And my lab studies transposable elements.

The title of these two presentations are The Dynamic Genome.

In the first talk I introduced transposable elements by describing their discovery by Barbara McClintock,

how they move, and how that discovery over the years was recognized as a major revolution in biology

as it became appreciated that transposable elements

are the major component of most of the genomes of higher eukaryotes.

In this talk I am going to go into detail about how my lab

studies the evolutionary impact of transposable elements on genomes.

And how we develop strategies to identify elements

that have an impact on transposable elements.

I have divided this talk into three parts.

In the first part I talk about the transition from genetic approaches to genomic approaches

in order to identify elements that in fact impact genome evolution.

The elements that were discovered in my lab are called MITEs,

and I will tell you about that discovery in the second part of this talk.

And in the final part of the talk I will tell you about how MITEs

are able to increase their copy number in the genome without harming the host significantly.

So to review the first part, we talked about the genetic analysis of transposable elements,

how genetic analysis led to the discovery of transposable elements,

and I used this spotted corn kernel as an example to tell you really how powerful the genetic analysis was.

So when you see a spotted corn kernel like this,

we know, the geneticist knows, that the reason that kernel is spotted

is because there are active transposable elements.

There are... that is the spots reflect the movement of transposable elements.

The other thing the genetics tells us is exactly where in the genome that active transposable element is.

So for example, here when we are looking at spotted corn kernels,

we know that there is an active transposable element, in other words, one that is capable of moving

in a gene responsible for kernel pigmentation.

The other thing that the genetics tells us is the type of element that's there.

And I described at the beginning of the first talk the difference between autonomous elements,

that is, ones that encode transposase, and non-autonomous elements.

Those are the elements that don't make transposase

but are able to move if there is an autonomous element in the genome.

McClintock and others were able to deduce this just by looking at the behavior

of transposable elements in crosses.

Now that is the good news about the genetic analysis.

Unfortunately the genetic analysis is limited in its scope.

And that is that by its very nature genetics depends on the analysis of mutant alleles,

and so the transposable elements that were being studied were the ones causing mutations.

They were mutagenic elements.

Now because these elements cause mutations, there aren't many copies of them in the genome.

So, I mentioned in the first talk that genomes are up to 50-80%

of the genome sequence is derived from transposable elements.

However, the elements that cause mutations are not those elements.

And you can understand that an element that causes mutation is eventually,

if its copy number increases too high, will kill the host.

So these are.... these are a special class of transposable elements that cause mutations.

And as such these elements really have a minimal impact on genome evolution.

They're bad. They are really bad.

So McClintock as I also described in the first talk,

not only discovered transposable elements,

but she hypothesized that they were also tools that diversify organisms.

And to review, she hypothesized that transposable elements that are in the genome

do not move around frequently,

that there are conditions, such as changes in climate for example,

that could activate transposable elements.

that this activation would generate genetic diversity in the population

by increasing the frequency of mutation,

and that some of these transposable element mutations may be adaptive.

I will come back to this scenario at the end of the talk

when I show you how the elements that we have identified

in plant genomes fit this scenario very, very nicely.

So to review, I described in the last talk the two general classes of transposable elements in the genome.

The first class, which is called Class 2,

are DNA transposons. These are the elements that were discovered by McClintock genetically.

We know these elements have a typical structure of terminal inverted repeats,

that they encode a single protein necessary for the movement of the element, and that is transposase.

The other class of elements which I am not going to go into extensively in this talk,

and nor did I in the last talk, are called retrotransposons.

These are elements that encode reverse transcriptase,

and they move through an RNA intermediate, again by mechanisms that were described in the last talk.

I also want to re-introduce you to transposable elements families

because we are going to revisit families in this talk.

That transposable element families contain autonomous elements.

That is the elements that encode transposase.

And non-autonomous elements these are elements that don't encode transposase,

but are able to move utilizing the transposase that's encoded by the autonomous element.

Something else that I discussed in the first talk is just how prevalent transposable elements are in genomes.

So this was a human gene where I showed you the exons that are in the gene,

and this is really a pretty typical gene. And what we find is that

in the non-coding regions, there are many, many, many transposable elements,

that some human genes have over a hundred transposable elements in their introns.

Now the element that I am referring to, most of the elements in the human genome are called Alu.

They are a class 2 retrotransposon which are present at an astonishing copy number of over a million copies.

It's almost ten percent of the human genome.

Now one of the things, if we are interested in the evolutionary impact of a particular transposable element,

one question we could ask is, so for example, if we picked out one of these elements,

we could ask what happened when it inserted?

Did it change the expression of the gene?

These are the questions, if we are interested in the evolutionary impact,

these are the questions that we would like to be able to address.

Unfortunately we can't do that with the human elements, most of the humans elements.

And that is that the insertions may have changed gene expression,

but we have no way to address that now. And the reason is

first of all in the human population, virtually all of us, 99.99% of us, have these insertions,

have exactly these insertions. That's because these elements moved millions of years ago.

So what that means is that if we want to know how did the insertion

of a particular element change the expression of a gene, if at all, we are too late.

So what we want to do is identify a group of organisms

where these high copy number elements are actively transposing.

And I am going to talk about that strategy- that's exactly what this talk is about.

So here is our strategy for analyzing the impact of transposable elements on genome evolution.

And this is a figure from the previous talk which is sort of a typical region of a genome, a grass genome,

and this is from the barley genome,

and the blue boxes are genes and the triangles are transposable elements.

So in barley about 85% of the genome is derived from transposable elements.

So the strategy that we would like to do to identify evolutionarily relevant transposons

is to find a species that is in the midst of genome expansion.

So where these high copy number elements are moving, are increasing their copy number.

And we would then go ahead and identify and isolate an active element.

So this is not one of the mutagens that was identified by the geneticists,

but in fact these are the high copy number elements that are now increasing in copy number.

Ok, so we would then ask the question,

how is this element able to increase its copy number so extensively without harming the host?

What are its strategies for success? And success in this case is defined by being able to increase

your copy number without killing or harming the host, and possibly even by benefiting the host in some way.

And we are going to address all of those issues in this talk.

So in the first talk we talked about the discovery of transposable elements in maize.

Well, maize is a member of a larger group of organisms. It's a grass.

These are the most important organisms for human health, for the human diet.

More calories come from members of the grass clade than any other group of organisms on this planet.

We are familiar with maize. The other members of the grass clade is: rice,

which actually is the most important source of human calories,

sorghum, which is also a very important crop plant especially in Africa,

and finally barley. Another member of this family, which I am not showing is wheat.

So what you would notice here, those numbers are the size of the genome.

The maize genome is about the same size as the human genome, at 2500 megabasepairs.

The rice genome is much, much smaller. It is almost ten-fold smaller.

And what is remarkable is that here are these plants that are so incredibly similar,

yet their genomes size differs dramatically, by more than ten-fold.

So these organisms diverged from a common ancestor only about 70 million years ago.

And the main reason for this difference in genome size is this dramatic amplification,

expansion of transposable elements.

And this slide helps explain in part how that can happen.

These organisms, the grasses, in fact have about the same gene number.

They have about 30,000 genes, give or take a few 1000.

And so the genomes of these organisms are largely syntenous. The genes are mostly in the same order.

So in rice you could see, with smallest genome, I've shown three genes there in pink, yellow and blue

that are pretty close together. In maize those genes are further apart.

And in rice those genes, I'm sorry, in barley those genes are even further apart.

So what's happening here is transposable elements,

which are the squares and circles and ellipses in between,

transposable elements are inserting massively between the genes and expanding the genome.

So this is largely responsible for the difference in genome size.

And it's the safe havens that transposable elements can go without harming the host.

So I want to show you a little bit at a higher resolution

to show you what elements are involved.

So I introduced to you before that there were two types of transposable elements.

There were Class 1 elements which are retrotransposons,

and then Class 2 elements which are DNA transposons.

The retrotransposons which are generally these big elements that make RNA copies.

That RNA copy is then made into double stranded DNA.

The double stranded DNA can insert back into the genome.

It almost make copies like a printing press, like an old fashioned mimeograph machine,

which most of you probably never experienced.

So what you see here is that the huge blocks could be hundreds of kb

that separate some genes in the grass genome

are largely retrotransposons that are inserted into each other,` literally driving the genes apart.

So as we say it's almost like genes sitting in a sea of transposable elements.

Now these are not the elements that we are going to talk about today.

Instead we are going to talk about elements that I think are probably more involved in diversifying the genome.

And that is, so what I have done here is I've blown up the area of one gene

and broken it up into its exons and introns.

And I've shown you that sitting in plant genes, much like the Alu elements that are inserted in human genes,

are little elements, little transposable elements called MITEs.

These are DNA transposons. They are non-autonomous elements

and in the next few slides I am going to tell you a little bit more about MITEs

because they were discovered in my lab at least 2 decades ago.

So the way the first MITE was discovered was it was an insertion sitting in a mutant gene.

And this is the work of several people in my lab, especially Tom Bureau and Rita Varagona.

What you see here is a maize gene, and sitting in it is a little DNA transposon,

disrupting the gene, causing a mutation.

When Tom Bureau isolated this transposon, like I said,

this was back in the early 90s, when he isolated the transposon,

he took the sequence and compared it to several other maize genes or plant genes

that had been deposited into databases. This was before you did BLAST searches.

This was in the early 90s when there were some wildtype genes that had been sequenced.

And what he found was that in fact sequences like this little element

were present in many of the other wildtype genes.

And I am showing that here. So what you see is here are some wildtype genes

that were revealed by this computer search.

And so for example, this gene has an insertion in an intron.

So these are normal genes. So these insertions are in non-coding regions.

They are not effecting the expression of the gene.

This one is in the 5' promoter region.

And the one at the end here is in the 3' region.

So he discovered these elements, they are called... MITEs stand

for miniature inverted repeat transposable elements.

What is similar about these elements is their structure.

Their sequence may not be similar and is not similar when you go from organism to organism.