Ooops! The human genome does not exist! Part III: a non-Platonic type specimen?

In the last couple of posts (here and here) we have been discussing the idea of 'type specimens', as that idea relates to genome sequences representative, in some way, of a species.  Stuffed robins and dodos seem to raise few scientific questions about the species they represent as type specimens, even though we know that each species varies.

Of course, we're engaging in a bit of tongue in cheek poetic license here, because there are lots of ways to view the notion of a type specimen (see Wikipedia on this, e.g.), and of course, every biologist knows that species vary.  The point here is that it is one or a few examples chosen in some way to represent a species, assuming adequate evidence that a type specimen does represent a proper species (such discussions are off-topic here).  People in genetics should be clear about what a reference sequence, an analogy to a morphological type specimen, is.  Not all of them are.

In the genomic age we now have whole genome sequences from one or more representative individuals, as DNA sequence type specimens of 'the' genome for that species.  Of prime interest of course is 'the' human genome sequence (HG), and we've pointed out that this must be viewed not as a real sequence carried by any individual, but as a sort of Platonic ideal: a reference or road map to the sequences we carry around but none of which is identical to--regardless of whether it does or ever did exist in any individual.  The latter is the central point.

Dodo

The reference sequence keeps changing as errors are found or problematic areas done again, to be clearer or sure about the sequence, yet it seems not to lose its status as 'the' reference. And now we have many sequences from different species--soon to be thousands for humans alone.  Since each differs, many in major ways but all in millions of minor ways, from any edition of the HG reference, we wonder if a single reference (or reference set) is the best way to characterize 'our' genome.  Since evolution works via populations, why not use a population-based way to represent our genome?

We suggested yesterday the idea of using a visual 'gene logo' to represent the variation found in existing sequences on file, augmenting that reference as more data become available. We show the example again here.  This is a 'population type' rather than a type specimen.  It would be 'the' referene instead of a single sequence, not as a supplement.  It shows the most common nucleotides as well as the variation, in every position along the genome.

This is not  a Platonic ideal, because it does not represent a single anchor sequence, but instead portrays genomic variation as it exists in available data--much more natural than a single reference and since genomes got here by evolution, and evolution is about variation,  a biologically more natural way to represent genomes.  There are problems, for example, that each instance of the HG has bits and pieces added, missing or rearranged, so not all the variation is in nucleotides in specific positions along the genome.  But ways could be found (there are, for example, graphical ways to show where segments of the genome have moved to and from).  And of course the underlying data are downloadable.  Still, could there be a better way?

An evolutionary Platonism?
Plato's idea of things like dogs or chairs or virtues is that there is an actual 'ideal' that really exists somehow but of which all actual ones we see are but imperfect instances.  Each new chair is an instance of 'chair', and one might liken this to being stamped out, never perfectly, by a chair-making machine.  Or perhaps every chair is a kind of real sibling produced by the same (ideal) parent.  In Plato's terms the parent may not even have physical existence.  That might give us an idea of how to handle biological types in a better, more physically real way.  The way is an evolutionary one.

Doll chair, The Children's
Museum of Indianapolis

The only reason we can even use a representative, reference HG sequence is that all humans are similar and the only reason that's true is that we evolved by descent with modification, Darwin's phrase, from common ancestry.  So could we represent a gene not by an arbitrary reference sequence of some person who happens to be in our lab, but in terms of ancestry?

If we look at large numbers of instances of a gene, we can make reasonable guesses as to what the common ancestral sequence was like.  If most people have an A in a given nucleotide position in the gene (even if, say, some have a T or G there), then A was likely the ancestral state of that position.   If we find that chimps also mainly have an A there, we can be even more confident.  Working site by site in this way, we can make an educated guess as to the ancestral sequence.  In fact, we do this all the time, and have routine, reasonably reliable software to do it on a large scale.

So why not use the ancestral sequence--which actually did exist, unlike some of the composite and hence artificial sequences we use today--as our reference?  Then, while nobody has the same exact sequence today, we can landmark what we do carry in reference to that 'ideal', a kind of real Platonism.

In many ways we do this in daily lab work and drawing evolutionary phylogenies.  We routinely use the idea of the common ancestral sequence, called the coalescent because all today's variation seems to 'coalesce' to the single ancestor as we look backward in time.  So the coalescent sequence of a gene could be our type specimen for that gene.

That seems like a sound way to 'Platonize' our genomic reference map without the arbitrary use of a randomly picked sequence today, or a sequence that has no material existence, since the coalescent is 'the' ancestor rather than just 'an' instance of the sequence.  But while this is evolutionary sensible, there are problems.

First, the ancestral gene may have been so different from today's that even if we reconstruct it accurately, using it as our reference would not work well from a functional, or perhaps even a structural point of view.

Second, while we can do this for a gene (even including inserted or deleted elements in the gene), what we need is the contextual arrangement of a whole genome sequence.  That means the sequence surrounding the genes (and that is about 95% of genomes!).  But when that is non-functional, or even when it has gene-regulating sequence elements, these can move around and evolve so fast that we often cannot easily guess their sequence.  Genes coding for proteins, are so highly conserved in sequence (because, evolutionarily, mutations were harmful and removed) that we can find and align them from other species.  But this doesn't work as well for surrounding sequence.

Third, the lack of such conservation in non-functional sequence might not matter so much if it weren't that the common ancestor of many genes existed long before there were humans and in some species often very distant from us--sometimes, even, an early vertebrate or even before that.  So the surrounding sequence can't even be guessed at.  And the ancestral gene sequence may have little interpretable relevance for human biology (or phylogeny).

Fourth, sequences are rearranged over evolutionary time, and the flanking bits aren't the same, so we can't really put together a very meaningful whole genome sequence (this has been attempted for mammals or vertebrates, but is very speculative and approximate, it never existed in an individual, and is essentially worthless as a reference to guide much of current work on genes and their functions).

Fifth, each gene's evolutionary-Platonic ancestral copy existed in a different individual, the ancestors for different genes living far apart and at very different times in history.  The animal (not even a person!) with 'the' original beta-globin gene would have been countless centuries and kilometers different from whoever had the ancestral blue-light receptor gene, and so on.  Unlike Platonic chairs, there may have been single ancestral genes, but not a single ancestral genome. 

So, while evolution is a better framework for discussing ideals than Plato's, because ancestors did actually exist, using ancestry to produce reference sequences--genome 'type specimens'--doesn't seem to lead to an obvious strategy.  This is strange, since a random instance of a genome sequence works as a reasonable reference because we're all so closely related--the same evolutionary reason that doesn't seem to work very well.

Our own view is that we should try some kind of variation-including method, such as the 'logo' approach above and that we discussed in Part II of this series, to represent our genomic 'type'.  It's just like using a single stuffed robin, or a single Neanderthal skeleton: imperfect, easily misleading, not actually 'representative' as a representative......but taking variation into account, perhaps the best we can do.

A few thoughts and updates about these topics will conclude this series, in Parts IV and V next week.

Ooops! The human genome does not exist! Part II. What kind of reference is a reference?

We posted yesterday about the nature of type specimens, one individual (or sometimes a few more) chosen by scientists to represent an entire species.  How that choosing was done, how the type specimens were sampled, or what they actually represent is variable but always a bit (or more than a bit) arbitrary.

Nessie

When the age of genomics arrived, and we had large-scale sequencing, the Human Genome Project gave us something similar, 'the' Human Genome sequence (HG).  This, like previous type specimens--the stuffed carcases shipped back home by globe-trotting naturalists largely in the past 2 or so centuries--is an arbitrary instance of something we believe exists elsewhere in profusion.  But while the stuffed robin or Loch Ness Monster on display was an actual robin--or a grainy picture of an actual LNM--even this degree of specificity doesn't apply to the HG:  it has generally been said that it was not derived from a single individual, but 8 or so, whose identity or identities or ethnic origin etc.  are confidential.  In addition, it is under revision as errors are found and so on.

That means if you do a study today, for example, looking at some human genome region in a sample of people, you'll have to identify that region in terms of hg19, the current edition of the sequence. But if you want to compare your work to past work, or if those following up on your work want to compare to yours, they'll have to go back (or forward) to hg19.  Yesterday we noted that the starting position for the human beta-globin gene has shifted about 43,000 nucleotides between the current and previous versions of the genome.  This is not just due to error correction, but would happen if you compared any two instances of a human genome.

Thus not only do the sequence details of any given gene vary, but so does its location. The same is true for any other functional or even nonfunctional aspect of the genome.  The beta-globin or any other gene no more exists in reality than does 'the' HG itself.  What about other details? Just as every robin differs (we don't know about Nessies), so every instance of the HG differs.

Albino Robin, photo from www.seed-solutions.com

How many chromosomes do we have?  22 (two copies of each) plus X and Y?  Well not everybody has that number.  People with Down syndrome have 3 copies of chromosome 21.  Well, you may say that's abnormal, but are they not human?   In fact, every instance of a human genome varies in the number and location of substantial chunks of DNA (called 'copy number variation'), and there are shorter bits that vary in their number of copies in millions of different places across the genome.  There are tens or hundreds of defunct genes, different sets of them in each 'normal' person.  Further, of course, what is 'normal' is a largely arbitrary notion, based on what is average (the 'norm') or some range that we think of as 'normal' in some vague and subjective way.

We've discussed when or how we decide that your behavior is not 'normal' (e.g. here), and how disease is sometimes an arbitrary classification that we choose to make.  Are people 'abnormally' tall or short not human?  Is that a 'disease'? 

Somehow we need to decide how to categorize species--their traits and their DNA.  What would be a better way to deal with a Platonic ideal, a single kind of reference genomic sequence--an ideal  that seems to make sense, while at the same time there is no actual instance of that ideal?

Reference sets?
Rather than a continually revised type sequence, one possibility is to use some set of DNA sequences to determine what 'the' human genome is like.  But then what would be included in that set, and how many different sequences?  If we want a road map, strictly as a reference for comparing sequences, how would a set be different from a single type specimen?


If we had a set of sequences, they would somehow capture both the typical elements and their variability, and these are vital to understanding both gene function and evolution.  One way might be to use 'gene logos', such as in this figure:

Here, each position along a sequence is represented by a visual, color-coded, letter whose size is proportional to the the fraction of the  sample that contains an A, C, G, or T. So in the first position, every sequence had a C, but in position 7, G predominates.  The height of the letters sum to 1 (meaning 100%, unlike in this particular image).

This would provide a quantitative 'stereotype' of the human genome sequence that is not a rigid Platonic ideal, but would still be useful.  A given sequence, like the beta-globin gene, varies but a computer search could easily find it in any whole genome sequence (a search and compare method called BLAST does just that).

We could also include missing nucleotides by having a proportionally sized N (for None) in the location could be added.  Major copy variation or rearrangements would be harder to include, but probably ways could be found.

With computer technology, even a digest of all available sequences in a species could be stored in this way.  Since individuals will always die and be born, at any given time this would be a 'type species' rather than type or reference specimen, but that would perhaps be a much better way to think of things.

Then, it would not be necessary to use the concept of 'the human genome sequence' as a somewhat misleading guide--where we use something that doesn't exist to try to understand what does exist.  In fact, something like this approach is already available, though not standard, and will be discussed in Part IV of this series.

How we would do the same with robins (or Loch Ness monsters) is less clear.

Ooops! The human genome does not exist! Part I. The notion of a type specimen

It's sultry summer, and of course the science hype machine motors on unabated, but we want to ignore it briefly and relax at least a little bit before Penn State resumes it's serious academic activities--that is, football season.  Still, while you're basking in the sun getting your tan (and skin cancer), along with your mint juleps (and undoubtedly some other ugly diseases), here's something to think about:

The human genome doesn't exist

René Magritte: This is not a pipe.
It's a painting.  But a painting of 'a'
 pipe--but not  'the' pipe, which doesn't
 actually exist in the real world.  Or does it?

Despite many claims to the contrary, that The Human Genome project sequenced the human genome and thus set in motion the most exciting era of fundamental new scientific discovery since Galileo, it has turned out that the HG doesn't exist, after all.  It no more exists than does 'the chair' or 'the dog' as Plato once asserted.  He said we have dogs and chairs but they are only imperfect instances of the real true dog and chair.

Now what everybody raves on about, the HG, is like that.  It is not from one person, not even one copy from one person's two. It's not clear whether this is even from one person or from several donors, or if from one, who that person is in terms of his origins (his, because there's a Y chromosome in the sequence). 

In any case, it's not a 'normal' genome, because while we believe the sequenced individual(s) was/were healthy at the time they bled for the cause, they won't be healthy forever.  Since we are told every day by the press that every disease without exception must be genetic, and hence we should GWAS it endlessly,  the donors will eventually become genetically abnormal.

Worse, 'the' genome keeps changing!  We are now in version 19, released in 2009. It's either more details from the same donors, or now has bits that couldn't be sequenced easily from those donors' DNA and so was sequenced from additional donors (we don't know which is the case).  There will be future revisions.  Not only that, but it is a 'haploid' sequence, with only a single-nucleotide per position reference, whereas any human has two copies (except for X and Y chromosomes).  Yet in any one person 1% or more of sites actually vary.

This is not Van Gogh's
kitchen chair

Now, this means clearly that nobody actually has  'the' currently posted human genome sequence any more than your kitchen chair is 'the' chair.  Of course, what some biologists who are more savvy than most people who talk about it know is that the HG is a reference sequence.  Nobody has and nobody has ever had, this incomplete reference HG. All of us have some variants of that sequence.  It is wrong, but standard, to refer to what we each carry as 'copies' of the HG.  They are not copies!  There is no copy of the HG!  No one person ever had that sequence, so no person could copy (replicate) it to transmit to its offspring.

Probably, we should use a term like 'instance' rather than copy, in the same sense that each chair is an instance of the concept of chair, even if no Platonic actual ideal chair exists, except as a reference concept in our minds.

What is good for the goose is good for the gander.  If the HG doesn't actually exist, neither does a given gene say 'the' beta-globin gene, which is expressed in red blood cells and of which some variants are involved in anemia (like sickle cell anemia).  Here is hg19's version of a very small part of that sequence (click to see details):

Any one instance of this in an actual person, like you, may or may not have this exact sequence.  But unless it's been deleted from your genome, you'll have something very similar, not because it is a copy of some Platonic idea, but because they are descendant copies handed down through the generations since we shared a common ancestor, and that--evolution--is the crucial difference.  It's why we should be careful about treating a reference as being real, or how we relate the existing instances of the 'same' (homologous) sequence.

We use 'the' HG sequence as a way of referring to a standardized set of nucleotide locations along the chromosomes in the individuals who were sequenced.  Each of us varies from that sequence in millions of places (in each copy that we carry). To find similar functional elements--genes and so on--in my instances of the human genome, we use 'the' HG as a kind of map.  If I had your DNA sequence, I could identify which is 'the' beta-globin gene by its similarity to that in the figure, even if you didn't have the exact same sequence.

This is very useful, but it's important to be aware that not only does nobody have the same sequence, but even the structural details--the locations of the 'same' elements in you and us and the donors of 'the' HG--differ among people.  So the coordinates (the number at the top line) will not be the same.  For example, the beta-globin gene starts at chromosome 11 position 5,246, 696 in draft hg19, but at 5,203, 400 in draft hg18, a difference of around 43,000 nucleotides!  Where did they come from?  Which draft should we believe--if any?  What will hg20 say? What is 'the' HG?

Type specimens

American Robin

Think about this in another context.  For centuries we have had the concept of a type specimen.  There is 'the' robin, 'the' monarch butterfly (or even, perhaps, 'the' Loch Ness monster??).  'The' Neanderthal hominid is an arbitrary individual whose luck it was to die and not rot, so we could discover him around 30,000 years later in Germany.  The lucky old sod gets to represent all of his contemporaries and ancestors and descendants for hundreds of thousands of years.  And 'the' Neanderthal DNA sequence is partial, a composite of several individuals who lived hundreds of kilometers and thousands of years apart.  Think about that when you hear people pronouncing on how 'the' Neanderthal lived, or what color hair 'it' had, or whether it had religion!

Robin you'll see in the UK (image: RSPB)

The international organization of systematists--biologists who are concerned with cataloging, characterizing, and name species and their relationships--has decided what counts as a reference specimen of an organism, in the same way we decide what counts as a reference specimen of the genome of a species.  Indeed, and interestingly, the genome of 'the' mouse or 'the' horse, is likely rarely if ever to be from the same official physical type specimen on display at some authorized museum.  Thus some poor (former) robin sits rigidly on some display branch for all to see, representing all robin-hood individuals it never saw or dreamt of.

This leads to interesting questions about how we treat our subjects, biology and evolution.  Are there alternatives to type specimens?  So, while you nurse your mint julep in suspense, think about this--the whole concept of a reference specimen, be it genetic or physical, because tomorrow we'll give a least a bit of thought to this question.