My what big eyes you have: how the Drosophila retina grows

The compound eye of the fruit fly, Drosophila melanogaster, has for decades been used extensively to study a number of critical developmental processes including tissue development, pattern formation, cell fate specification, and planar cell polarity. To a lesser degree it has been used to examine the cell cycle and tissue proliferation. Discovering the mechanisms that balance tissue growth and cell death in developing epithelia has traditionally been the realm of those using the wing disc. However, over the last decade a series of observations has demonstrated that the eye is a suitable and maybe even preferable tissue for studying tissue growth. This review will focus on how growth of the retina is controlled by the genes and pathways that govern the specification of tissue fate, the division of the epithelium into dorsal-ventral compartments, the initiation, and progression of the morphogenetic furrow and the second mitotic wave.     

FGF signal interpretation is directed by Sprouty and Spred proteins during mesoderm formation

Vertebrate gastrulation requires coordination of mesoderm specification with morphogenetic movements. While both of these processes require FGF signaling, it is not known how mesoderm specification and cell movements are coordinated during gastrulation. The related Sprouty and Spred protein families are recently discovered regulators of receptor tyrosine kinase signaling. We identified two genes for each family in Xenopus tropicalis: Xtsprouty1, Xtsprouty2, Xtspred1, and Xtspred2. In gain- and loss-of-function experiments we show that XtSprouty and XtSpred proteins modulate different signaling pathways downstream of the FGF receptor (FGFR), and consequently different developmental processes. Notably, XtSproutys inhibit morphogenesis and Ca(2+) and PKCdelta signaling, leaving MAPK activation and mesoderm specification intact. In contrast, XtSpreds inhibit MAPK activation and mesoderm specification, with little effect on Ca(2+) or PKCdelta signaling. These differences, combined with the timing of their developmental expression, suggest a mechanism to switch FGFR signal interpretation to coordinate mesoderm formation and cell movements during gastrulation.     

Frodo proteins: modulators of Wnt signaling in vertebrate development

The Frodo/dapper (Frd) proteins are recently discovered signaling adaptors, which functionally and physically interact with Wnt and Nodal signaling pathways during vertebrate development. The Frd1 and Frd2 genes are expressed in dynamic patterns in early embryos, frequently in cells undergoing epithelial-mesenchymal transition. The Frd proteins function in multiple developmental processes, including mesoderm and neural tissue specification, early morphogenetic cell movements, and organogenesis. Loss-of-function studies using morpholino antisense oligonucleotides demonstrate that the Frd proteins regulate Wnt signal transduction in a context-dependent manner and may be involved in Nodal signaling. The identification of Frd-associated factors and cellular targets of the Frd proteins should shed light on the molecular mechanisms underlying Frd functions in embryonic development and in cancer.     

Epithelial differentiation in Drosophila

Our understanding of epithelial development in Drosophila has been greatly improved in recent years. Two key regulators of epithelial polarity, Crumbs and DE-cadherin, have been studied at the genetic and molecular levels and a number of additional genes are being analyzed that contribute to the differentiation of epithelial cell structure. Epithelial architecture has a profound influence on morphogenetic movements, patterning and cell-type determination. The combination of embryological and genetic/molecular tools in Drosophila will help us to elucidate the complex events that determine epithelial cell structure and how they relate to morphogenesis and other developmental processes.     

Planar cell polarity regulators in asymmetric organogenesis during development and disease

The phenomenon of planar cell polarity is critically required for a myriad of morphogenetic processes in metazoan and is accurately controlled by several conserved modules. Six “core” proteins, including Frizzled, Flamingo (Celsr), Van Gogh (Vangl), Dishevelled, Prickle, and Diego (Ankrd6), are major components of the Wnt/planar cell polarity pathway. The Fat/Dchs protocadherins and the Scrib polarity complex also function to instruct cellular polarization. In vertebrates, all these pathways are essential for tissue and organ morphogenesis, such as neural tube closure, left–right symmetry breaking, heart and gut morphogenesis, lung and kidney branching, stereociliary bundle orientation, and proximal–distal limb elongation. Mutations in planar polarity genes are closely linked to various congenital diseases. Striking advances have been made in deciphering their contribution to the establishment of spatially oriented pattern in developing organs and the maintenance of tissue homeostasis. The challenge remains to clarify the complex interplay of different polarity pathways in organogenesis and the link of cell polarity to cell fate specification. Interdisciplinary approaches are also important to understand the roles of mechanical forces in coupling cellular polarization and differentiation. This review outlines current advances on planar polarity regulators in asymmetric organ formation, with the aim to identify questions that deserve further investigation.     

Targeted expression of the dominant-negative FGFR4a in the eye using Xrx1A regulatory sequences interferes with normal retinal development

Molecular analysis of vertebrate eye development has been hampered by the availability of sequences that can selectively direct gene expression in the developing eye. We report the characterization of the regulatory sequences of the Xenopus laevis Rx1A gene that can direct gene expression in the retinal progenitor cells. We have used these sequences to investigate the role of Fibroblast Growth Factor (FGF) signaling in the development of retinal cell types. FGFs are signaling molecules that are crucial for correct patterning of the embryo and that play important roles in the development of several embryonic tissues. FGFs and their receptors are expressed in the developing retina, and FGF receptor-mediated signaling has been implicated to have a role in the specification and survival of retinal cell types. We investigated the role of FGF signaling mediated by FGF receptor 4a in the development of retinal cell types in Xenopus laevis. For this purpose, we have made transgenic Xenopus tadpoles in which the dominant-negative FGFR4a (Delta FGFR4a) coding region was linked to the newly characterized regulatory sequences of the Xrx1A gene. We found that the expression of Delta FGFR4a in retinal progenitor cells results in abnormal retinal development. The retinas of transgenic animals expressing Delta FGFR4a show disorganized cell layering and specifically lack photoreceptor cells. These experiments show that FGFR4a-mediated FGF signaling is necessary for the correct specification of retinal cell types. Furthermore, they demonstrate that constructs using Xrx1A regulatory sequences are excellent tools with which to study the developmental processes involved in retinal formation.     

Overexpression of poly(ADP-ribose) polymerase disrupts organization of cytoskeletal F-actin and tissue polarity in Drosophila.

Poly(ADP-ribose) polymerase (PARP) may play important roles in nuclear events such as cell cycle, cell proliferation, and maintenance of chromosomal stability. However, the exact biological role played by PARP or how PARP is involved in these cellular functions is still unclear. To elucidate the biological functions of PARP in vivo, we have constructed transgenic flies that overexpress Drosophila PARP in the developing eye primordia. These flies showed mild roughening of the normally smooth ommatidial lattice and tissue polarity disruption caused by improper rotation and chirality of the ommatidia. To clarify how this phenotypical change was induced, here we analyzed transgenic flies overexpressing PARP in the developing eye, embryo, and adult in detail. PARP mRNA level and the phenotype were enhanced in flies carrying more copies of the transgene. Developing eyes from third instar larvae were analyzed by using the neural cell marker to examine the involvement of PARP in cell fate. Morphological disorder of non-neuronal accessory cells was observed in PARP transgenic flies. Interestingly, overexpression of PARP did not interfere with the cell cycle or apoptosis, but it did disrupt the organization of cytoskeletal F-actin, resulting in aberrant cell and tissue morphology. Furthermore, heat-induced PARP expression disrupted organization of cytoskeletal F-actin in embryos and tissue polarity in adult flies. Because these phenotypes closely resembled mutants or transgenic flies of the tissue polarity genes, genetic interaction of PARP with known tissue polarity genes was examined. Transgenic flies expressing either PARP or RhoA GTPase in the eye were crossed, and co-expression of PARP suppressed the effect of RhoA GTPase. Our results indicate that PARP may play a role in cytoskeletal or cytoplasmic events in developmental processes of Drosophila.     

Redundant functions of Rac GTPases in inner ear morphogenesis

Development of the mammalian inner ear requires coordination of cell proliferation, cell fate determination and morphogenetic movements. While significant progress has been made in identifying developmental signals required for inner ear formation, less is known about how distinct signals are coordinated by their downstream mediators. Members of the Rac family of small GTPases are known regulators of cytoskeletal remodeling and numerous other cellular processes. However, the function of Rac GTPases in otic development is largely unexplored. Here, we show that Rac1 and Rac3 redundantly regulate many aspects of inner ear morphogenesis. While no morphological defects were observed in Rac3(-/-) mice, Rac1(CKO); Rac3(-/-) double mutants displayed enhanced vestibular and cochlear malformations compared to Rac1(CKO) single mutants. Moreover, in Rac1(CKO); Rac3(-/-) mutants, we observed compromised E-cadherin-mediated cell adhesion, reduced cell proliferation and increased cell death in the early developing otocyst, leading to a decreased size and malformation of the membranous labyrinth. Finally, cochlear extension was severely disrupted in Rac1(CKO); Rac3(-/-) mutants, accompanied by a loss of epithelial cohesion and formation of ectopic sensory patches underneath the cochlear duct. The compartmentalized expression of otic patterning genes within the Rac1(CKO); Rac3(-/-) mutant otocyst was largely normal, however, indicating that Rac proteins regulate inner ear morphogenesis without affecting cell fate specification. Taken together, our results reveal an essential role for Rac GTPases in coordinating cell adhesion, cell proliferation, cell death and cell movements during otic development.     

Early stages of zebrafish eye formation require the coordinated activity of Wnt11, Fz5, and the Wnt/beta-catenin pathway

During regional patterning of the anterior neural plate, a medially positioned domain of cells is specified to adopt retinal identity. These eye field cells remain coherent as they undergo morphogenetic events distinct from other prospective forebrain domains. We show that two branches of the Wnt signaling pathway coordinate cell fate determination with cell behavior during eye field formation. Wnt/beta-catenin signaling antagonizes eye specification through the activity of Wnt8b and Fz8a. In contrast, Wnt11 and Fz5 promote eye field development, at least in part, through local antagonism of Wnt/beta-catenin signaling. Additionally, Wnt11 regulates the behavior of eye field cells, promoting their cohesion. Together, these results allow us to postulate a model in which Wnt11 and Fz5 signaling promotes early eye development through the coordinated antagonism of signals that suppress retinal identity and promotion of coherence of eye field cells.     

JNK signaling controls border cell cluster integrity and collective cell migration

Collective cell movement is a mechanism for invasion identified in many developmental events. Examples include the movement of lateral-line neurons in Zebrafish, cells in the inner blastocyst, and metastasis of epithelial tumors [1]. One key model to study collective migration is the movement of border cell clusters in Drosophila. Drosophila egg chambers contain 15 nurse cells and a single oocyte surrounded by somatic follicle cells. At their anterior end, polar cells recruit several neighboring follicle cells to form the border cell cluster [2]. By stage 9, and over 6 hr, border cells migrate as a cohort between nurse cells toward the oocyte. The specification and directionality of border cell movement are regulated by hormonal and signaling mechanisms [3]. However, how border cells are held together while they migrate is not known. Here, we show that a negative-feedback loop controlling JNK activity regulates border cell cluster integrity. JNK signaling modulates contacts between border cells and between border cells and substratum to sustain collective migration by regulating several effectors including the polarity factor Bazooka and the cytoskeletal adaptor D-Paxillin. We anticipate a role for the JNK pathway in controlling collective cell movements in other morphogenetic and clinical models.     

Strabismus requires Flamingo and Prickle function to regulate tissue polarity in the Drosophila eye

Tissue polarity in Drosophila is regulated by a number of genes that are thought to function in a complex, many of which interact genetically and/or physically, co-localize, and require other tissue polarity proteins for their localization. We report the enhancement of the strabismus tissue polarity phenotype by mutations in two other tissue polarity genes, flamingo and prickle. Flamingo is autonomously required for the establishment of ommatidial polarity. Its localization is dynamic throughout ommatidial development and is dependent on Frizzled and Notch. Flamingo and Strabismus co-localize for several rows posterior to the morphogenetic furrow and subsequently diverge. While neither of these proteins is required for the other's localization, Prickle localization is influenced by Strabismus function. Our data suggest that Strabismus, Flamingo and Prickle function together to regulate the establishment of tissue polarity in the Drosophila eye.