High Polymorphism of TPE Repeats Within Natural Populations of
Drosophila melanogaster: A Gradient of the 5TPE hobo Element in
Western Europe
Eric Bonnivard,* Claude Bazin,† and Dominique Higuet*
*Laboratoire Dynamique du Génome et Evolution, Institut J. Monod, Paris Cedex, France; and †Laboratoire Population,
Génétique et Evolution, Gif sur Yvette, France
To find out whether the polymorphism of TPE repeats of the hobo transposable element observed in some populations results from polymorphism within flies or from variability between flies, or both, we carried out isofemale
line analyses of 25 populations. We found that polymorphic populations result from the presence of polymorphic
flies combined with interfly variability within these populations. The fact that populations display different levels
of polymorphism, i.e., different types of element and different frequencies of polymorphic flies, can be used to
differentiate between qualitatively identical populations. This showed that the geographical structuring previously
observed is reinforced and, in particular, that the western European populations, which have 3TPE and 5TPE
elements, display a centrifugal decrease in the frequency of 5TPE hobo elements which start in western France.
This gradient supports the hypothesis of a dynamic invasion by this type of elements: a total invasion by 3TPE
elements, followed by further invasions involving other types of hobo elements. Moreover, the analysis of numerous
sequences in current populations revealed the existence of seven types of never-previously described hobo elements
with regard to TPE repeats. This diversity, which contrasts with the conservation of other parts of the element,
highlights the high mutation rate of the S region.
Introduction
Temporal surveys of natural populations of Drosophila melanogaster have revealed historical patterns,
suggesting that worldwide invasions of this species by
at least three transposable elements, I, hobo, and P, have
occurred during the last century (Bregliano and Kidwell
1983; Kidwell, Frydryk, and Novy 1983; Anxolabéhère,
Kidwell, and Périquet 1988; Périquet et al. 1989; Pascual and Périquet 1991). These transposable elements,
which are currently found in all natural populations, are
not found in long-established laboratory strains. The dynamics of the P element in natural populations is now
clearly understood (Anxolabéhère et al. 1984, 1985,
1990; Boussy and Kidwell 1987; Boussy et al. 1998;
Bonnivard and Higuet 1999; Itoh et al. 1999), but the
dynamics of I and hobo remain unclear. In the case of
the hobo element, previous studies, based on the correlation between collection date and the presence of the
full-size element, suggested that a recent worldwide invasion started in America some time before about 1950
(Périquet et al. 1989; Boussy and Daniels 1991; Pascual
and Périquet 1991). To clarify the dynamics of hobo
elements, and thus to help understand the hobo invasion,
current, natural populations were assessed with regard
to the repetition polymorphism of a tandem ‘‘Threonine
(T) Proline (P) Glutamic acid (E)’’ motif. These TPE
repeats consist of repetitions of a 9-bp ‘‘actccagaa’’ sequence (Streck, Mac Gaffey, and Beckendorf 1986; Calvi et al. 1991; Bazin and Higuet 1996), corresponding
to the polymorphic S region localized in the ORF1 of
the element. In hobo108, Streck, Mac Gaffey, and BeckKey words: hobo element, TPE repeats, microsatellites, Drosophila melanogaster, isofemale lines, structuring populations.
Address for correspondence and reprints: Eric Bonnivard, Laboratoire Dynamique du Génome et Evolution, Institut J. Monod, 2 place
Jussieu, 75251 Paris Cedex 05, France. E-mail: [email protected].
Mol. Biol. Evol. 19(12):2277–2284. 2002
q 2002 by the Society for Molecular Biology and Evolution. ISSN: 0737-4038
endorf (1986) described 10 perfect copies, flanked by
five degenerate ones, three in 59 and two in 39. The
autonomous hobo element of reference, Hfl1 (Calvi et
al. 1991), has only three perfect copies, which are also
flanked by degenerate ones.
Using TPE repeats as molecular markers revealed
a characteristic geographical distribution, which seems
to have been stable since the early 1960s (Bonnivard et
al. 2000). Most populations are monomorphic and contain only hobo elements with three TPE repeats (3TPE
elements). These populations belonging to the [3] class
(with solely 3TPE elements) are found worldwide. In
contrast, polymorphic populations (with several types of
element) are restricted to three regions: western Europe,
South America, and Equatorial Africa. These polymorphic populations display marked geographical structuring, with adjacent populations showing similar polymorphism. In the light of this stable distribution, we
proposed a new scenario for the dynamics of hobo elements based on there having been two distinct invasion
stages: a successful and total invasion by 3TPE elements
followed by the start of new invasions involving other
types of hobo elements, especially 5TPE elements (Bonnivard et al. 2000).
The existence of polymorphic populations raised
questions about the origin of this polymorphism. Does
it result from existence of polymorphic flies (i.e., different types of hobo sequences within a fly) or from
variability between flies within the same population (i.e.,
flies belong to different classes with regard to TPE repeats), or from both? To describe the components of the
polymorphism that was observed previously and to test
our model, we investigated 25 current, natural populations through an analysis of the isofemale lines. In current populations, this analysis revealed the existence of
seven new types of hobo elements with regard to the
TPE repeats. Moreover, we observed interfly variability
within the populations that made it possible to distin2277
2278
Bonnivard et al.
Table 1
Location and molecular characteristics in the H-E system of 25 natural populations
collected between 1992 and 1999.
POPULATIONS
a...
b...
c...
d...
e...
f ...
g...
h...
i ...
j ...
k...
l ...
m ..
n...
o...
p...
q...
r ...
s ...
t ...
u...
v...
w. . .
x...
y...
STRAIN
LATITUDE
LONGITUDE
Mass
culture
analysesa
France, Brest
France, Le Châtelet
France, Château Thierry
France, Naussac
France, Perpignan
Germany, Freiburg
Germany, Frankenthal
Germany, Kircherhenbach
Italy, Firenze
Poland, Poznan
Norway, Stavenger
USA, Boston
Costa Rica
Colombia, Carthagena
West Indies
Guyana, Kourou
Bolivia
Morocco, Marrakech
Algeria, Skikda
Dubai
Senegal, Dakar
Guinea Bissau, Bissau
Liberia, Nimba
Kenya
Madagascar
48.24
46.38
49.03
44.43
42.41
48.00
49.00
49.43
43.46
52.25
58.58
42.21
10.20
10.25
16.15
05.09
217.00
31.49
36.52
25.15
14.40
11.50
07.30
01.00
219.00
204.29
02.17
03.24
03.50
02.53
07.50
08.21
11.09
11.15
16.55
05.45
271.03
284.37
275.32
261.30
252.39
263.00
208.00
06.54
55.16
217.25
215.34
208.40
38.00
46.00
[3, 5]
[3, 5]
[3, 5]
[3, 5]
[3, 5]
[3, 5]
[3, 5]
[3, 5]
[3, 5]
[3]
[3]
[3]
[3]
[3, 5]
[3, 5]
[3, 5]
[3, 5]
[3]
nd
[3]
[3, 5]
[3]
[3, 5]
[3, 5]
[3, 5]
STATUS
hobo
Isofemale
lines analysesb
FULL-SIZE
COPY
NUMBER
[3, 5]
[3, 5, 8]
[3, 5, 8]
[3, 5]
[3, 5]
[3, 5]
[3, 5, 8]
[3, 5]
[3, 5]
[3, 3 1 1SLE]
[3]
[3]
[3, 4]
[3, 5]
[3, 4]
[21TPR 2, 3, 5, 6]
[3, 4, 5]
[3]
[3, 4]
[3]
[3, 5]
[3]
[2, 3, 3 11SLE]
[3, 4, 5]
[3, 4, 9]
6
10
10
12
4
5
4
12
6
16
20
5
10
8
2
13
5
7
4
5
7
6
3
5
9
a
Status results from Bonnivard et al. (2000) corrected with regard to the 7TPE repeats artifact (see Results).
Status of the population according to the different types of hobo elements observed in single fly analyses. The
different types of elements confirmed by sequence analysis (Fig. 1) are indicated in bold.
b
guish between populations that belong to the same polymorphic class. In particular, we describe in western Europe a centrifugal decrease in the frequency of 5TPE
hobo elements that start from western France.
Materials and Methods
Strains
Twenty-five natural populations, collected between
1992 and 1999, derived from various localities around
the world, were studied using isofemale lines (table 1).
The choice of these populations was based on the distribution described in Bonnivard et al. (2000) and their
maintenance as isofemale lines. Four of the chosen populations (k, l, r, and t) displayed unvarying regions,
where only [3] monomorphic populations are found. We
then chose three [3] populations (j, m, and v), located
in or near the regions where polymorphic populations
were found, as well as eight polymorphic populations,
four populations each from South America (n, o, p, and
q) and Equatorial Africa (u, w, x, y). Finally, nine polymorphic populations (a to i) were used to investigate the
marked geographical structuring seen in western Europe. In all these populations, the TPE motif characteristics were investigated in 10 isofemale lines (eight lines
for Liberia, Nimba [w] and Algeria, Skikda [s]).
Determining the TPE Status of Isofemale Lines
For each isofemale line, DNA was extracted from
a single fly, using the method described by Di Franco
et al. (1995). PCR amplification was performed using
the h6 and h11 internal primers of the Hfl1 element, as
described in Bazin and Higuet (1996). Preliminarily,
PCR products were electrophoretically segregated by
size in 2% agar gel on a Mupid-21 (Eurogentec), migration 3 h at 100 V. This made it possible to identify
the different profiles present in a population by comparing the length of the PCR product with the sequenced
control fragments. Then, for each of the populations, we
identified lines that were representative of the different
migration profiles observed.
For the 50 representative lines identified, PCR
products were also separated on polyacrylamide gel to
confirm the different numbers of TPE repeats observed.
For this purpose, PCR amplification was performed using a 33P-labeled h6 primer. After amplification, samples
were diluted with one volume of loading dye (95%
formamide, 0.005% xylene cyanol FF, and 0.005%
bromophenol blue), heat-denatured at 948C for 5 min,
and then directly cooled on ice. For each sample, a 5to 7-ml aliquot was loaded onto a 6% denaturing polyacrylamide gel. After migration, gels were transferred to
Whatman 3 MM paper and vacuum dried at 808C for 1
h, and the dried gels were exposed to X-ray film.
hobo TPE-Repeats Polymorphism Within Populations
More than 35 TPE repeat bands were chosen on
the basis of their size and the population from which
they had come, such that at least an example of each
type of TPE repeat was included from each of the geographical regions. PCR amplifications were performed
on these selected TPE repeat bands excised from the
dried gels, according to Melayah et al. (2001), and sequenced using the specific amplifying primer h11.
Southern Blot
The Southern blot method was used to estimate the
number of full-size hobo elements (2.6 kb XhoI fragments) using the method described by Bonnivard and
Higuet (1999). For this purpose we used a hobo probe
corresponding to the 1756–2168 internal sequence obtained by PCR amplification using the h6 and h11 primers. The full-size elements were quantified using the
CyHBL1 strain as a reference (one Hfl copy per diploid
genome, Calvi and Gelbart 1994). This enabled us to
estimate the number of these elements present in the
different strains.
Sequence Polymorphism Analysis
To research polymorphism beside the S region, we
first used the sequences of h6-h11 PCR amplification
products. Second, we investigated part of the no-coding
region in 39 of the ORF1 (Streck, Mac Gaffey, and
Beckendorf 1986) using two primers, h21 and h4, which
have sequences corresponding to bases 2267–2286 (59ACAAAAACCTAAACAACTCG-39) and 2879–2859
(59-ACCCTACTTGCGGCAACACA-39), respectively,
in the Hfl1 element (Calvi et al. 1991). PCR amplifications were performed on DNA extracted from single fly
with only one type of element with regard to the TPE
repeats and were sequenced using the specific amplifying primer h21.
Results
High Polymorphism of TPE Repeats
PCR amplifications of the S region were carried out
on 246 isofemale lines obtained from 25 different natural populations. Nine different hobo elements were de-
2279
tected (fig. 1). They correspond to 3TPE or 5TPE elements, which have already been reported elsewhere, and
to seven new types of hobo elements, which have never
previously been described. Five of them correspond to
variations in the number of TPE motifs, resulting in
2TPE, 4TPE, 6TPE, 8TPE, or 9TPE elements, respectively. Our results do not support the existence of a
7TPE hobo element suggested by Bonnivard et al.
(2000). In two cases, the sequenced S regions did not
match the expected number of TPE copies due to two
particular types of hobo elements (fig. 1). The first element, the size of which corresponds to four TPE repeats, actually contains only three perfect TPE repeats
linked to a duplication of the 39-adjacent 9-bp sequence
that encodes a ‘‘Serine (S) Leucine (L) Glutamic acid
(E)’’ motif. This element is defined as 3TPE 1 1SLE
element (simplified notation: 3 1 1SLE). The second
element, corresponding in size to one TPE repeat, in fact
has two perfect TPE repeats that are linked to a deletion
of the 59-adjacent 9-bp sequence encoding a ‘‘Threonine
(T) Proline (P) Arginine (R)’’ motif. This element is
defined as 21TPR 2TPE element (simplified notation:
21TPR 2).
Characterization of Natural Populations with Regard to
TPE Repeats
To determine how to characterize an isofemale line,
16 lines were analyzed, each using three to five flies
separately. Whatever the conditions (geographical origin, monomorphic or polymorphic flies, flies with rare
types of hobo elements, flies from the same or different
generations), no difference in TPE profile could be detected between flies from the same isofemale line. Thus,
a single fly corresponds to an isofemale line.
Existence of Polymorphism Within Flies
This study of isofemale lines enables us to define
a new class of monomorphic flies, the [5] class, represented by two French lines (fig. 2). Hence, it appears
that some flies do not contain the 3TPE hobo element.
However, this element type is still found to be predominant, as it is present in all the other 244 lines tested
(being sequenced in 14 flies representing 11 populations,
FIG. 1.—Sequence variability of the S region of hobo elements reported in current, natural populations. Sequences have been aligned on
the Hfl1 element sequence (Calvi et al. 1991) chosen as a reference (at the foot). The TPE-repeats motif is indicated in bold, and the adjacent
motifs are indicated by gray boxes. Dashes indicate sequence deletion, and blanks have been introduced in the reference sequence to allow for
nucleotide insertions.
2280
Bonnivard et al.
FIG. 2.—Distribution of flies within natural populations with regard to their TPE status. For each population (bars), 10 flies (single boxes)
from different isofemale lines are analyzed (only eight flies for Nimba [w] and Algeria [s]). Flies belong to six different classes, according to
both the number and the type of elements detected. For clarity, types of elements present in less than two flies or populations are not distinguished
but are grouped in the [3 & others] class. (A) Populations from America and Africa. (B) Focus on populations from western Europe. Letters
refer to table 1.
table 1). The 137 other monomorphic flies all belonged
to the [3] class. In addition, several different polymorphic classes of flies can be defined (fig. 2). Two of them
seem to be preponderant: 5TPE elements were found in
75 flies from 14 different populations (sequenced in
eight of them, table 1), and 4TPE elements were found
in 18 flies from six different populations (sequenced in
all of them, table 1). All the other types of elements
seemed to occur at low frequencies, as they occurred in
less than two lines (six lines for the 8TPE element).
Moreover, the flies that harbor these types of elements
present a higher signal for 3TPE, 4TPE, or 5TPE elements in agarose gel. Such differences in the intensity
of the PCR product can be interpreted as differences in
the ratio of the number of elements of each type, according to Brunet et al. (1996). To confirm this hypothesis, we used two transformed lines, which have a single
3TPE or 5TPE hobo element, respectively. DNA extractions were performed on different mixes of these
flies (ratio 1:9, 2:8, 5:5, 8:2, and 9:1). PCR amplification
results show a difference in the intensity of the PCR
products that correlates with the ratio of the transformed
flies. Hence, according to the intensity of the PCR product, these particular types of element also seem to be in
the minority within the genome.
Interfly Variability Within Populations Explains the
Variability Between Populations
The level of polymorphism of the 25 populations
studied is shown in table 1 and figure 2. Five populations were monomorphic and consisted solely of flies
with 3TPE elements. Four of these populations (Stavenger [k, not shown in fig. 2], Boston [l], Marrakech [r],
and Dubai [t]) are located in regions where only monomorphic populations had previously been observed. This
confirms that such populations present only 3TPE hobo
elements. The fifth population (Bissau [v]) came from
western Africa, where polymorphic populations also
were found.
Polymorphic populations show different levels of
interfly variability, depending on the frequency of polymorphic flies that they display. Some populations have
only a few fractions of polymorphic flies, about 1 or 2
hobo TPE-Repeats Polymorphism Within Populations
FIG. 3.—Decrease in the frequency of 5TPE elements in western
European populations. The number of flies that present 3TPE elements
(rhombus) or 5TPE elements (square) was plotted against the geographic distance from Brest. (Letters refer to populations in table 1
and fig. 2B.)
out of 10. They are mainly located around the edge of
the region where TPE-repeat polymorphism is observed
(e.g., Poznan [j] or Costa [m]). For the sake of clarity,
the uncommon classes of flies are not shown in figure
2. They harbor: the 2TPE element in one fly in Nimba
(v); the 9TPE element in one fly in Madagascar (y); the
3 1 1SLE element in two independent flies in Poznan
(j) and Nimba (v); and 21TPR, 2TPE, or 6TPE elements in few flies from Kourou (p), some of which harbored up to four different types of hobo elements, belonging to the [21TPR 2, 3, 5, 6] class. In contrast,
several populations contain a majority of polymorphic
flies. This polymorphism mainly results from 4TPE or
5TPE elements in addition to 3TPE elements. For example, elements with four and five TPE repeats are accountable for the high polymorphism level observed in
eastern Africa and western Europe, respectively.
A Gradient of 5TPE hobo Elements in Western
European Populations
In western Europe, populations contain a majority
of flies of the [3,5] class (fig. 2B), especially in France,
where only two monomorphic [5] flies were observed.
The other 48 French flies were polymorphic, and all but
four belonged to the [3,5] class. The remaining four flies
also contained 8TPE elements. In the three German populations, monomorphic flies of the [3] class were observed but were still a minority. They make up a majority only in the Italian population Firenze (i), and, to
the east, in Poland, Poznan (j). This implies that the
western European populations display marked geographically determined structuring, as geographically
adjacent populations show similar kinds of interfly
variability.
To study the marked geographical structuring,
3TPE or 5TPE hobo elements were considered separately, and their frequency in each of the European populations was determined (fig. 3). 3TPE hobo elements
were present in all flies, except one in Brest (a) and one
in Naussac (d). On the other hand, there was a marked
decrease in the number of isofemale lines harboring
5TPE elements that correlated with the distance from
Brest. 5TPE hobo elements were present in all French
flies but were present in progressively fewer and fewer
2281
FIG. 4.—Number of Hfl elements in western European populations
versus their geographic distance from Brest. (Letters refer to populations in table 1 and fig. 2B.)
flies in the German and Italian populations and were
completely absent from the Polish flies. The data therefore revealed a relationship between the number of flies
presenting 5TPE elements and the location of each population. This conclusion is reinforced when we consider
the intensity of the different PCR products. Indeed, the
eight [3,5] flies in which the 3TPE repeats product could
not be detected easily were all French. This fits in with
the fact that two French flies belonged to the [5] class.
In contrast, the 11 [3,5] flies in which the 5TPE repeat
product could not be detected easily were all German.
In conclusion, the frequency of 5TPE hobo elements
seems to decline steadily and centrifugally with distance, irrespective of the direction from Brest.
Number of Full-Size Elements
The presence of 5TPE elements, or of other types
of elements, in addition to 3TPE elements raises the
question of how many hobo elements are found within
polymorphic populations. Indeed, the fact that populations present different levels of polymorphism, i.e., different types of elements and different frequencies of
polymorphic flies, could influence their number of hobo
elements. For example, populations consisting of a majority of polymorphic flies (or populations that include
many different types of elements) could have a greater
number of elements. This implies that the gradient detected in western Europe could be reflected by the number of elements. To examine this possibility, we looked
at the estimated number of full-size hobo elements (Hfl)
because they are the main hobo elements carrying TPE
repeats (Bonnivard et al. 2000). The data (table 1) show
that there were between 2 and 20 copies of Hfl, with an
average of eight, which is in agreement with our previous results (Bonnivard, Higuet, and Bazin 1997). The
wide range of numbers of Hfl makes it impossible to
detect any particular distribution of either the geographical position of the populations or their invasion by new
types of elements with more than three TPE repeats.
First, in western Europe, there is no correlation between
the number of Hfl and the location of the populations
(Spearman rank correlation: rs 5 0.2, n 5 10, P . 0.05).
Hence, there is no particular gradient (fig. 4) to match
the one found for the frequency of flies with 5TPE elements (fig. 3). Second, the number of Hfl does not reflect the level of interfly variability. For example, Per-
2282
Bonnivard et al.
pignan, where all the flies seemed to be polymorphic,
has only four copies (i.e., only about one-third the number of copies as other French populations), whereas
Kircherhenbach, where only half of the flies are polymorphic, has about 12 copies. This may be because
there is a considerable variability in the number of copies, which ranges from 4 to 20 in populations of the [3]
class.
Sequence Polymorphism
To find out whether each type of element observed
on different continents has a single origin or not, we
were interested in finding a linkage disequilibrium between the number of TPE repeats and other polymorphic
loci in the hobo sequence. For this purpose, the sequences of h6-h11 PCR products were compared with the Hfl1
sequence. These PCR products come from 18 different
populations and correspond to all the different types of
elements described. Our data do not reveal polymorphism for any type of hobo element anywhere other than
in the S region. Then, we investigated about 600 bp of
the no-coding region in 39 of the ORF1 of the hobo
element. Four monomorphic isofemale lines were studied, three belonging to the [3] class (from Freiburg, Costa, and Dubai) and the other to the [5] class (from Brest).
Sequences performed directly on h21-h4 PCR products
(see Materials and Methods) did not reveal any polymorphism, even though these flies came from widely
distant regions. These findings show that even though
some hobo elements could display sequence polymorphism other than the TPE repeats, most of them have
the same sequence that Hfl1 has. The same consensus
sequence therefore is always found beside the S region,
and no polymorphism of hobo elements in natural populations can be used to infer the origin of 4TPE or 5TPE
hobo elements.
Discussion
Results reveal that polymorphic populations result
from the existence of polymorphic flies combined with
interfly variability. Actually, a given population can contain monomorphic flies as well as polymorphic flies belonging to different classes. It is the ratio between these
different flies that accounts for the polymorphism observed at the population level. Moreover, the polymorphism also results from an unexpected diversity of the
hobo elements with regard to TPE repeats, which raises
the question of their origin, both at the molecular and
geographical levels.
Characterization of Populations with Regard to TPE
Repeats
The polymorphism observed in some populations
using mass culture can be redefined on the basis of the
interfly variability. Of the seven populations of the [3]
class, chosen to confirm that they really are monomorphic, only populations from Costa Rica and Poznan contained one polymorphic fly. Hence, if we assume that
mass cultures had been established at least two gener-
ations before the molecular analysis, it is quite conceivable that they had escaped detection in the previous
study. It also appears that these two populations were
chosen for their particular location, on the borders of
regions where polymorphic populations were found (see
Materials and Methods). On the other hand, all the populations located in regions where only monomorphic
populations were found displayed only 3TPE hobo elements, as expected.
Considering polymorphic populations, isofemale
lines analyses not only make it possible to achieve better
distinction between populations in different classes but
also are able to discriminate between populations belonging to the same polymorphic class. Some differences appear between mass culture and isofemale lines data
(table 1). Such differences have two causes. The first
cause is that the isofemale line approach is able to detect
types of elements that are only present at a low frequency within a population, such as 2TPE, 8TPE, or
9TPE elements, and may not be detected using mass
culture analysis. The second cause is that polyacrylamide gel analysis can be used for directly sequencing
elements of interest. This can account for the detection
of 4TPE elements that were probably mistaken for 5TPE
elements in earlier studies using only agarose gel.
Molecular Origin of the Different Types of Elements
According to the length of the repeats unit, TPE
repeats can be consider as microsatellites or minisatellites, depending on the authors (Tautz 1993; Debrauwere, Gendrel, and Dutreix 1997), even though their size
corresponds more closely to microsatellites. Two mechanisms have been proposed to account for the instability
of such repetitive sequences: DNA polymerase slippage,
which could account for the low modification of the
number of repeats, and unequal recombination, which
reshuffles repeat variants (for review see Debrauwere,
Gendrel, and Dutreix 1997). The existence of 3TPE,
4TPE, and 5TPE elements in the same region (e.g., Bolivia or Kenya), as well as the deletion of the degenerate
TPR motif in 59 or the duplication of the degenerate
SLE motif in 39, can be used to argue for a mechanism
linked to polymerase slippage. On the other hand, the
absence of elements with an intermediate number of
TPE repeats, between five and eight in Europe or between four and nine in Madagascar, conflicts with such
an evolutionary model and could argue for recombination events. The instability of microsatellites depends on
numerous parameters such as the size and sequence of
the repeated motif, the number of repeats, the monotony
of the series, the locus, and positive mutational bias (Estoup and Cornuet 1999). All these parameters can also
interact with the mutation rate; hence, the evolution processes of microsatellites depend on the locus considered.
In this special case of a microsatellite within the coding
region of a transposable element, more information is
needed to infer the evolution of the number of TPE repeats. However, some particularities of this locus can
already be pointed out. First, TPE repeats present unexpectedly great diversity, given that their worldwide
hobo TPE-Repeats Polymorphism Within Populations
invasion of D. melanogaster is supposed to be a recent
event, occurring within the past 70 years. This diversity
reflects a high mutation rate that contrasts with the conservation of other parts of the element and with the average mutation rates of microsatellite loci in D. melanogaster (6.5 3 1026 per locus per generation; Schug,
Mackay, and Aquadro 1997). This high mutation rate
could be related to the number of copies and to replication resulting from the transposition of the elements.
Second, variations always involve sequences of nine nucleotides, usually TPE motifs but also motifs that are
different at the protein level. Third, it is necessary to
consider that the S region may not be neutral because it
is located in the coding sequence. Thus, it is also possible than some types of elements were undetected because they were eliminated by selection pressure.
The Two-Step Invasion Model
The results obtained in this study are still consistent
with the scenario of a two-stage invasion, consisting of
a total invasion by 3TPE elements followed by new invasions involving other types of hobo elements (Bonnivard et al. 2000). Each of the less common types of
hobo element is found only in restricted geographical
regions; as a result, at present, no data can argue for
their invasion capacity. Indeed, elements may be considered as invasive only if they are found in several
populations on a large geographical area, for example,
5TPE elements in Europe.
The centrifugal gradient of 5TPE hobo elements
described in western Europe can reflect a new invasion
of the European populations by these elements, corresponding to the second invasion stage. An alternative
hypothesis is that 5TPE elements occurred earlier in Europe, but if this were so, there is no way of explaining
the origin of these elements or how they have faded out
in many European flies. Fly migrations are preponderant
factors in the transposable-element invasion of D. melanogaster (Quesneville 1996; Bonnivard 1999). Stepby-step recurrent migrations play a major role in establishing centrifugal gradients such as the one described
in this study or the one described in the investigation of
natural populations with regard to the number of fullsize P elements (Bonnivard and Higuet 1999). Hence,
if we compare the two systems, the stable distributions
in western Europe present a remarkable spatial likeness
with the centrifugal gradient centered in the west of
France that decreases sharply in central Europe. This
similarity is difficult to interpret because it cannot result
from fly migration alone. Indeed, in the P-M system, the
distribution also results directly from the activity and
repression properties of the elements (Bonnivard and
Higuet 1999). Indeed, mobility of the element within the
genome is a second factor that may be implicated in
their invasion capacity because elements that present activity can be amplified within the genome. Considering
5TPE elements, this mobility could reflect the activity
of the elements themselves or their mobilization in trans
because we are still not sure that a hobo element with
more than three TPE repeats corresponds to complete
2283
elements. However, some data argued for the likely activity properties of 5TPE elements. First, most of the
hobo elements carrying TPE repeats are full-size elements, as deleted elements that display the S region remain a minority, especially in the European populations
(Bonnivard et al. 2000). Second, according to the preponderance of these elements in France, it seems that
these elements not only invade French populations but
also replace 3TPE elements.
Considering 4TPE elements, their geographical distribution, especially their high frequencies in eastern Africa, suggests that these elements are also potentially
invasive and could participated in the second invasion
stage. However, at present, the analysis of natural populations suffers from the lack of samples from South
America and South Africa. Studying natural populations
will allow us to determine if there is a continuum between these two regions and a gradient such as the one
described for 5TPE elements.
Acknowledgments
We would like to thank Cécile Balanant for her
helpful technical assistance, Monika Ghosh for language
revision, and anonymous referees for their helpful comments. This work was supported by GDR 2157-CNRS—
‘‘Evolution des éléments transposables: du génome aux
populations’’ and Paris 6 University.
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Accepted August 22, 2002