Transforming growth factor beta at 60A
Mutations have been isolated in the Drosophila melanogaster gene glass bottom boat (gbb), which encodes a TGF-ß signaling molecule (formerly referred to as 60A) with the highest sequence similarity to members of the bone morphogenetic protein (BMP) subgroup including vertebrate BMPs 5–8. Genetic analysis of both null and hypomorphic gbb alleles indicates that the gene is required in many developmental processes, including embryonic midgut morphogenesis, patterning of the larval cuticle, fat body morphology, and development and patterning of the imaginal discs. In the embryonic midgut, gbb is required for the formation of the anterior constriction and for maintenance of the homeotic gene Antennapedia in the visceral mesoderm. In addition, a requirement has been shown for gbb in the anterior and posterior cells of the underlying endoderm and in the formation and extension of the gastric caecae. gbb is required in all the imaginal discs for proper disc growth and for specification of veins in the wing and of macrochaete in the notum. Significantly, some of these tissues have been shown to also require the Drosophila BMP2/4 homolog decapentaplegic (dpp), while others do not. These results indicate that signaling by both gbb and dpp may contribute to the development of some tissues, while in others, gbb may signal independent of dpp (Wharton, 1999).
Defects in the embryonic midgut are observed in both dpp and gbb mutants, but each BMP appears to play a different role in midgut morphogenesis. gbb is required for the formation of the anterior midgut constriction, while dpp is required for the central constriction. Previous work has indicated that in the developing midgut, the localized visceral mesoderm expression of homeotic genes Antp, Ubx, and abd-A is required for the correct positioning of the anterior, central, and posterior constrictions, respectively. The homeotic genes have been shown to provide regional specification through their regulation of genes encoding secreted factors, such as dpp and wg, which subsequently act on the underlying midgut endoderm. dpp is activated directly by Ubx in a discrete band of cells in PS 7 of the visceral mesoderm from which Dpp is secreted, resulting in the induction of labial expression in underlying endodermal cells. It has been shown that Ubx expression is, in turn, maintained in the visceral mesoderm via a regulatory feedback loop through the action of dpp. In a manner similar to this regulation of Ubx by dpp, the expression of the homeotic gene Antp is regulated by gbb in the visceral mesoderm cells of PS 5 and 6. However, as is true of the regulation of dpp by Ubx, a reciprocal regulation of gbb by Antp is unlikely. The broad expression of gbb throughout the midgut indicates that gbb cannot be regulated exclusively by Antp (Wharton, 1999 and references).
gbb is expressed in both the visceral mesoderm and endoderm, and, as indicated by the regulation of Antp, gbb signaling is required in the visceral mesoderm. gbb signaling is also required in specific regions of the endoderm. The absence of gbb function eliminates the expression of the endodermal marker P-1 from cells in both the anterior and posterior midgut, as well as from cells in the ventriculus, the site from which the gastric caecae bud. The absence of P-1 staining in the primordia of the gastric caecae in gbb mutant embryos is consistent with gastric caecae defects observed in gbb mutant first instar larvae. It appears that although no gastric caecae are evident in stage 17 gbb mutant embryos, gastric caecae do form, albeit abnormally, by the end of the first larval instar. In summary, this analysis indicates, as is true for dpp, that gbb signaling is required in both the visceral mesoderm and endoderm of the Drosophila midgut. At this time, it is unknown which germ layer or layers serve as the source of the gbb signal (Wharton, 1999).
The specification of positional identity often arises from the localized expression of genes or factors controlling that particular process. It is of interest that although gbb does not exhibit a localized expression pattern, it is involved in regional specification of the midgut. The role of gbb in this process can be explained by two different models. In one model, gbb acts throughout the midgut, but with a partner that provides specific positional information. This partner or cofactor could be another BMP-type ligand or some other signaling component that is specifically localized. Given that the loss of gbb signaling has profound effects, for example, on the formation of the anterior midgut constriction, it would be predicted that a gbb partner would be localized to the anterior region of the midgut if this model were true. In the second model, gbb signaling does not specifically require a novel partner to provide positional information, but instead, cells within the midgut respond differently to varying levels of gbb and dpp signaling. This model is consistent with the paradigm that has been proposed for gbb and dpp signaling in the wing. Specification of different regions of the gut could result from the interpretation of different relative levels of gbb to dpp signaling. The total level of BMP signaling may be important, and the localized expression of dpp could provide a source of asymmetry necessary for the establishment of different positional information throughout the midgut. Low levels of signaling provided by gbb alone would specify anterior and posterior midgut vs. the high levels of signaling provided by both gbb and dpp that would specify the central domain of the midgut. Alternatively, differences in the responses elicited by a putative Gbb/Dpp heterodimer and Gbb and Dpp homodimers could be responsible for the assignment of different positional values. Other factors could certainly be involved in refining or elaborating the coarse pattern laid out by gbb and dpp. wg is an example of such a factor, as it has been shown that both wg and dpp are required to activate the expression of certain target genes in several tissues, At this time, it is not possible to distinguish between these two simple models, but these models provide a framework within which to investigate further the contribution of multiple BMP signaling to a specific developmental process: midgut morphogenesis (Wharton, 1999 and references).
In addition to the gbb mutant phenotypes that resemble dpp mutant phenotypes or those that affect tissues also affected by dpp mutations, several phenotypes have been identified that have not been observed in dpp mutants. Defects in the development of the telson and fat body of the larva have not been reported as aspects of dpp mutants, suggesting that in some developmental processes, gbb may function independently of dpp. It is interesting to note that mutations in the Drosophila BMP signaling components Mad, Medea, and sax can produce a clear larva phenotype. Mad and Medea encode Smad proteins shown to mediate dpp signaling in the midgut and the wing imaginal disc. It is possible that in the formation of the telson and the fat body, gbb may be the only BMP signal mediated by Mad, Medea, and sax: gbb is known to signal independent of dpp in these tissues. Alternatively, the earlier requirement for dpp in dorsal/ventral patterning of the embryo may have precluded the identification of dpp involvement in fat body or telson differentiation, and in fact, both gbb and dpp signaling are required for proper development of these structures. Without the ability to bypass the early requirement for dpp, it is not possible at this time to distinguish between these two possibilities (Wharton, 1999 and references).
The analysis of gbb alleles has also identified a requirement for gbb in the proper specification or positioning of bristles on the notum of the adult fly. A reduction in gbb activity results in the formation of ectopic macrochaete, most frequently on the scutellum. Such a phenotype has not previously been reported for dpp mutants. However, a recent report describing the ubiquitous activation of Tkv, a proposed Dpp receptor, results in ectopic macrochaete formation within the dorsolateral region of the notum. In this case, ectopic macrochaete formation results from the proposed activation of Dpp signaling via the Tkv receptor. In contrast, ectopic macrochaete has been observed with a reduction of gbb function. These opposite phenotypes could reflect a fundamental difference in the role of gbb signaling vs. dpp signaling in the formation or patterning of sensory mother cells, the precursor cells of the macrochaete. Furthermore, the appearance of ectopic macrochaete in the dorsocentral vs. scutellar regions of the notum may reflect a different positional or spatial requirement for dpp vs. gbb. Further analysis will reveal whether the requirement for gbb in scutellar macrochaete formation is independent of the potential role for dpp in the dorsocentral region (Wharton, 1999 and references).
The phenotypic analysis indicates that gbb and dpp participate in many of the same developmental processes; in some tissues the functions of gbb and dpp appear to be the same or very similar, while in others, their functions appear to be distinct. It is clear that while both gbb and dpp signaling contribute to the proper formation of the embryonic midgut and to patterning of the wing veins in the adult, the relative contribution of each BMP must be different. It is possible that overall, gbb and dpp participate in the development of certain tissues, and this could be accomplished by both cooperative or synergistic interactions and/or antagonistic interactions. As the different mutant phenotypes indicate, the mechanism by which gbb and dpp signaling each contribute to a developmental process must differ depending on the tissue. Understanding the different mechanisms by which these signals are sent and how these differences are regulated in Drosophila will provide significant insight into signaling by multiple TGF-ß/BMP ligands in both invertebrates and vertebrates (Wharton, 1999).
Formation of the longitudinal veins (LVs) of the Drosophila wing involves the interplay among Dpp, Egf and Notch pathways. Formation of crossveins (CVs: see Derivatives of the wing disc) present a paradoxical problem. As shown both morphologically and using molecular markers,
the definitive CVs are not formed until long after the initial
specification of the LVs. The CVs therefore must form within
territory that has already been specified as intervein. The CVs
must also interconnect with existing LVs at a time when the
Delta expressed by the LVs is thought to inhibit vein formation
in adjacent cells.
Mechanisms must exist that override both intervein
specification and the lateral inhibition of veins, allowing the
formation of continuous, interconnected vein tissue.
BMP-like signaling plays a special role
in the formation of the CVs from within intervein territory.
BMP-like signals also help maintain the connections between
the LVs and the margin of the wing. crossveinless 2 (cv-2) is a critical factor in
these processes, as it is expressed more highly in the CVs and
the ends of the LVs and is required for the high levels of BMP-like
signaling observed in these regions (Conley, 2000). The cv-2 mutation was first identified by Benedetto Nicoletti in 1962 (FlyBase: Cv-2 site). The structure of the
Cv-2 protein strongly suggests that these effects are direct, and
that Cv-2 is a novel player in the BMP-like signaling pathway (Conley, 2000).
Both Dpp and Gbb vein signals are mediated largely
by the type I receptor Thickveins, rather than the
alternate type I receptor Saxophone. Cells lacking Tkv
do not form veins, but removal of Sax does not reliably remove veins.
However, not all veins are equally sensitive to reductions in
Dpp and Gbb signaling. The hypomorphic gbb4 mutation
shows complete loss of the cross veins (CVs), but only slight
loss of the ends of the LVs. Sog encodes
a Chordin-like molecule that inhibits BMP-like signaling; both
Sog and Chordin are thought to bind to and sequester ligands,
preventing the activation of receptors. Overexpressing Sog
in the wing specifically blocks formation of the CVs and the
ends of the LVs. The secreted Tolloid
proteases, similar to vertebrate BMP1s, can increase BMP
signaling by cleaving and inactivating Chordin or Sog. Loss of tolkin (also known as tolloid-related) blocks
formation of the CVs and the tips of the LVs. Overexpressing a dominant negative
form of Sax again induces a similar phenotype (Conley, 2000 and references therein).
Such phenotypes are very reminiscent of the crossveinless
class of mutations in Drosophila (reviewed in Garcia-Bellido, 1992). Strong reductions
in crossveinless 2 (cv-2) function have been shown to remove the posterior CV
(PCV), the anterior CV (ACV), and the ends of the LVs. However, despite the possibility that the crossveinless
genes encode novel players in BMP-like signaling, none have
been characterized and the sensitivity of CVs to BMP-like
signaling has not been explained. Evidence is presented that cv-2 encodes a novel member
of the BMP-like signaling pathway, expressed in and required
for high levels of BMP-like signaling in the developing cross
veins. The Cv-2 protein contains five cysteine-rich domains
similar to those known to bind BMP-like ligands, strongly
suggesting that Cv-2 directly modulates Dpp or Gbb activity (Conley, 2000 and references therein).
dpp and gbb mutations both disrupt CV
formation. Weak cv-2 alleles are strengthened
by dpp and gbb loss-of-function mutations. cv-2225-3/cv-23511
flies never lack the entire PCV, but 50% of gbb 4 cv-2225-3/cv-2
3511 flies lack the entire PCV. Similarly, cv-23511/Df(2R)Pu-D17
only rarely disrupt the ACV, but dppd6 cv-23511/Df(2R)Pu-D17 commonly does. However, cv-2
cannot dominantly enhance earlier dpp-dependent patterning in
the wings: dppd5 Df(2R)Pu-D17 /dpphr4 wings look no worse
than dppd5/dpphr4 wings.
To provide a more direct link between cv-2 and Dpp and Gbb
signaling, Mad activation was examined in mutant pupal wings.
In cv-21 adults, the PCV is more reliably disrupted than the
ACV; the anti-p-Mad staining normally found near the PCV in
19, 22, 26 and 36 hours after pupariation wings is lost or disrupted in cv-21 homozygotes, as is the reduction of anti-DSRF
in the PCV. In adults of the stronger allelic combination cv-21/Df(2R)Pu-D17, the ACV is also often lost along with the
ends of some of the LVs. Interestingly, no
disruption of the ACV or LV anti-p-Mad staining cv-21/Df(2R)Pu-D17 pupal wings is detected at 21 or 25 hours after pupariation;
only at 36 hours after pupariation is staining lost from the ACV.
This indicates that cv-2 is required not only to initiate Mad
activity in the PCV, but also to maintain that activity in the ACV (Conley, 2000).
The Drosophila BMP5/6/7/8 homolog, glass bottom boat (gbb), has been shown to be involved in proliferation and vein patterning in the wing disc. To better understand the roles for gbb in wing development, as well as its relationship with decapentaplegic, clonal analysis was used to define the functional foci of gbb during wing development. gbb has both local and long-range functions in the disc that coincide both spatially and functionally with the established functions of dpp, suggesting that both BMPs contribute to the same processes during wing development. Indeed, comparison of the mutant phenotypes of dpp and gbb hypomorphs and null clones shows that both BMPs act locally along the longitudinal and cross veins to affect the process of vein promotion during pupal development, and long-range from a single focus along the A/P compartment boundary to affect the processes of disc proliferation and vein specification during larval development. Moreover, duplications of dpp are able to rescue many of the phenotypes associated with gbb mutants and clones, indicating that the functions of gbb are at least partially redundant with those of dpp. While this relationship is similar to that described for dpp and the BMP screw (scw) in the embryo, the mechanisms underlying both local and long-range functions of gbb and dpp in the wing are different. For the local foci, gbb function is confined to the regions of the veins that require the highest levels of dpp signaling, suggesting that gbb acts to augment dpp signaling in the same way as scw is proposed to do in the embryo. However, unlike scw-dependent signals in the embryo, these gbb signals are not transduced by the Type I receptor saxophone (sax), thus, the cooperativity between gbb and dpp is not achieved by signaling through distinct receptor complexes. For the long-range focus along the A/P compartment boundary, gbb function does not appear to affect the high point of the dpp gradient, but, rather, appears to be required for low points, which is the reciprocal of the relationship between dpp and scw in the embryo. Moreover, these functions of gbb also do not require the Type I receptor sax. Given these results, it is concluded that the relationships between gbb and dpp in the wing disc represent novel paradigms for how multiple BMP ligands signal during development, and that signaling by multiple BMPs involves a variety of different inter-ligand relationships that depend on the developmental context in which they act (Ray, 2001).
gbb has two distinct types of functions: local and long range. The local foci are confined to the posterior compartment, and affect the promotion of the posterior cross-vein (PCV) and the distal tips of the longitudinal veins L4 and L5. The long-range focus lies in the anterior compartment comprising a broad band of cells along the A/P compartment boundary and affects disc proliferation and the specification of L5. These gbb foci are coincident with the foci for dpp in the disc, and many of the phenotypes associated with the gbb clones are rescued by additional copies of the dpp locus. Thus, gbb and dpp contribute to the same functions in the disc and gbb functions are to some extent redundant with those of dpp. Comparison of the foci and phenotypes of gbb and dpp mutants and clones indicates that the relationship between gbb and dpp is different for different functions. For promotion of distal tips of L4 and L5, gbb function is restricted to those areas that require the highest levels of dpp signaling, and since these phenotypes can be rescued with additional copies of dpp, it is concluded that gbb is required to augment the levels of dpp signaling. For promotion of the PCV, the case is not so clear. Both gbb and dpp are required for PCV promotion. However, since dpp duplications do not rescue this phenotype, it is possible that gbb and dpp act independently or that the contribution of gbb to this process is sufficiently great that it cannot be compensated for by the additional doses of dpp. The requirement for gbb in the specification of L5 is not consistent with an augmentation of dpp signaling, since gbb mutants and clones do not affect structures specified by the high point of the dpp gradient. Rather, gbb clones affect structures far from the source along the A/P compartment boundary, suggesting that gbb signaling contributes to the low levels of BMP signaling at the extremes of the gradient (Ray, 2001).
Mutant phenotypes are observed only in gbb clones when the mutant tissue encompasses the entirety of the focus on both the dorsal and ventral surfaces of the wing. For example, clones that occupy the dorsal-anterior quadrant of the wing exhibit no defects in the patterning or size of the wing, while clones that occupy both the dorsal-anterior and ventral-anterior quadrants affect both these aspects of wing development. One explanation for this phenomenon is that Gbb exhibits long-range non-autonomy in the disc, and, in fact, there is some evidence for this, since it has been found that small patches of wild-type cells along the A/P compartment boundary in the context of a large mutant clone are able to rescue loss of L5 completely in the posterior compartment. However, gbb clearly does not act in a broadly non-autonomous fashion in all of its functions: gbb clones that cover the PCV or distal L5 exhibit vein defects that respect the clone boundaries, indicating that the presumptive vein cells within the clone cannot be rescued by the wild-type Gbb present in the adjacent cells. For these functions, the 'rescue' observed in single-sided clones implies pattern regulation occurring between the two wing surfaces. Indeed, it has long been asserted that there are signaling events between the dorsal and ventral surfaces of the wing (as have been shown for several genes) whereby loss of veins on one surface can be compensated for by the wild-type pattern in the opposing surface. The requirement for dorsal-ventral overlap observed with gbb mutant clones is indicative of such a signaling mechanism, and given these results, as well as those from previous studies that have shown a requirement for dorsal-ventral overlap in clones of dpp and sog, it is plausible that the BMPs themselves might be responsible for mediating these signaling processes (Ray, 2001).
Perhaps the most striking result from clonal analysis is that the requirements for gbb in the wing disc are localized even though the gene is widely expressed. This result implies that Gbb activity is in some way restricted post-transcriptionally. Two models seem the most likely to account for this effect. First, since it has been shown that all gbb foci are coincident with sites of dpp expression in the disc, it is possible that Gbb and Dpp form heterodimers, and that Gbb is only active in this form. Heterodimer formation has been documented for a number of different TGFß superfamily members, and in some cases heterodimers and homodimers have been shown to have distinct properties. For example, heterodimers of BMP2 or BMP4 and BMP7 are much more potent in the induction of ventral mesoderm and bone induction than their respective homodimers. Activins and Inhibins illustrate a different relationship: the homodimeric Activins having the opposite biological effects of the heteromeric Inhibins (Ray, 2001).
An alternative model is that the restriction of gbb function in the disc is achieved through local activation of Gbb homodimers, which may be achieved by specific agonists expressed within the foci or antagonists expressed everywhere else. Possible agonists include the Drosophila BMP-1 homologs tolloid and tolkin, or Drosophila homologs of the subtilisin-like proprotein convertases or furins, that are thought to be involved in the cleavage of BMP pro-proteins into the active ligand. In addition, the recently characterized secreted protein crossveinless 2 (cv-2) may act as an agonist of BMP signaling specifically in the presumptive crossveins. The antagonist sog is a likely candidate for restricting BMP activity during pupal development (i.e. for vein promotion functions) as it has been shown to be expressed in all intervein cells at this time. Moreover, there is some evidence that sog function in the wing may specifically antagonize gbb, and thus may very well account for the restriction of gbb function to the presumptive veins (Ray, 2001).
Clonal analysis has identified four processes that require gbb during wing development, disc proliferation, specification of the L5 vein territory, promotion of the PCV and promotion of the longitudinal veins L4 and L5. Based on the criteria of comparisons of gbb clone phenotypes with dpp and sax phenotypes, the ability for the gbb mutant phenotypes to be rescued by additional copies of dpp, and the spatial requirements for gbb during wing development, it is clear that each of these functions employs a different relationship between dpp and gbb, and each of these relationships is distinct from that which has been established for dpp and scw in the embryo (Ray, 2001).
BMP signaling is essential for promoting self-renewal of mouse embryonic stem cells and Drosophila germline stem cells and for repressing stem cell proliferation in the mouse intestine and skin. However, it remains unknown whether BMP signaling can promote self-renewal of adult somatic stem cells. In this study, BMP signaling is shown to be necessary and sufficient for promoting self-renewal and proliferation of somatic stem cells (SSCs) in the Drosophila ovary. BMP signaling, via the ligand Glass bottom boat, is required in SSCs to directly control their maintenance and division, but is dispensable for proliferation of their differentiated progeny. Furthermore, BMP signaling is required to control SSC self-renewal, but not survival. Moreover, constitutive BMP signaling prolongs the SSC lifespan. Therefore, this study clearly demonstrates that BMP signaling directly promotes SSC self-renewal and proliferation in the Drosophila ovary. This work further suggests that BMP signaling could promote self-renewal of adult stem cells in other systems (Kirilly, 2005)
FLP-mediated FRT recombination has revolutionized studies on diverse developmental processes in Drosophila. The mosaic clones marked by loss of armadillo (arm)-lacZ or ubiquitin (ubi)-GFP are routinely used to study Drosophila oogenesis. Two positive labeling methods, the tubulin-lacZ positive labeling system and the gal80-based mosaic analysis with a repressible cell marker (MARCM), have been developed to facilitate visualization of marked cells. The lacZ-positive labeling system is effective for identification of marked cells, but it is not ideal for manipulating gene function, while stable GAL80 protein may not allow rapid visualization of marked cells after one or two divisions due to its persistence. A new positively marked mosaic lineage (PMML) method has been developed to positively mark cells and allow for rapid expression of the UAS-GFP marker and any other UAS construct in the marked cells by using a combination of the GAL4-UAS and FLP-FRT systems. This PMML system uses the heat shock-inducible FLP to reconstitute a functional actin5C-gal4 gene from two complementary inactive alleles, actin5C FRT52B and FRT52B gal4. The actin5C-gal4 gene drives GFP expression to mark cells and can also activate or knock down gene function by using UAS constructs in the marked cells (Kirilly, 2005)
To test whether PMML is also suitable for marking SSCs and assisting in SSC identification in the Drosophila ovary, ovaries were immunostained with anti-GFP and anti-Fasciclin III (Fas3) antibodies 1 week after clone induction (ACI). Fas3 is expressed in SSCs at low levels and in differentiated follicle cell progenitor cells at higher levels. It takes about 4-5 days for transiently labeled GFP-positive follicle cells to completely exit the germarium. One week ACI, a typical GFP-positive SSC clone was easily observed with the GFP-marked follicle cells present in regions 2b and 3 of the germarium and in egg chambers. The marked SSC could be identified by its location (the GFP-positive somatic cell at the 2a/2b junction), low Fas3 expression, and the presence of GFP-marked follicle cells in the germarium and/or in the egg chambers. The GFP-marked inner germarial sheath (IGS) cells could also be readily identified by their location (the germarial regions 1 and 2a), the absence of marked differentiated follicle cells in the same ovarioles, and also the absence of Fas3 expression, since the IGS descendants do not pass beyond the 2a/2b junction. Therefore, this system can be applied effectively for labeling SSCs and their progeny and for further studying the function of any gene in the marked SSCs and their progeny by overexpression (Kirilly, 2005).
This study shows that SSCs in the adult ovary are capable of responding to BMP signaling. Genetic mosaic analyses demonstrate that known BMP downstream components are also required for SSC self-renewal, but not survival. Hyperactive BMP signaling enhances SSC self-renewal capacity. Glass bottom boat (Gbb) is essential for controlling SSC maintenance, at least in the GSC niche. Furthermore, BMP signaling appears to be specific to stem cells, since follicle cells mutant for BMP-specific downstream components proliferate and differentiate normally. In addition to participation in BMP signaling, Medea (Med) is likely involved in other TGF-β-like pathway(s) to control proliferation and size of differentiated follicle cells. The results from this study led to the proposal of a working model that Gbb perhaps as well as Dpp from neighboring somatic cells function as stem cell growth factors in vivo for promoting self-renewal of ovarian SSCs (Kirilly, 2005).
gbb and dpp are expressed in cap cells, inner germarial sheath (IGS) cells, and follicle cells. SSCs are located in the middle of the germarium and are likely exposed to both BMPs, since both Dpp and Gbb are diffusible molecules. gbb mutants exhibit severe SSC/follicle cell proliferation defects and SSC loss. Furthermore, SSCs mutant for BMP downstream components such as tkv, punt, and mad are lost faster and divide slower than wild-type ones. Although dpp mutants show much weaker mutant defects, it is still possible that it plays as important a role as does gbb, since only weak dpp mutations could be used for studying the regulation of adult SSCs due to its stringent requirements during early development. Therefore, these findings support the idea that Gbb, perhaps together with Dpp, controls SSC self-renewal and division. Studies on GSCs in the Drosophila ovary have shown that BMPs control GSC self-renewal by directly repressing transcription of differentiation-promoting genes such as bam. Possibly, BMP signaling also represses differentiation-promoting genes and thereby maintains SSC self-renewal. Meanwhile, BMP signaling could also positively regulate other genes that are important for maintaining the undifferentiated state of SSCs. This study also shows that BMP signaling also promotes SSC division. It has been shown that BMP signaling promotes GSC division. In order to better understand how BMP signaling controls SSC self-renewal and division, it is critical to identify the BMP target genes in SSCs, that are either repressed or activated by BMP signaling (Kirilly, 2005)
This study also shows that tkv is a major type I BMP receptor for controlling SSC self-renewal in the Drosophila ovary. The SSCs mutant for sax4, a null allele of sax, behave close to normal wild-type ones, while the SSCs mutant for a strong tkv allele, tkv8, are lost rapidly, indicating that Tkv is a major functional receptor to control SSC self-renewal. Given the evidence that gbb signaling is essential for maintaining SSCs, this study strongly supports the idea that Gbb signals mainly through Tkv to control SSC self-renewal in the Drosophila ovary. A recent study on Drosophila spermatogenesis also suggests that Gbb signaling primarily functions through Tkv, but not Sax. In the Drosophila testis, gbb and tkv are both essential for maintaining GSCs, but sax is not. Although one study on dominant-negative tkv and sax receptors suggests that dpp and gbb signal preferentially through tkv and sax, respectively, another more recent study has shown that both dpp and gbb use tkv, but not sax, control the process of vein promotion during pupal development and disc proliferation and vein specification during larval development. Taken together, the results from this study and the previous studies indicate that Gbb can use Tkv as a major receptor for its signal transduction in Drosophila (Kirilly, 2005).
Although Gbb/BMP signaling plays a critical role in controlling SSC self-renewal and division, it appears that it is dispensable for SSC survival, follicle cell proliferation, and cell size control. For example, expression of the baculovirus antiapoptotic gene p35 could not rescue the mutant punt SSC loss; the follicle cell clones mutant for strong tkv and mad alleles, tkv8 and mad12, proliferate normally, and the sizes of the mutant follicle cells are quite normal. In contrast, p35 expression can rescue the Med26 SSC loss to the levels of the mutant punt, tkv, and mad mutant SSC loss. The partial rescue indicates that Med is required for SSC survival in a BMP-independent pathway. The Med mutant follicle cell clones proliferate slower than wild-type, and the size of follicle cells is also smaller than that of wild-type, suggesting that Med is required for follicle cell proliferation and growth. Since BMP signaling is not involved in the control of SSC survival, follicle cell proliferation, and growth, these findings further suggest that Med must participate in other TGF-β-like pathways controlling these processes. In mammalian systems, SMAD4 has been shown to be a common SMAD for all TGF-β-like signaling pathways, including TGF-β, Activin, and BMP. A likely candidate TGF-β-like signaling pathway includes Activin and TGF-β. Activin and TGF-β molecules exist in Drosophila. Activin-like signaling has been shown to be involved in regulating growth control and neuronal remodeling. However, the role of TGF-β signaling in Drosophila remains a mystery. It could not be completely ruled out, however, that Med is involved in other signaling pathways unrelated to TGF-β-like pathways to control SSC survival, follicle cell proliferation, and growth. In the future, it is very important to figure out which pathway Med takes part in for controlling SSC survival, follicle cell proliferation, and growth control (Kirilly, 2005).
In a variety of systems, stem cells have been proposed to be regulated by signals from niches. SSCs are anchored to the posterior group of IGS cells through DE-cadherin-mediated cell adhesion. Elimination of the anchorage leads to rapid SSC loss, suggesting that the posterior IGS cells function as a SSC niche. This study shows that gbb is expressed in the somatic cells, including IGS cells and follicle cells, and plays an important role in maintaining SSCs. Hh and Wg are expressed in the cap cells and play essential roles in controlling SSC self-renewal, suggesting that the SSC niche is composed of IGS cells and cap cells. In Drosophila imaginal development, these three pathways often regulate one another to control patterning, cell proliferation, and differentiation. In the Drosophila ovary, disruption of Hh, Wg, and BMP signaling cascades causes rapid SSC loss, while hyperactive signaling results in abnormal proliferation and differentiation of SSC progeny. Interestingly, their downstream transcriptional factors are also required for controlling SSC maintenance, suggesting that integration of these pathways likely takes place at or after transcription of their target genes. This study has shown that hyperactive BMP signaling can substitute for Wg signaling, but not Hh signaling, in controlling SSC self-renewal. However, it still remains unclear how hyperactive BMP signaling bypasses Wg signaling in SSCs. An important task in the future is to define their target genes in SSCs and to further figure out how these three signal transduction pathways interact with each other to control expression of these target genes (Kirilly, 2005).
In mammals, Shh, Wnt, and BMP pathways have been shown to regulate stem cell behavior directly or indirectly. BMP signaling directly represses activities of stem cells in the intestine and the hair follicle and promotes self-renewal of ES cells and spermatogonial stem cells. BMP signaling can also indirectly regulate haematopoeitic stem cells (HSCs) by controlling niche size. Wnt signaling has been shown to control self-renewal of HSCs, ES cells, intestinal stem cells, and possibly hair follicle stem cells. Shh signaling is required for proliferation of stem cells/progenitor cells in the lung airway. Studies from Drosophila and mice have shown that different stem cell types may utilize a combination of different growth factors to control their self-renewal, proliferation, and differentiation. Interestingly, Wnt and BMP signaling pathways promote ES self-renewal in mice and ovarian SSC self-renewal in Drosophila. Future studies of how different signaling pathways are integrated in Drosophila ovarian SSCs may also shed light on how these same pathways control stem cell self-renewal in mammals (Kirilly, 2005).
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Transforming growth factor beta at 60A:
Biological Overview
| Evolutionary Homologs
| Regulation
| Developmental Biology
| References
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