Stephen DiNardo

arrow encodes an LDL-receptor-related protein essential for Wingless signalling.

The Wnt family of secreted molecules functions in cell-fate determination and morphogenesis during development in both vertebrates and invertebrates (reviewed in ref. 1). Drosophila Wingless is a founding member of this family, and many components of its signal transduction cascade have been identified, including the Frizzled class of receptor. But the mechanism by which the Wingless signal is received and transduced across the membrane is not completely understood. Here we describe a gene that is necessary for all Wingless signalling events in Drosophila. We show that arrow gene function is essential in cells receiving Wingless input and that it acts upstream of Dishevelled. arrow encodes a single-pass transmembrane protein, indicating that it may be part of a receptor complex with Frizzled class proteins. Arrow is a low-density lipoprotein (LDL)-receptor-related protein (LRP), strikingly homologous to murine and human LRP5 and LRP6. Thus, our data suggests a new and conserved function for this LRP subfamily in Wingless/Wnt signal reception.

Escherichia coli DNA topoisomerase I mutants have compensatory mutations in DNA gyrase genes.

Escherichia coli deletion mutants lacking DNA topoisomerase I have been identified previously and shown to grow at a normal rate. We show that such strains grow normally only because of spontaneously arising mutations that compensate for the topoisomerase I defect. Several of these compensatory mutations have been found to map at or near the genes encoding DNA gyrase, gyrA and gyrB. DNA gyrase assays of crude extracts show that strains carrying the mutations have lower gyrase activity. Thus the mutations are in the gyrase structural genes or in nearby regulatory sequences. These results, in conjunction with DNA supercoiling measurements of others, indicate that in vivo DNA superhelicity is a result of a balance between topoisomerase I and gyrase activities. An excess of negative supercoils due to an absence of topoisomerase I is deleterious to the cell, but a moderate gyrase deficiency is not harmful.

Wg/Wnt Signal Can Be Transmitted through Arrow/LRP5,6 and Axin Independently of Zw3/Gsk3β Activity

Activation of the Wnt signaling cascade provides key signals during development and in disease. Here we provide evidence, by designing a Wnt receptor with ligand-independent signaling activity, that physical proximity of Arrow (LRP) to the Wnt receptor Frizzled-2 triggers the intracellular signaling cascade. We have uncovered a branch of the Wnt pathway in which Armadillo activity is regulated concomitantly with the levels of Axin protein. The intracellular pathway bypasses Gsk3beta/Zw3, the kinase normally required for controlling beta-catenin/Armadillo levels, suggesting that modulated degradation of Armadillo is not required for Wnt signaling. We propose that Arrow (LRP) recruits Axin to the membrane, and that this interaction leads to Axin degradation. As a consequence, Armadillo is no longer bound by Axin, resulting in nuclear signaling by Armadillo.

Drosophila hedgehog acts as a morphogen in cellular patterning

The patterning of cell types in embryogenesis is specified by signals emanating from specialized organizer regions. We demonstrate that engrailed-expressing cells in the Drosophila epidermis have organizer properties. These cells influence the pattern of cell type differentiation across the segment. We show that this function is mediated by the hedgehog (hh) gene. The results of modulating the levels of hh in the embryo suggest that hh acts as a morphogen, specifying distinct cell fates by a concentration-dependent mechanism. We present a model that integrates the role of hh with that of the wingless signal in establishing the segmental array of cell type diversity.

Somatic control over the germline stem cell lineage during Drosophila spermatogenesis

Stem cells divide both to produce new stem cells and to generate daughter cells that can differentiate. The underlying mechanisms are not well understood, but conceptually are of two kinds. Intrinsic mechanisms may control the unequal partitioning of determinants leading to asymmetric cell divisions that yield one stem cell and one differentiated daughter cell. Alternatively, extrinsic mechanisms, involving stromal cell signals, could cause daughter cells that remain in their proper niche to stay stem cells, whereas daughter cells that leave this niche differentiate. Here we use Drosophila spermatogenesis as a model stem cell system to show that there are excess stem cells and gonialblasts in testes that are deficient for Raf activity. In addition, the germline stem cell population remains active for a longer fraction of lifespan than in wild type. Finally, raf is required in somatic cells that surround germ cells. We conclude that a cell-extrinsic mechanism regulates germline stem cell behaviour.

Zfh-1 controls somatic stem cell self-renewal in the Drosophila testis and nonautonomously influences germline stem cell self-renewal.

The ability of adult stem cells to maintain their undifferentiated state depends upon residence in their niche. While simple models of a single self-renewal signal are attractive, niche-stem cell interactions are likely to be more complex. Many niches have multiple cell types, and the Drosophila testis is one such complex niche with two stem cell types, germline stem cells (GSCs) and somatic cyst progenitor cells (CPCs). These stem cells require chemokine activation of Jak/STAT signaling for self-renewal. We identified the transcriptional repressor Zfh-1 as a presumptive somatic target of Jak/STAT signaling, demonstrating that it is necessary and sufficient to maintain CPCs. Surprisingly, sustained zfh-1 expression or intrinsic STAT activation in somatic cells caused neighboring germ cells to self-renew outside their niche. In contrast, germline-intrinsic STAT activation was insufficient for GSC renewal. These data reveal unexpected complexity in cell interactions in the niche, implicating CPCs in GSC self-renewal.

The making of a maggot: patterning the Drosophila embryonic epidermis

Cell fates are instructed by signals emitted from specialized cell populations called organizers. The study of epidermal patterning in Drosophila is contributing novel insights concerning the establishment and action of such organizers. Juxtaposed rows of cells express either the wingless or hedgehog signaling molecules and thereby act as organizers of segment pattern. These signals mediate a mutually re-enforcing interaction between the two rows of cells to sustain organizer function. In a distinct and subsequent phase, wingless and hedgehog act to specify the fates of cells.

Dynamics of the male germline stem cell population during aging of Drosophila melanogaster

Drosophila melanogaster has emerged as an important model system for the study of both stem cell biology and aging. Much is known about how molecular signals from the somatic niche regulate adult stem cells in the germline, and a variety of environmental factors as well as single point mutations have been shown to affect lifespan. Relatively little is known, however, about how aging affects specific populations of cells, particularly adult stem cells that may be susceptible to aging‐related damage. Here we show that male germline stem cells (GSCs) are lost from the stem cell niche during aging, but are efficiently replaced to maintain overall stem cell number. We also find that the division rate of GSCs slows significantly during aging, and that this slowing correlates with a reduction in the number of somatic hub cells that contribute to the stem cell niche. Interestingly, slowing of stem cell division rate was not observed in long‐lived methuselah mutant flies. We finally investigated whether two mechanisms that are thought to be used in other adult stem cell types to minimize the effects of aging were operative in this system. First, in many adult tissues stem cells exhibit markedly fewer cell cycles relative to transit‐amplifying cells, presumably protecting the stem cell pool from replication‐associated damage. Second, at any given time not all stem cells actively cycle, leading to ‘clonal succession’ from the reserve pool of initially quiescent stem cells. We find that neither of these mechanisms is used in Drosophila male GSCs.

A somatic role for eyes absent (eya) and sine oculis (so) in drosophila spermatocyte development

Interactions between the soma and the germline are a conserved feature of spermatogenesis throughout the animal kingdom. In this report, we find that the transcription factors eyes absent (eya) and sine oculis (so), previously shown to play major roles during eye development [Cell 91 (1997), 881] are each required in the somatic cyst cells of the testis for proper Drosophila spermatocyte development. eya mutant testes exhibit degenerating young spermatocytes. Mosaic analysis reveals a somatic requirement for both eya and so, in that neither gene is required in the germline for spermatocyte development. Immunolocalization analysis supports this somatic role, since both proteins are localized within cyst cell nuclei as spermatocytes differentiate from amplifying spermatogonia. Using antibodies against known cyst cell markers, we demonstrate that cysts of degenerating spermatocytes in eya mutant testes are encysted, ruling out a role for eya in cyst cell viability. Finally, we have uncovered a genetic interaction between eya and so in the testis, suggesting that, as in the eye, eya and so may form a transcription complex responsible for the activation of target genes involved in cyst cell differentiation and spermatocyte development.

Divide and conquer: pattern formation in Drosophila embryonic epidermis

The pattern of differentiated cell types within tissues and organs is often established by organizers, the localized sources of secreted ligands. Although the mechanisms underlying organizer function have been extensively studied, only in a few cases is it clear how an organizer ultimately controls each individual cells fate across a field of progenitor cells. One of these cases involves the establishment of a precise pattern of cell differentiation across the embryonic epidermis in Drosophila. Here, we review several recent reports that help to elucidate the regulatory principles used to control this pattern. Because organizers are conserved, the same fundamental principles might operate in other organizers.