1 Stowers Institute for Medical Research, 1000 East 50th Street, Kansas City, MO
64110, USA
2 Department of Anatomy and Cell Biology, University of Kansas School of
Medicine, 3901 Rainbow Boulevard, Kansas City, KS 66160, USA
* Author for correspondence (e-mail: tgx{at}stowers-institute.org)
Accepted 19 March 2003
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SUMMARY |
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Key words: Germline stem cell, Niche, Drosophila, Ovary
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INTRODUCTION |
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The Drosophila ovary is an excellent system to study stem cells
and their relationships to niches (Lin,
1998; Xie and Spradling,
2001
). Each ovary is composed of 12-16 individual ovarioles, in
which both germline and somatic stem cells are located at the tip of the
ovariole also known as the germarium. Two or three somatic stem cells are
located at the middle of each germarium and are responsible for producing
somatic follicle cells that surround germline cells in developing egg chambers
(Margolis and Spradling, 1995
;
Zhang and Kalderon, 2001
). Two
or three GSCs that situate at the tip of each germarium generate
differentiated germline cysts (Wieschaus
and Szabad, 1979
; Lin and
Spradling, 1993
). These GSCs have recently been shown to be
located in a niche, which is composed of three differentiated somatic cell
types: terminal filament (TF) cells, cap cells and inner germarium sheath
(IGS) cells (Cox et al., 1998
;
Cox et al., 2000
;
King and Lin, 1999
;
King et al., 2001
;
Xie and Spradling, 1998
;
Xie and Spradling, 2000
). A
GSC divides asymmetrically to generate one stem cell and one cystoblast
daughter. The cystoblast divides precisely four times to produce 16
interconnected cystocytes that are then encapsulated by a layer of follicle
cells to form an egg chamber (Spradling,
1993
).
Germline stem cells in each niche can be easily recognized by their size,
location and the presence of a special intracellular organelle known as a
spectrosome. Spectrosomes, like their counterparts in differentiated germ cell
cysts known as fusomes, are rich in cytoskeletal proteins such as Huli-tai
shao (Hts) (Lin et al., 1994).
The spectrosome is found in GSCs and cystoblasts usually as a spherical
structure; the fusome in cysts is branched thereby connecting individual
cystocytes. In the germarium, GSCs are in direct contact with cap cells, and
their spectrosome is invariably anchored to the cap cell contact site.
Adherens junctions exist in the interface between cap cells and GSCs, and
their disruption causes GSC loss (Song et
al., 2002
). Furthermore, GSCs divide along the anteroposterior
germarial axis so that the anterior GSC daughter remains anchored to cap cells
and maintains stem cell identity, while the posterior daughter that fails to
contact cap cells differentiates into a cystoblast. However, when one of the
GSCs in a niche is lost, its neighboring stem cell divides perpendicular to
the germarial axis, causing both daughter cells to contact cap cells and to
retain stem cell identity and thus repopulate the niche
(Xie and Spradling, 2000
).
Consistently, it has been shown that in the adult ovary, TFs/cap cells express
many genes that are known to be important for maintaining GSCs, such as
hedgehog (hh), piwi, fs(1) Yb and dpp
(Cox et al., 1998
;
Cox et al., 2000
;
King and Lin, 1999
;
King et al., 2001
;
Xie and Spradling, 1998
;
Xie and Spradling, 2000
).
Therefore, direct interactions with niche cells, especially cap cells, are
essential for maintaining GSC identity.
Relatively little is known about germ cell development in the female gonads
of larvae and pupae. GSCs in the adult Drosophila ovary originate
from embryonic pole cells. The pole cells proliferate and migrate from the
posterior end of the embryo to the gonadal mesoderm to form the primitive
gonad (Mueller, 2002). In
females, the primordial germ cells (PGCs) and somatic cells in the gonad
increase dramatically in number during the larval period. Individual TFs that
consist of eight or nine disc-shaped cells are formed 2 hours before pupation
(King, 1970
). With the aid of
molecular markers, it has been shown that TFs form in a progressive manner
from medial to lateral across the ovary, with the number of terminal filament
cells increasing gradually during the second half of the third instar larval
stage (Sahut-Barnola et al.,
1995
). We have recently shown that adherens junctions establish
between cap cells and newly formed GSCs during early pupation
(Song et al., 2002
). A genetic
study suggests that the establishment of GSCs takes place during the early
pupal stage (Bhat and Schedl,
1997
). As there are more PGCs than needed for the formation of
12-16 ovarioles, each of which contains two or three GSCs, it has been
proposed that the extra PGCs directly enter the germ cell differentiation
pathway without passing through the stem cell stage
(King, 1970
;
Bhat and Schedl, 1997
).
However, nothing is known about how PGCs are selected to become GSCs or to
differentiate directly, or how the selected PGCs populate the niche. In this
study, we show that PGCs are selected to become GSCs based on their
juxtaposition to TFs/cap cells. We further demonstrate that GSCs in a niche
can come from one PGC and that dpp signaling probably controls the
proliferation of GSCs in the niche.
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MATERIALS AND METHODS |
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Developmental staging of larvae and pupae
Morphological definitions of the developmental stages of
Drosophila followed those of King
(King, 1970). In this study,
late third instar larvae were referred to as the third instar larvae that
remained in food. At this stage, terminal filament stacks started to form. The
larvae at the larval-to-pupal transition were referred to as the larvae that
moved out of food but pupation had not started. At this stage, most of TFs
were still forming and cap cells were starting to form. The early pupal stage
was the stage at which pupation had already started but pupae were still pale
and clear. At this stage, all TFs were finished and a few cap cells had
already formed.
Immunohistochemistry and microscopy
Ovary dissection, fixation and immunohistochemistry were performed as
described previously (Song et al.,
2002). The following antibodies were used: monoclonal anti-Hts
antibody 1B1(1:3) (Developmental Studies Hybridoma Bank at the University of
Iowa); rat anti-Bam antibody (1:100)
(McKearin and Ohlstein, 1995
);
polyclonal anti-ß-galactosidase antibody (1:100) (Molecular Probes);
polyclonal anti-GFP antibody (1:100) (Molecular Probes); and monoclonal
anti-BrdU antibody (1:20) (Oncogene). All micrographs were taken using a Leica
confocal NT II microscope.
Clonal analysis and calculations
To generate positively marked germ cells by tubulin-lacZ in the
developing female gonads, larvae from the cross between X-15-29 females and
hsFLP; X-15-33/CyO males were heatshocked at or before the late third
instar larval stage in a 37°C water bath for 4 hours, and were then
allowed to develop into adults. The ovaries from one- to two-day-old females
of hsFLP; X-15-33/X-15-29 were dissected and immunostained with
anti-Hts and anti-ß-gal antibodies. The percentage of germaria containing
only marked GSCs was determined by dividing the number of germaria containing
only marked GSCs by the number of germaria containing any marked GSCs.
To generate marked germ cells by loss of armadillo-lacZ expression in the developing female gonads, larvae from the cross between FRT40A males and hsFLP; FRT40A armadillo-lacZ females were heatshocked at or before the late third instar stage in a 37°C water bath, and were then allowed to develop into adults. The ovaries from one- to two-day-old females of hsFLP; FRT40A/FRT40A armadillo-lacZ were dissected and immunostained with anti-Hts and anti- ß-gal antibodies. For generating the marked mutant tkv germ cells at the late third instar larval stage, the FRT40A tkv8 strain was used instead of the FRT40A strain.
Whole-mount mRNA in situ hybridization
The dpp probe was labeled with digoxigenin following the DIG RNA
Labeling Kit instruction (Roche). The mRNA in situ hybridization was performed
according to the protocol described by Tautz and Pfeifle
(Tautz and Pfeifle, 1989),
except fluorescent tyramide detection was used. The combination of
immunostaining with an anti-Hts antibody and fluorescent dpp mRNA in
situ hybridization was performed according to a published protocol
(Wilkie and Davis, 2001
).
BrdU and TUNEL labeling
BrdU labeling was performed for 1 hour in Grace's medium as described
previously (Lilly and Spradling,
1996). The TUNEL cell death assay was performed following the
ApopTag apoptosis detection kit manual (Intergen Company).
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RESULTS |
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To determine whether TF/cap cell formation correlates with initial PGC
differentiation during Drosophila ovarian development, we used a
hh-lacZ line to mark TFs/cap cells in the developing female gonads in
addition to using the bam-GFP as a marker for germ cell
differentiation. In the hh-lacZline (the bacterial lacZ gene
expression controlled by the hh promoter), lacZ is
specifically expressed in TFs and cap cells of the adult ovary
(Forbes et al., 1996). As
expected, hh-lacZ was expressed in newly formed TF cells and cap
cells (Fig.
1E,F).
As hh-lacZ was expressed in both TF cells and cap cells, we
distinguished them based on their morphology. Eight or nine oval shaped TF
cells were packed very tightly against each other in each stack, while cap
cells were rounder and did not line up with TF cells. Consistent with previous
studies (King, 1970
;
Sahut-Barnola, 1995
), TF cells
started to form at the late third-instar larval stage, and the number of TF
cells in a developing stack increased gradually in a progressive manner across
the ovary until early pupal stage (Fig.
1E). bam-GFP was not expressed in the PGCs at the late
third instar larval stage regardless of their location
(Fig. 1E; 12 female gonads
examined). During the larval-pupal transition, bam-GFP was expressed
in 1.5% of the PGCs adjacent to TFs/cap cells, but its expression in the rest
of the PGCs was heterogenous, ranging from 3.5% to 99.0% with an average of
71.5% (12 female gonads examined). After TF formation, cap cell
differentiation occurred from the larval-pupal transitional stage to the early
pupal stage (about 0-4 hours after pupation)
(Fig. 1F). At the early pupal
stage, all eight or nine oval TF cells were packed tightly against each other
along their anteroposterior axis, and rounder lacZ-positive cap cells
accumulated at the posterior end of TFs. By then, bam-GFP had been
expressed at high levels in 93.0% of the germ cells that were not in contact
with TFs/cap cells, but only 2.7% of the PGCs that were close to TFs/cap cells
expressed bam-GFP (Fig.
1F; 12 female gonads examined). The remaining 7.0% of the
posterior germ cells probably represented newly produced germ cells from
anterior PGCs. Consistent with this interpretation, newly produced cystoblasts
also fail to express bam-GFP at high levels in the adult ovary
(Chen and McKearin, 2003
). The
formation of TF/cap cells prior to bam expression during early
ovarian development suggests that signals from TFs/cap cells are important for
preventing anterior PGCs from differentiating (Bam expression) and for then
allowing them to become GSCs.
GSCs in one niche can originate from one PGC
To gain further evidence supporting GSC establishment at the early pupal
stage, we carefully examined spectrosome positioning and division patterns of
PGCs that were in contact with cap cells. Owing to the lack of a definitive
GSC marker, two criteria are often used to determine GSC identity in the adult
ovary (Lin, 1998;
Xie and Spradling, 2001
). One
is that the spectrosome of GSCs is anchored to the cap cell contact site; the
other is that a GSC divides asymmetrically and generates two daughters with
only one of them remaining in contact with cap cells. At the larval-pupal
transitional stage, most of the female gonads did not have obvious cap cells
but had only eight or nine precisely packed oval TF cells, and the spectrosome
of the germ cells in the anterior row was not positioned to the anterior side
(Fig.
2A,B).
This observation indicates that before the early pupal stage, GSCs have not
yet been established. At the early pupal stage, cap cells were evident by a
few rounder lacZ-positive cells that were positioned posterior to the
eight or nine nicely packed TF cells (Fig.
2C,D).
After cap cell formation, spectrosomes in some of the PGCs that were
juxtaposed with cap cells started to be anchored to the anterior side that was
in contact with cap cells (Fig.
2C), suggesting that the establishment of GSCs takes place during
this period. Furthermore, the putative GSCs juxtaposing cap cells continued to
divide (Fig. 2D), exhibiting
two distinct division patterns: one division pattern generated two daughters
in which only one was in contact with cap cells, while the other pattern
generated two daughters that were both in contact with cap cells. To exclude
the possibility that two germ cells connected by an elongated fusome in either
division pattern are a two-cell cyst, we examined the expression of
bam- GFP in the gonad in which TFs/cap cells were identified
by hh-lacZ expression. The differentiated cysts always express
bam-GFP in the adult ovary (Chen
and McKearin, 2003
). Interestingly, the two cells generated by
either division pattern failed to express bam-GFP (Fig.
2E,F),
indicating they were two daughters of a newly established GSC rather than a
two-cell cyst. The division pattern that generates two daughters contacting
cap cells that then become two GSCs can be seen in the adult ovary during stem
cell replacement (Xie and Spradling,
2000
). Therefore, we predicted that GSCs in some niches might come
from one PGC.
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|
Dpp signaling is capable of stimulating PGC proliferation
Among many possible explanations for clonal expansion of GSCs in a niche,
the most attractive is that mitogenic signals from TFs/cap cells stimulate
PGCs to divide and produce two daughters that will directly contact cap cells
and become GSCs before other PGCs enter the niche. If this prediction is
correct, we would expect that the PGCs adjacent to TFs/cap cells are more
mitotically active than the rest of the PGCs. To determine the mitotic
activity of PGCs during niche formation, we examined the distribution of PGCs
in the S phase of the cell cycle after the incorporation of a nucleotide
analog, bromodeoxyuridine (BrdU), into the gonads ranging from the late third
instar larval stage to the early pupal stage. The gonads were further
immunostained with anti-BrdU and anti-Vasa antibodies to visualize
BrdU-positive cells and germ cells, respectively. The presence of BrdU in the
nucleus indicates a cell in the S phase of the cell cycle. At the late
third-instar larval stage, a few PGCs were positive for BrdU and were randomly
distributed in the PGC zone (data not shown). At the early pupal stage, we
observed that more PGCs were positive for BrdU and that those cells were
located preferentially close to TFs/cap cells (Fig.
4A,B).
Of the PGCs that were adjacent to TFs/cap cells, 31.1±5.5% were
BrdU-positive, in contrast to only 11.6±1.6% for the remaining PGCs
(total of nine gonads examined), indicating that the anterior PGCs are more
mitotically active than the rest of the PGCs. Interestingly, many of the
somatic cells throughout the gonads, including the somatic cells mingled with
the PGCs, were positive for BrdU, indicating that the somatic cells are also
very active in proliferation at this developmental stage (Fig.
4A,B).
These results suggest that a mitogenic signal(s) from TF/cap cells stimulates
PGC division during niche formation.
|
To investigate further whether dpp can stimulate PGC proliferation, we used the GAL4-UAS expression system to overexpress dpp in the somatic cells of female gonads and examined PGC proliferation. The GAL4 line, C587-gal4, was used to overexpress dpp in the somatic cells throughout the developing female gonad. The C587-gal4 line drives expression of UAS-GFP in most of the somatic cells but not in germ cells in the developing female gonads (Fig. 5A,B). To test whether we could achieve dpp overexpression throughout the developing female gonads, we used the Dad-lacZ line to detect dpp action in gonadal cells. dpp overexpression caused Dad-lacZ to be expressed in all the somatic cells and PGCs, indicating dpp activity everywhere in the gonads (Fig. 5C,D). It appeared that the somatic cells expressed Dad-lacZ at higher levels than the germ cells after dpp was overexpressed (Fig. 5C,5D). However, most of the somatic cells normally express Dad-lacZ at much lower levels than anterior PGCs (Fig. 4E), suggesting that the somatic cells are more sensitive to elevated dpp expression. This also raises a possibility that dpp could indirectly affect germ cell proliferation by regulating the production of other signals that are important for germ cell proliferation. To determine quantitatively the effect of dpp overexpression on the accumulation of PGCs in the gonad, we counted PGCs based on the number of spectrosomes in the gonad. The female gonads of the early pupae overexpressing dpp or GFP (control) by C587-gal4 were labeled with an anti-Hts antibody to visualize the spectrosomes. In female gonads overexpressing dpp, the number of PGCs per gonad was increased, averaging 245±92 per gonad (total of 10 gonads examined), in contrast to 136±26 (total of 14 gonads examined) observed in control female gonads. These results indicate that the increase in dpp signaling causes the accumulation of more PGCs in the developing female gonads.
|
tkv is essential for GSC clonal expansion in a niche
Owing to the stringent requirement of dpp during embryogenesis, it
is difficult to test directly the effect of dpp mutations on the PGC
proliferation and clonal expansion of GSCs. To determine whether dpp
is required for GSC clonal expansion, we tested the requirement of
dpp downstream components, thick veins (tkv) and
mothers against dpp (mad), for populating adult GSC niches.
tkv, which encodes a serine/threonine kinase receptor, is essential
for transducing the dpp signal in all tissues that require
dpp (Nellen et al.,
1994; Penton et al.,
1994
; Brummel et al.,
1994
). Mad is a transcription factor that is phosphorylated upon
dpp signaling and is responsible for activating dpp target
genes (Sekelsky et al., 1995
;
Newfeld et al., 1997
). To test
the requirement of tkv and mad for clonal GSC expansion, we
removed their function from PGCs just before they were recruited into their
niches by using the FLP-mediated FRT recombination and strong tkv and
mad alleles, tkv8 and mad12.
In the control, 11.2% of the ovarioles carried marked wild-type GSC clones
(total 331 ovarioles examined). Under the exactly same conditions, only 2.5%
of the ovarioles carried mutant tkv GSC clones, and instead many
marked mutant tkv GSCs were lost before adulthood, which was evident
by the presence of mutant tkv cysts but the absence of marked mutant
tkv GSCs in the germarium (total 937 ovarioles examined; Fig.
6A,B).
No ovarioles carrying mutant mad12 GSC clones were
recovered but the ovarioles with mutant mad12 germ cells
in egg chambers were observed (total 500 ovarioles examined), indicating that
marked mad12 GSCs could not be maintained before
adulthood. These results suggest that dpp signaling may be involved
in maintaining GSCs before adulthood. In the ovarioles that carried marked
wild-type GSC clones (total of 37 marked GSC clones examined), 36.5% of them
contained only marked GSCs, indicating the clonal expansion of GSCs. By
contrast, none of the ovarioles that carried marked tkv mutant GSC
clones were clonally populated (total of 23 marked tkv GSC clones
examined; Fig. 6B).
tkv mutant GSCs divide slower than the wild type in the adult ovary
(Xie and Spradling, 1998
). The
mitotic potential of tkv mutant PGCs in the developing gonad is
probably also compromised. A slow division rate of mutant tkv PGCs
allows more time for adjacent wild-type PGCs to contact cap cells directly and
become GSCs. Therefore, a mutant tkv PGC cannot effectively populate
a niche by itself probably because of its proliferation defects. This result
demonstrates that tkv is essential for GSC expansion, thus allowing
one PGC to populate a niche. It also further suggests that dpp
signaling is required for this process.
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DISCUSSION |
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Positional information helps select stem cells from GSC
precursors
How stem cell identity is established initially remains elusive even in the
well-studied stem cell systems: Drosophila ovary and testis. In the
primitive female gonads before the pupal stage, PGCs appear to undergo
symmetric division to generate germ cells with the identical pre-stem cell
fate. Several studies suggest that GSCs were established at the early pupal
stage (Bhat and Schedl, 1997;
Song et al., 2002
). At the
early pupal stage, there are 136 germ cells on average in each gonad. The
adult ovary, which is composed of 12-16 ovarioles with two or three GSCs per
ovariole (average of 2.5), contains about 30 to 40 GSCs. Therefore, at the
most, 20-30% of PGCs in the early pupal gonad are recruited to niches and turn
into GSCs.
How is a particular germ cell selected and recruited to niches, and how
does it become a GSC? Positional information is known to be very important for
cell-fate determination in various developmental processes. In this study, we
have taken a developmental approach to investigate when key niche components
form, and how PGCs are subdivided into GSCs and differentiated germ cells. The
expression of bam is associated with germ cell differentiation in the
adult ovary (McKearin and Ohlstein,
1995; Ohlstein and McKearin,
1997
). Using bam expression as an indicator for germ cell
differentiation, we have shown that no PGCs in late third instar larval gonads
have differentiated. In early pupal gonads (about 0-4 hours after pupation),
all the PGCs that are not in contact with TFs/cap cells are differentiated;
therefore, the PGCs that contact newly formed cap cells remain
undifferentiated and become GSCs (Fig.
1F). Possibly, newly formed TFs/cap cells directly prevent the
most anterior PGCs from differentiation when an unknown developmental signal
triggers PGC differentiation around the larval-pupal transition stage. Our
study demonstrates that the stem cell fate of PGCs is determined by their
position, i.e. juxtaposition to TFs/cap cells.
Stem cells can originate from one GSC precursor by clonal
expansion
The next important question is how these anterior PGCs populate niches. In
this study, we show that the PGCs in contact with newly formed cap cells at
the early pupal stage divide more frequently than the rest of the PGCs. The
division patterns are very interesting: one division pattern generates two
daughters that are both in contact with cap cells; the other pattern generates
only one daughter that is in contact with cap cells. As in the adult ovary,
two daughters that are in contact with cap cells can both become GSCs. This is
verified by the observation that one marked PGC in the gonad at the late
third-instar larval stage can generate two or three GSCs in a niche. Our
results also indicate that the stem cells in a niche can come from multiple
PGCs. Whether GSCs in a niche come from one or multiple PGCs probably depends
on whether one or multiple PGCs directly contact cap cells within the
developing niche. If only one PGC contacts cap cells, it probably has an
opportunity to generate two or three germ cells that contact cap cells and
become GSCs. This study shows that newly formed niches do not simply recruit
existing PGCs and turn them into GSCs, but also stimulate PGCs to proliferate
and produce more GSCs.
The orientation of stem cell divisions seems to be very important for
self-renewal and expansion of a stem cell pool. This strategy does not seem
unique to the Drosophila ovarian GSCs. In the ventricular zone of the
developing mammalian brain, neural stem cells divide either parallel or
perpendicular to the ventricular surface
(Chenn and McConnell, 1995).
Normally, neural stem cells are in close contact with the surface of the
ventricular zone, and differentiated daughters move away from the ventricular
zone. It has been suggested that the stem cell divides along the ventricular
surface to give rise to two stem cells, while the perpendicular division
generates one stem cell and one differentiated neuronal cell. Controlling the
orientation of the stem cell division plane could be a general mechanism for
maintaining stem cell homeostasis and generating needed differentiated
cells.
BMP-like signaling stimulates clonal expansion of GSCs during early
ovarian development
The clonal expansion of GSCs in a niche clearly requires the newly
established stem cell to divide rapidly and generate a daughter that occupies
the same niche, which further prevents other neighboring precursor cells from
entering it. Consistent with this prediction, we observed that the anterior
row of germ cells at the early pupal stage was more mitotically active than
the rest of the germ cells based on the BrdU incorporation assay. dpp
is known to be important for maintaining GSCs and stimulating their division
in the adult ovary (Xie and Spradling,
1998). We show that dpp is expressed in TFs/cap cells and
other anterior somatic cells, and that PGCs close to cap cells are capable of
responding to dpp. Furthermore, overexpressing dpp promotes
PGC proliferation. To demonstrate the necessity of dpp signaling in
stimulating GSC clonal expansion, we have shown that a PGC mutant for
tkv, an essential dpp receptor, fails to clonally populate a
niche. All these results demonstrate that dpp is probably a signal
for stimulating GSC clonal expansion. However, we cannot rule out that other
BMP-like molecules, such as gbb-60A, could also play a similar role
because tkv could also be involved in other BMP-like signaling
pathways.
As in the adult ovary, hh is also expressed in terminal filaments
and cap cells in developing female gonads. Hh has recently been shown to play
a minor role in modulating GSC division
(King et al., 2001). Wingless
(Wg) protein is expressed in terminal filaments and cap cells (X. Song and
T.X., unpublished). Its expression in developing female gonads has not been
examined. Because wg, dpp and hh often work together to
regulate many developmental processes in Drosophila, it is possible
that hh and wg could also cooperate with dpp to
regulate PGC proliferation and modulate GSC clonal expansion in niches.
What signal(s) keeps anterior GSC precursors from
differentiation?
PGCs in the gonad do not show any signs of differentiation until the
larval-to-pupal transition. At the early pupal stage, only the PGCs in the
anterior row remain undifferentiated, but the rest have already
differentiated. It seems that a developmental signal(s) starts to appear and
then induces the differentiation of PGCs during the transition from larva to
pupa. Such a developmental signal could be mediated by a steroid-like hormone
ecdysone. Interestingly, during most of the third instar larval stage, the
ecdysteroid levels are very low but begin to rise and peak just before
pupation (Riddiford, 1993).
The ecdysteroid peak could be potentially responsible for the initial
differentiation of germ cells in the gonad of the larva ready for pupation. It
is also possible that the hormone is not a direct signal but controls the
production of the signal(s). Somehow, the signals from the anterior somatic
cells antagonize the differentiating signals and thus prevent the anterior row
of the PGCs from differentiation. One of the signals that prevent PGCs from
differentiation could be encoded by dpp. Dpp is known to prevent GSCs
from differentiation in the adult ovary
(Xie and Spradling, 1998
). In
this study, 2.5% of the marked tkv mutant PGCs and none of the marked
mad mutant PGCs before the third instar larval stage were recruited
to niches or were maintained as GSCs before adulthood. The failure of
tkv and mad mutant GSCs to be maintained in niches could be
explained by the role of dpp in preventing PGCs from differentiation.
It could also be explained by other possibilities, such as defects in the
formation of adherens junctions between cap cells and GSCs. Whether
dpp is a signal for maintaining the undifferentiated state of PGCs
during early ovarian development remains undetermined. Therefore, the signals
that maintain the undifferentiated state of PGCs from TFs/cap cells remain to
be identified.
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ACKNOWLEDGMENTS |
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