Germ-line stem cells (GSCs) serve as the source for gametogenesis in diverse organisms. We cloned and characterized the Drosophila piwi gene and showed that it is required for the asymmetric division of GSCs to produce and maintain a daughter GSC but is not essential for the further differentiation of the committed daughter cell. Genetic mosaic and RNA in situ analyses suggest that piwi expression in adjacent somatic cells regulates GSC division. piwi encodes a highly basic novel protein, well conserved during evolution. We isolated piwi homologs in Caenorhabditis elegans and humans and also identified Arabidopsis piwi-like genes known to be required for meristem cell maintenance. Decreasing C. elegans piwi expression reduces the proliferation of GSC-equivalent cells. Thus, piwi represents a novel class of genes required for GSC division in diverse organisms.

The dumpy gene encodes a gigantic extracellular molecule that we predict to be a membrane-anchored fibre of almost a micrometer in length. Insertion and crosslinking of this fibre within the cuticle may provide a strong anchor for the underlying tissue, allowing it to maintain mechanical tension at sites under stress. This would explain its contribution to tissue morphogenesis through its regulation of mechanical properties.

Summary Regeneration is a complex chain of events that restores a tissue to its original size and shape. The tissue-wide coordination of cellular dynamics needed for proper morphogenesis is challenged by the large dimensions of regenerating body parts. Feedback mechanisms in biochemical pathways can provide effective communication across great distances 1 - 5 , but how they might regulate growth during tissue regeneration is unresolved 6 , 7 . Here, we report that rhythmic traveling waves of Erk activity control the growth of bone in time and space in regenerating zebrafish scales, millimetre-sized discs of protective body armour. We find that Erk activity waves travel as expanding concentric rings, broadcast from a central source, inducing ring-like patterns of osteoblast tissue growth. Using a combination of theoretical and experimental analyses, we show that Erk activity propagates as excitable trigger waves able to traverse the entire scale in approximately two days, with the frequency of wave generation controlling the rate of scale regeneration. Furthermore, periodic induction of synchronous, tissue-wide Erk activation in place of travelling waves impairs tissue growth, indicating that wave-distributed Erk activation is key to regeneration. Our findings reveal trigger waves as a regulatory strategy to coordinate cell behaviour and instruct tissue form during regeneration.

The 2B5 region of the X‐chromosome in Drosophila melanogaster plays a developmentally important role in the ecdysterone‐triggered response of the late third instar salivary gland. Using a combination of transposon‐tagging and chromosomal walking techniques, we have isolated 231 kb of contiguous genomic DNA sequences corresponding to this region. We have more precisely aligned this DNA to the 2B1,2 to 2B5‐6 interval of the cytogenetic map by locating the position of three well‐characterized chromosomal breakpoints by in situ hybridization and genomic DNA blotting experiments. Labeled cDNA, synthesized from poly(A)+ RNA isolated from hormone‐induced salivary gland and imaginal disc tissues and hybridized to the cloned DNA, demonstrated that the ecdysterone‐inducible sequences mapped to DNA segments corresponding to the 2B3,4 to 2B5‐6 interval. Although some of these sequences were inducible in only one tissue type, many were found to be inducible in both salivary glands and imaginal discs. RNA blotting experiments have detected a major 4.5‐kb RNA which is hormone inducible in the larval salivary gland and whose quantitative induction is not inhibited by cycloheximide. Thus, the 4.5‐kb RNA represents at least one product from the ecdysterone‐responsive 2B5 ‘early’ puff.

Wnt signaling specifies cell fates in many tissues during vertebrate and invertebrate embryogenesis. To understand better how Wnt signaling is regulated during development, we have performed genetic screens to isolate mutations that suppress or enhance mutations in the fly Wnt homolog, wingless (wg). We find that loss-of-function mutations in the neural determinant SoxNeuro (also known as Sox-neuro, SoxN) partially suppress wg mutant pattern defects. SoxN encodes a HMG-box-containing protein related to the vertebrate Sox1, Sox2 and Sox3 proteins, which have been implicated in patterning events in the early mouse embryo. In Drosophila, SoxN has previously been shown to specify neural progenitors in the embryonic central nervous system. Here, we show that SoxN negatively regulates Wg pathway activity in the embryonic epidermis. Loss of SoxN function hyperactivates the Wg pathway, whereas its overexpression represses pathway activity. Epistasis analysis with other components of the Wg pathway places SoxN at the level of the transcription factor Pan (also known as Lef, Tcf) in regulating target gene expression. In human cell culture assays, SoxN represses Tcf-responsive reporter expression, indicating that the fly gene product can interact with mammalian Wnt pathway components. In both flies and in human cells, SoxN repression is potentiated by adding ectopic Tcf, suggesting that SoxN interacts with the repressor form of Tcf to influence Wg/Wnt target gene transcription.

piwi represents the first class of genes known to be required for stem cell self-renewal in diverse organisms. In the Drosophila ovary, piwi is required in somatic signaling cells to maintain germline stem cells. Here we show that piwi encodes a novel nucleoplasmic protein present in both somatic and germline cells, with the highly conserved C-terminal region essential for its function. Removing PIWI protein from single germline stem cells significantly decreases the rate of their division. This suggests that PIWI has a second role as a cell-autonomous promoter of germline stem cell division. Consistent with its dual function, over-expression of piwi in somatic cells causes an increase both in the number of germline stem cells and the rate of their division. Thus, PIWI is a key regulator of stem cell division - its somatic expression modulates the number of germline stem cells and the rate of their division, while its germline expression also contributes to promoting stem cell division in a cell-autonomous manner.

Wg/Wnt signals specify cell fates in both invertebrate and vertebrate embryos and maintain stem-cell populations in many adult tissues. Deregulation of the Wnt pathway can transform cells to a proliferative fate, leading to cancer. We have discovered that two Drosophila proteins that are crucial for cytokinesis have a second, largely independent, role in restricting activity of the Wnt pathway. The fly homolog of RacGAP1, Tumbleweed (Tum)/RacGAP50C, and its binding partner, the kinesin-like protein Pavarotti (Pav), negatively regulate Wnt activity in fly embryos and in cultured mammalian cells. Unlike many known regulators of the Wnt pathway, these molecules do not affect stabilization of Arm/β-catenin (βcat), the principal effector molecule in Wnt signal transduction. Rather, they appear to act downstream of βcat stabilization to control target-gene transcription. Both Tum and Pav accumulate in the nuclei of interphase cells, a location that is spatially distinct from their cleavage-furrow localization during cytokinesis. We show that this nuclear localization is essential for their role in Wnt regulation. Thus, we have identified two modulators of the Wnt pathway that have shared functions in cell division, which hints at a possible link between cytokinesis and Wnt activity during tumorigenesis.

SUMMARYThe specification of the body plan in vertebrates and invertebrates is controlled by a variety of cell signaling pathways, but how signaling output is translated into morphogenesis is an ongoing question. Here, we describe genetic interactions between the Wingless (Wg) signaling pathway and a nonmuscle myosin heavy chain, encoded by the crinkled (ck) locus in Drosophila. In a screen for mutations that modify wg loss-of-function phenotypes, we isolated multiple independent alleles of ck. These ck mutations dramatically alter the morphology of the hook-shaped denticles that decorate the ventral surface of the wg mutant larval cuticle. In an otherwise wild-type background, ck mutations do not significantly alter denticle morphology, suggesting a specific interaction with Wg-mediated aspects of epidermal patterning. Here, we show that changing the level of Wg activity changes the structure of actin bundles during denticle formation in ck mutants. We further find that regulation of the Wg target gene, shaven-baby (svb), and of its transcriptional targets, miniature (m) and forked (f), modulates this ck-dependent process. We conclude that Ck acts in concert with Wg targets to orchestrate the proper shaping of denticles in the Drosophila embryonic epidermis.