drinking can disrupt circadian rhythms.

EBRS/WCC Manchester Conference

EBRS

I’ve been in Manchester this week attending the joint European Biological Rhythms Society (EBRS) and World Congress of Chronobiology (WCC). I presented a short communication entitled ‘An hnRNP isoform switch links temperature perception to regulation of the Arabidopsis circadian clock’. It was a wide and varied programme – although plant-clock research was thin on the ground (I was one of only two plant-clock presentations).

The highlight for me though was Michael Menaker’s plenary talk – the ‘grandfather’ of chronobiology research gave an overview of his journey through chronobiology research from when he was a graduate student in the USA  the 1950’s. It was interested to hear him say how easy he felt it was to get funding back then – “…you would need to be a real idiot not to get a grant funded back then” and how the money sloshing about then was largely due to the space race. Another nugget of insight was “If you haven’t done any failed experiments, you haven’t tried hard enough”

Another quote that stuck with me was from Ueli Schibler’s Plenary lecture – the quote is, I think, attributed to Bertrand Russell – Mathematician/Philosopher/Writer and Nobel Laureate for Literature in 1950 – when he said “One of the symptoms of an approaching nervous breakdown is the belief that one’s work is terribly important”.

Welcome

Bower Building chimney, University of Glasgow - view from just outside the lab
Bower Building chimney, University of Glasgow – view from just outside the lab

I suppose ‘It’s About Time’ we actually posted something!

It’s taken us a while but hopefully this is the start of an interesting way to disseminate what we get up to in our labs and provide a sort of diary of our activities.

During the establishment of this blog we’ve talked a lot about how time and seasons seemed to be a huge influence on artists and songwriters (I’ve been introduced to the delights of Fairport Convention & The Incredible String Band, for example) and I’m sure that this will be a subject of future posts – Time and the passage of Time sure seems to get under our skins, no?

Anyway, welcome to our blog! We are chronobiologists and we like splicing!