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Molecular Regulation of Germline Development and Organogenesis
Summary: Judith Kimble studies controls of growth, patterning, and morphogenesis during animal development.
Animal development is a remarkable feat of biological regulation. How are cells, tissues, and organs governed to reach their proper size, shape, and pattern? We have embarked on an in-depth analysis of a single organ, the gonad, to learn how controls of growth, patterning, and morphogenesis are integrated to generate a complex biological structure. Using the nematode Caenorhabditis elegans, we have uncovered genes, proteins, and pathways fundamental to the development of all animals, including humans.
The Control of Germline Stem Cells by Wnt and Notch Signaling
Stem cells are responsible for replenishing tissues as differentiated cells are depleted. Defective stem cell controls can lead to serious problems in humans, including cancer. In addition, reduced stem cell function appears to be a major factor in aging. Our laboratory has focused on the cellular and molecular mechanisms that control germline stem cells (GSCs). Our analyses of nematode GSCs have uncovered fundamental mechanisms of stem cell control that are broadly conserved.
Stem cells are established and maintained within a stem cell "niche," a microenvironment specialized for nurturing stem cells. For C. elegans GSCs, a single somatic cell, called the distal tip cell, is responsible for creating the stem cell niche (Figure 1). The distal tip cell provides a particularly simple model for understanding both niche development and function. We have found that the distal tip cell arises from an asymmetric cell division during early postembryonic development, and that its fate is specified by Wnt signaling and a conserved homeodomain transcription factor, CEH-22, which is the nematode homolog of Drosophila tinman and vertebrate Nkx2.5. Moreover, we found that Notch signaling from the distal tip cell controls GSC maintenance. In mammals, both Wnt and Notch signaling have been implicated in stem cell controls, and in humans, Notch signaling defects are associated with several diseases, including leukemia. The well-defined roles of these two pathways in controlling C. elegans GSCs may provide insight into how they work to control vertebrate stem cells.
An RNA Regulatory Network Controls Germline Stem Cells
Notch signaling controls GSCs, at least in part, by regulating the expression of FBF, which is a PUF (Pumilio and FBF) RNA-binding protein. FBF is a key regulator of stem cell maintenance: mutant germlines that lack FBF are no longer capable of maintaining GSCs. Pumilio similarly promotes GSC maintenance in the Drosophila ovary, and the human PUM2 homolog localizes to GSCs in the testis. Therefore, PUF RNA-binding proteins appear to be an ancient mechanism of stem cell control.
FBF controls GSC maintenance by repressing regulators designed to drive germ cells from the stem cell state toward differentiation. Specifically, FBF binds regulatory elements within the 3'-untranslated region of target mRNAs and down-regulates their translation or stability. (These molecular studies of FBF regulation have been done in collaboration with Marvin Wickens [University of Wisconsin–Madison].) Over the past few years, we have delineated a regulatory network that controls the switch between stem cell maintenance and differentiation. That network employs a battery of conserved RNA regulators [FBF, GLD-1/quaking, GLD-2/poly(A) polymerase, GLD-3/Bicaudal-C, FOG-1/CPEB, FOG-3/Tob, NOS-3/Nanos], as well as the LIP-1 dual-specificity phosphatase. The network is robust, yet plastic: no single component is essential for GSC maintenance, but alterations of single components can contract or expand the number of mitotic germ cells and also modulate the kinetics of the switch from mitotic cycling to differentiation.
Integration of the Mitosis/Meiosis and Sperm/Oocyte Decisions
Throughout the animal kingdom, somatic signals tell germ cells whether to differentiate as sperm or oocyte. Our work has focused on how germ cells respond to the somatic signal and make the sperm/oocyte decision. Some years ago, we identified FOG-1 and FOG-3, two regulators that specify germ cells as sperm. Upon loss of either FOG-1 or FOG-3, germ cells sexually transform into oocytes (hence the name FOG, for feminization of germline). FOG-1 is a homolog of the CPEB (cytoplasmic polyadenylation binding) RNA-binding protein, whereas FOG-3 is related to vertebrate Tob/Btg antiproliferative proteins. More recently, we discovered that FOG-1 has dose-dependent effects: abundant levels of FOG-1 promote the sperm fate; lower levels stimulate mitotic divisions. The fog-1 mRNA is a direct target of FBF repression. Therefore, FBF promotes continued mitoses within the niche, at least in part, by keeping FOG-1 levels low; then, as germ cells leave the niche and escape FBF repression, FOG-1 levels rise and direct sperm differentiation. In frogs, CPEB also promotes mitotic divisions at low levels but meiotic maturation at higher levels. This parallel suggests that establishment of a FOG-1 gradient in the nematode germline may be a widely used mechanism for patterning growth and differentiation.
Wnt Signaling, a Two-Pronged Control of the β-Catenin:TCF Ratio
A few years ago, we discovered that the sys-1 gene encodes the major C. elegans β-catenin transcriptional coactivator for the POP-1/TCF DNA-binding protein. Over the past year, we found that SYS-1/β-catenin and POP-1/TCF are reciprocally asymmetric in daughters of asymmetric cell divisions, including that of the EMS blastomere in early embryos as well as later embryonic and postembryonic divisions. In addition, we used both in vitro and in vivo assays to show that the SYS-1:POP-1 ratio is crucial for transcriptional activation or repression. Our work, together with that of others, delineates a two-pronged Wnt signaling pathway with one regulatory branch controlling POP-1/TCF abundance and a different branch controlling SYS-1/β-catenin abundance. A future challenge will be to learn whether this two-pronged Wnt signaling pathway, identified in nematodes, plays a similar role in vertebrates.
Regulation of Organ Polarity and Sexual Dimorphism
One unique feature of gonadogenesis is its sexual dimorphism. Female and male gonads are morphologically and functionally distinct, even though they have a common origin in the embryo. Our work on gonadogenesis is aimed not only at fundamental mechanisms of organogenesis but also at how those mechanisms can be modified to generate diverse structures.
The gonad begins as a cluster of cells known as a primordium. In C. elegans, the gonad primordium contains only two somatic gonadal progenitor cells (SGPs), which produce somatic structures (e.g., uterus, vas deferens). The first sign of sexual dimorphism during gonad development occurs at the first SGP cell division: that division is asymmetric in both sexes, but its daughter cells adopt sex-specific fates and sizes. Over the past few years, we have demonstrated that Wnt signaling controls cell fate asymmetry in both sexes, whereas a forkhead transcription factor, called FKH-6 (discovered in collaboration with David Zarkower [University of Minnesota]), specifies cell size asymmetry typical in males.
Most recently, we learned that a conserved cell cycle regulator, known as cyclin D, is also a key regulator of the SGP division. When cyclin D levels are lowered, the SGP always divides in a female (or more correctly hermaphrodite) mode, regardless of the sex of the animal, and it often generates daughter cells of equivalent fate. The molecular explanation is that fkh-6 expression is silenced and Wnt signaling components are symmetrically distributed. The function of cyclin D in metazoa is poorly understood. Our findings with the SGP division suggest that cyclin D may be specialized to coordinate the activities of multiple regulators with the cell cycle at pivotal junctures of development.
Last updated: July 19, 2007
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