89 232 Eukaryotic Transposons

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23.2 Eukaryotic Transposons
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step, the cointegrate is resolved into two DNA
circles, each of which bears a copy of the transposon. An alternative pathway, not used by Tn3, is
nonreplicative transposition, in which no replication
of the transposon occurs.
Nick donor DNA at arrows
23.2 Eukaryotic Transposons
It would be surprising if bacteria were the only organisms
to harbor transposable elements, especially because these
elements have powerful selective forces on their side. First,
many transposons carry genes that are an advantage to
their hosts. Therefore, their host can multiply at the expense of competing organisms and can multiply the transposons along with the rest of their DNA. Second, even if
transposons are not advantageous to their hosts, they can
replicate themselves within their hosts in a “selfish” way.
Indeed, transposable elements are also present in eukaryotes. In fact, they were first identified in eukaryotes.
+
Double-strand
gap repair
Filling gaps, sealing nicks
Figure 23.7 Nonreplicative transposition. The first two steps are just
like those in replicative transposition, and the structure at the top is the
same as that between steps 2 and 3 in Figure 23.6. Next, however, new
nicks occur at the positions indicated by the arrows. This liberates the
donor plasmid minus the transposon, which remains attached to the
target DNA. Filling gaps and sealing nicks completes the target plasmid
with its new transposon. The free ends of the donor plasmid may or
may not join. In any event, this plasmid has lost its transposon.
in the donor DNA on either side of the transposon. This
releases the gapped donor DNA but leaves the transposon
still bound to the target DNA. The remaining nicks in the
target DNA can be sealed, yielding a recombinant DNA
with the transposon integrated into the target DNA. The
donor DNA has a double-stranded gap, so it may be lost or,
as shown in Figure 23.7, here, the gap may be repaired.
SUMMARY Many transposons contain genes aside
from the ones necessary for transposition. These are
commonly antibiotic resistance genes. For example,
Tn3 contains a gene that confers ampicillin resist ance. Tn3 and its relatives transpose by a two-step
process (replicative transposition). First the transposon replicates and the donor DNA fuses to the
target DNA, forming a cointegrate. In the second
The First Examples of Transposable
Elements: Ds and Ac of Maize
Barbara McClintock discovered the first transposable elements in a study of maize (corn) in the late 1940s. It had
been known for some time that the variegation in color
observed in the kernels of so-called Indian corn was caused
by an unstable mutation. In Figure 23.8a, for example, we
see a kernel that is colored. This color is due to a factor
encoded by the maize C locus. Figure 23.8b shows what
happens when the C gene is mutated; no purple pigment is
made, and the kernel appears almost white. The spotted
kernel in Figure 23.8c shows the results of reversion in
some of the kernel’s cells. Wherever the mutation has reverted, the revertant cell and its progeny will be able to
make pigment, giving rise to a dark spot on the kernel. It is
striking that so many spots occur in this kernel. That means
the mutation is very unstable: It reverts at a rate much
higher than we would expect of an ordinary mutation.
In this case, McClintock discovered that the original
mutation resulted from an insertion of a transposable
element, called Ds for “dissociation,” into the C gene
(Figure 23.9a and b). Another transposable element, Ac for
“activator,” could induce Ds to transpose by a nonreplicative mechanism out of C, causing reversion (Figure 23.9c).
In other words, Ds can transpose, but only with the help of
Ac. Ac on the other hand, is an autonomous transposon.
It can transpose itself and therefore inactivate other genes
without help from other elements.
Now that molecular biological tools are available, we
can isolate and characterize these genetic elements decades
after McClintock found them. Nina Fedoroff and her collaborators obtained the structures of Ac and three different
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Chapter 23 / Transposition
Ds are derived from Ac by deletion. Ds-a is very similar to
Ac, except that a piece of the transposase gene has been
deleted. This explains why Ds is unable to transpose itself.
Ds-b is more severely shortened, retaining only a small
fragment of the transposase gene, and Ds-c retains only the
inverted repeats and transposase-binding subterminal repetitive regions in common with Ac. These inverted repeats
and transposase-binding sequences are all that Ds-c needs
to be a target for transposition directed by Ac.
It is interesting that the first pea gene described by
Mendel himself (R or r), which governs round versus wrinkled seeds, seems to involve a transposable element. We
now know that the R locus encodes an enzyme (starch
branching enzyme) that participates in starch metabolism.
The wrinkled phenotype results from a malfunction of this
gene; this mutation is in turn caused by an insertion of an
800-bp piece of DNA that seems to be a member of the
Ac/Ds family.
(a)
(b)
(c)
SUMMARY The variegation in the color of maize
Figure 23.8 Effects of mutations and reversions on maize kernel
color. (a) Wild-type kernel has an active C locus that causes
synthesis of purple pigment. (b) The C locus has mutated, preventing
pigment synthesis, so the kernel is colorless. (c) The spots correspond
to patches of cells in which the mutation in C has reverted, again
allowing pigment synthesis. (Source: F.W. Goro, from Fedoroff, N.,
Transposable genetic elements in maize. Scientific American 86 (June 1984).)
forms of Ds. Ac resembles the bacterial transposons we
have already studied (Figure 23.10). It is about 4500 bp
long, contains a transposase gene, and is bounded by short,
imperfect inverted repeats, and adjacent subterminal repetitive regions that bind transposase. The various forms of
(a)
kernels is caused by multiple reversions of an unstable mutation in the C locus, which is responsible for
the kernel’s color. The mutation and its reversion result from a Ds (dissociation) element, which transposes into the C gene, mutating it, and then
transposes out again, causing it to revert to wildtype. Ds cannot transpose on its own; it must have
help from an autonomous transposon called Ac (for
activator), which supplies the transposase. Ds is an
Ac element with more or less of its middle removed.
All Ds needs in order to be transposed is a pair of
inverted terminal repeats and adjacent short
sequences that the Ac transposase can recognize.
(b)
Ds
C
(c)
Ds
C
Ac
Ds
Ac
C
Ds
Mutant C
Figure 23.9 Transposable elements cause mutations and reversions in maize. (a) A wild-type maize kernel has an uninterrupted, active C
locus (blue) that causes synthesis of purple pigment. (b) A Ds element (red) inserts into C, inactivating it and preventing pigment synthesis. The
kernel is therefore colorless. (c) Ac (green) is present, as well as Ds. This allows Ds to transpose out of C in many cells, giving rise to groups of cells
that make pigment. Such groups of pigmented cells account for the purple spots on the kernel. Of course, Ds must have transposed into C before
it (Ds) became defective, or else it had help from an Ac element.
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Transposase
Ac
Delete
Ds-a
Delete
Ds-b
Replace DNA between inverted terminal repeats
Ds-c
Figure 23.10 Structures of Ac and Ds. Ac contains the transposase gene (purple) and two imperfect inverted terminal repeats (blue), including the
subterminal repetitive regions. Ds-a is missing a 194-bp region from the transposase gene (dashed lines); otherwise, it is almost identical to Ac. Ds-b
is missing a much larger segment of Ac. Ds-c has no similarity to Ac except for the inverted terminal repeats and subterminal repetitive regions.
P Elements
The phenomenon called hybrid dysgenesis illustrates another obvious kind of mutation enhancement caused by a
eukaryotic transposon. In hybrid dysgenesis, one strain of
Drosophila mates with another to produce hybrid offspring that suffer so much chromosomal damage that they
are dysgenic, or sterile. Hybrid dysgenesis requires a contribution from both parents; for example, in the P-M system
the father must be from strain P (paternal contributing)
and the mother must be from strain M (maternal contributing). The reverse cross, with an M father and a P mother,
produces normal offspring, as do crosses within a strain
(P 3 P or M 3 M).
What makes us suspect that a transposon is involved in
this phenomenon? First of all, any P male chromosome can
cause dysgenesis in a cross with an M female. Moreover,
recombinant male chromosomes derived in part from
P males and in part from M males usually can cause dysgenesis, showing that the P trait is carried on multiple sites
on the chromosomes.
One possible explanation for this behavior is that the
P trait is governed by a transposable element, and that is
why we find it in so many different sites. In fact, this is the
correct explanation. The transposon responsible for the
P trait is called the P element and it is found only in wild
flies, not in laboratory strains—unless a biologist puts it
there. Margaret Kidwell and her colleagues investigated the
P elements that inserted into the white locus of dysgenic
flies. They found that these elements had great similarities
in base sequence but differed considerably in size (from
about 500 to about 2900 bp). Furthermore, the P elements
had direct terminal repeats and were flanked by short direct repeats of host DNA, both signatures of transposons.
Finally, the white mutations reverted at a high rate by losing the entire P element—again a property of a transposon.
If P elements act like transposons, why do they transpose and cause dysgenesis only in hybrids? The answer is
that the P element also encodes a suppressor of transposition, which accumulates in the cytoplasm of the developing
germ cells. Thus, in a cross of a P or M male with a P female, the female cytoplasm contains the suppressor, which
binds to any P elements and prevents their transposition.
But in a cross of a P male with an M female, the early embryo contains no suppressor, and none is made at first, because the P element becomes active only in developing
germ cells. When the P element is finally activated, the
transposase and suppressor are both made, but the transposase alone goes to the nucleus, where it freely stimulates
transposition.
In 2009, Gregory Hannon and colleagues identified a
good candidate for the suppressor: a group of anti-P element piRNAs. Recall from Chapter 16 that piRNAs target
transposons in germ cells and suppress their transposition.
Hannon and colleagues found that P females had abundant P element-specific piRNAs in their germ cells, while
M females did not. The same principle applies to the similar
I element: Crosses between “inducer” (I) males, carrying the
I element and “reactive” (R) females lacking an I suppressor gave sterile progeny, whereas crosses involving I females, which had the suppressor, gave fertile offspring.
And, in accord with the hypothesis, Hannon and colleagues
showed that I females contained piRNAs targeting the
I element, while R females did not.
Hybrid dysgenesis may have important consequences
for speciation—the formation of new species that cannot
interbreed. Two strains of the same species (such as P and M)
that frequently produce sterile offspring will tend to become genetically isolated—their genes no longer mix as
often—and eventually will be so different that they will not
be able to interbreed at all. When this happens, they have
become separate species.