The same, yet different

6 December 2013 by Malcolm Campbell, posted in Biology

With such old counsellors they did advise, and by frequenting sacred groves grew wise.” From On St. James Park by Edmund Waller  (1606-1687)

On first look, it is merely a grove of trees.

It’s a fairly ordinary, nondescript grove at that – a couple of dozen deciduous trees, long and lean, grey barked, with arrowhead-shaped leaves.

They are uneven in age and diameter. A yearling here. Another, maybe a decade old, a few meters away.

Kind of scraggly.

“Nothing to write home about”, as they say.

But the little grove of trees is so much more than that.

It’s a patch of clones.

The small stand is composed of a group of genetically identical individuals. Clones. A botanist would call the stand a genet, and each of the individuals, ramets.

The trees are members of the poplar family – the genus Populus. This particular stand is balsam poplar, Populus balsamifera. In the summer, it perfumes the air with its distinct resinous aroma.

Trees in the poplar family, and those of the related willows, fill the landscape by two means. In the first instance, they create offspring by standard sexual reproduction – pollen fertilises eggs to produce seeds that are dispersed, generation after generation.

Poplars also spread asexually, through what is known as vegetative propagation. Here, new saplings arise from the roots. Alternatively, stems that are snapped off of the main tree take root in the ground where they fall. Either way, the new individuals are genetically identical to the tree from which they arose. They are clones.

Stands of poplar clones pepper the landscape in the boreal zones of the northern hemisphere. One member of the poplar family, trembling aspen, is so effective in reproducing clonally that it creates forests that are patchworks of clones. The patchwork is clearly visible in the autumn, especially from the air, as each patch has a distinctive appearance when the leaves change colour.

Clonal propagation is a remarkably effective strategy for colonising a landscape. In fact, some clonal stands attain notable sizes, occupying a dozen or more hectares each. One such stand is famously large, covering over 40 hectares. This genet, known as “Pando”  (Latin for “I spread”), has been described as both one of the oldest and the heaviest organism on the planet.

Clonal reproduction may have its advantages in spreading across a landscape, but it carries a significant risk. As all of individuals within the clonal stand are genetically identical, they are all susceptible to the same environmental insults. Whether it be pests or pathogens, cold weather or drought, if one individual within the genet is susceptible, it is likely that everyone is. Given this, why are poplar genets so successful? Why don’t clonal stands of poplar succumb to one malady or another before attaining such large sizes, and persisting for so long?

Of course, the secret to a poplar genet’s success resides within its DNA, its genome. The poplar genome is contained on 19 chromosomes. Like all organisms, its genome comprises pairs of the “letters”, the nucleotide bases – A, C, G, and T. Where the human genome has 3 billion of these base pairs, the poplar genome has only approximately 500 million. These 500 million bases contain more than 45000 pieces of code, genes, which provide the instructions for making a poplar tree.

The genome, and its component genes, have frequently been described as a “blueprint”. This is a bad analogy.

The genome, or a gene, is not a blueprint. A blueprint is a two dimensional representation of a three dimensional structure. By contrast, a gene is a linear code, like letters affixed to a piece of string. A gene is read in a linear fashion, using the quaternary letters of the code, in a manner more analogous to reading the binary code of a computer algorithm, or the alphabet used to write a recipe.

Each gene provides the instructions to make something, in a particular place, at a particular time. Gene after gene is attached together in a long string of different instructions. Each of these long strings is a chromosome.

The instructions encoded along the length of the chromosome must be read, and translated into a piece of molecular machinery that fulfils a function within the cell. That molecular machinery is generally a protein – a macromolecule that can catalyse a chemical reaction, or build a cellular structure.

The challenge of converting the DNA code of the genome into pieces of cellular machinery is that the instructions it contains are held like non-circulating books in a library. The genome resides in the nucleus, and its instructions must be read there.

A specialised group of proteins function to read the code within the nucleus, and transcribe it into a form, a transcript, that can be translated into a piece of molecular machinery elsewhere in the cell. The proteins that fulfil this transcribing function are like monks of yore transcribing segments of ancient text. Like the monks, these proteins are informed by external conditions to enter the library, the nucleus, to transcribe specific parts of the text. Just as different monks specialised on transcribing particular segments of text, so to do the proteins, these “transcription factors”, home in on particular regions of the genome, transcribing it in response to external cues.

The extent to which a gene is transcribed will ultimately shape the extent to which its instructions are used in the cell. Depending on the conditions – the time, the tissue, the prevailing environment – some genes will not be transcribed at all, while others will be transcribed abundantly, and still others at levels in between. In poplar, with its 45000 genes, the extent to which all the genes are transcribed at any given point in time will shape the capacity of the tree to survive in the environment – its ability to undertake photosynthesis, to transport water throughout the plant, to make a new leaf, to grow new roots, to mount a defence against a pathogen.

As their genetic code is identical, all members of a clonal stand of poplar theoretically share the same capacity of their genes to be transcribed at a particular point in time, in response to a particular set of environmental conditions. It is for this reason that clones share the same risk when adverse conditions arise – they all are saddled with identical sets of instructions of how to respond to adversity.

Sherosha Raj and her colleagues demonstrated the extent to which gene transcription in clones responds in an identical fashion in response to adverse conditions. Raj and colleagues took advantage of the fact that some poplar clones are very popular for forestry practices. These clones have been planted in multiple locations, sometimes at quite a distance from each other.

Raj and her colleagues used three different varieties of poplar. Each different variety was obtained from two different locations in the country – literally hundreds of kilometres apart – from nurseries with very different meteorological conditions.

Using stem segments collected from genetically identically clones grown at the two different locations, Raj and colleagues established new ramets in one site – a common garden. The common garden was a growth room with controlled growing conditions, so ramets established from stem segments obtained from one part of the country could be grown side-by-side those established from stem segments from another part of the country – all in one location, under identical conditions.

After approximately six weeks, once the ramets were around a meter tall, Raj and colleagues subjected one half of the ramets to drought and continued to water the other half. So, for each variety tested, for each geographical origin, half of the plants were subjected to drought stress while the others were not. All of this was done in the same growth room, and the treatment was done until all of the plants experienced the same extent of physiological stress. At this point, Raj and colleagues examined the extent to which each of 45000 genes were transcribed in the drought-treated plants relative to those that still received water. They did this at two times of the day – before dawn, and midday.

Raj and colleagues were interested in testing the hypothesis that a clone always behaves like a clone. That is, regardless of whether the clone was grown in one part of the country versus another part of the country, once the genetically identical individuals were grown together, their genes would respond in the same manner in response to drought stress. After all, if they have identical genes, shouldn’t clones always behave the same way?

For one variety, Raj and colleagues discovered exactly what was predicted. Many genes showed differences in transcript abundance in response to drought, but the extent of these changes was the same regardless of whether the clone had been sourced from one geographical region versus another. For another variety, a very small number of genes responded to drought differently in clones from one geographical region versus the other. For the third variety, there were remarkable differences, hundreds of genes, that responded to the drought stimulus differently in clones obtained from one region versus the other. Drought-induced changes in transcript abundance were consistent within a geographical region this latter variety, but changes in transcript abundance in response to drought were markedly different when the two regions were compared.

Raj and colleagues were surprised by the findings that genetically identical individuals could respond differently to the same stimulus, even when everything was done to ensure that the clones received identical treatment. What could be going on? Shouldn’t clones behave like clones – identically?

In the past two decades, a picture has emerged that helps explain how clones might respond differently to the same stimulus.

Experiences during one’s lifetime can leave an indelible “mark” on the genome, changing how genes are subsequently transcribed. We have an incomplete picture of how these marks are made. We know that it involves a change to the chemistry of the genome, but not a mutation of the DNA sequence.  As such, these marks are said to be epigenetic – they fall outside of heritable genetic mutations. What we don’t know is how experience, environmental conditions during one’s lifetime, precisely determine where these marks are laid down in the genome.

Raj and colleagues were able to show that clones that behaved identically had indistinguishable quantitative changes in the epigenetic chemistry of the genome. By contrast, they found that there were marked differences in the quantity of epigenetic chemical marks between the genomes of the clones that responded differently to the drought stimulus. While they are currently determining the precise genes that are impacted by these changes in chemistry, the evidence suggests that clones can respond differently to a stimulus if their past experience has induced an epigenetic change in their genome. This is not an entirely surprising find, but it suggests how clones could persist even though they are genetically identical.

Epigenetic mofications offer a remarkable way to create a form of genetic diversity even when diversity is low at the level of the DNA code itself. Like us, poplar trees inherit one copy of every gene from their paternal parent, and one from their maternal parent. These gene copies frequently vary in the details of the code, such that one copy may be transcribed more than the other, or the molecular machinery derived from one copy is more effective than the other.

Epigenetic modification can influence the extent to which maternal and paternal copies of genes are used. In some instances the paternal copy may be silenced – prevented from being transcribed. In other instances the maternal copy may be silenced, and, in yet other instances, either both or neither may be silenced. All told, epigenetic modifications can generate four possible ways of expressing the DNA information. And even this is an oversimplification – such modifications may vary from cell to cell, from one tissue to another, at one point in time versus another. As such, epigenetic modification provides a plasticity in the transcription of the genome, a way of nuancing the expression of the genome, that creates great flexibility for even clonal organisms. If ramets within a poplar genet have different epigenetic modifications relative to others, it is almost as though the ramets are siblings, not clones. Consequently, they will be less susceptible to the risks of leading a clonal existence.

But there’s an important consideration here.

Despite the wonderful plasticity in gene transcription provided by epigenetic modification, it is the code of the genes themselves that still ultimately determines the fate of the individual and its progeny. Epigenetic modification and gene transcription are not masters of inheritance, they are captive to it. At the end of the day, it will be the code that is passed from one generation to the next. The code will determine the extent to which genes are transcribed, and the code will shape its susceptibility to epigenetic modification. Epigenetic modification and gene expression don’t usurp the process of inheritance and evolution, they are mechanisms that are employed by them.

Epigenetic modification enables the clonal poplar stand to have the flexibility to live out one lifetime, but ultimately, mutation and sex will rule the day.  Indeed, evidence suggests that clonal poplar stands participate in sexual reproduction with relatively high frequency, despite their success as genets.

The poplar grove shows us that plasticity may help us contend with adversity through our lifetimes, but, ultimately, it’s the influence of a more consistent set of codes that make us who we are.

Images: All photographs by Malcolm M. Campbell.


DeWoody, J, Rowe CA, Hipkins VD, & Mock KE (2008) “Pando” lives: molecular genetic evidence of a giant aspen clone in Central Utah. Western North American Naturalist 68: 493-497

Mock KE, Rowe CA, Hooten MB, Dewoody J, & Hipkins VD (2008) Clonal dynamics in western North American aspen (Populus tremuloides). Molecular Ecology 17: 4827-4844

Raj S, et al. (2011) Clone history shapes Populus drought responses. Proceedings of the National Academy of Sciences 108: 12521-12526

Tuskan GA, et al. (2006) The genome of black cottonwood, Populus trichocarpa (Torr. & Gray). Science 313: 1596-1604


2 Responses to “The same, yet different”

  1. Jack Schultz Reply | Permalink

    There's another contributor to the divergence in behavior within clonal plants.
    Nice treatment of this topic! There is another side to this story, though.

    Somatic cell mutation generates within-genotypie variation in plants. The entire plant is generated by a very few stem cells (meristems) at growing points (and a few other places). Mutations occur as those stem cells divide, and they divide a lot to generate an entire tree. So the branches on a single tree - and ramets in a clone - can look and behave differently and have rather different genotypes. A good example is the phenomenon of 'sports'. A 'sport' is a phenotypic variant within a single plant whose germ cells can produce offspring that breed 'true' to the variant's phenotype. For example, the occasional (spiny) honey locust branch is spineless. This unusual variant can be cut off, planted, and all subsequent parts will be spineless and it will produce spineless offspring. In fact, that's how many horticultural varieties are established. The person who knows most about and promotes this view most effectively is Tom Whitham, at Northern Ariz Univ. Like Tom, many of us know that the parts of individual plants are very variable, and some of that variation is genetic. The genetic differences within an organism, even among adjacent cells, is a topic of growing interest lately. While epigenetic influences are also very important, individuals are not as uniform genetically as we usually think, either. (That's true in animals, too, but not so visible.)

    • Malcolm Campbell Reply | Permalink

      Thank you very much for the great reply, and for highlighting the importance of somaclonal variation in clonal stands of trees, Jack. The importance of somaclonal variation, including point mutations and alterations in ploidy, are discussed in the Mock et al. (2008) paper in the reference list. Somaclonal variation was also explored in the Raj et al. (2011) paper, but not comprehensively. I know that some of Raj's co-authors have now actually sequenced the genomes of the clones from the two locations for the poplar variety that showed the differences in gene transcripts. There is some somaclonal variation there, but none that explains the differences in transcripts that were observed. They are now using bisulfite sequencing to examine differences in the methylated genes in the genomes of the clones in question, to better understand the epigenetic differences that may explain the differences in transcript abundance between the clones. They plan to publish this in the coming year. This said, somaclonal variation of the sort you describe can still be a contributing factor in such differences, and can give rise to dramatic changes, like the "sports" you mention. I wanted to restrict the discussion of changes in clones to epigenetic differences in this particular piece (it was already getting to be rather long!), and will return to somaclonal variation in the future! Thanks though, for bringing this important phenomenon to the fore here. Greatly appreciated!

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