Antonio Baonza

The Orientation of Cell Divisions Determines the Shape of Drosophila Organs

Organ shape depends on the coordination between cell proliferation and the spatial arrangement of cells during development. Much is known about the mechanisms that regulate cell proliferation, but the processes by which the cells are orderly distributed remain unknown. This can be accomplished either by random division of cells that later migrate locally to new positions (cell allocation) or through polarized cell division (oriented cell division; OCD). Recent data suggest that the OCD is involved in some morphogenetic processes such as vertebrate gastrulation, neural tube closure, and growth of shoot apex in plants; however, little is known about the contribution of OCD during organogenesis. We have analyzed the orientation patterns of cell division throughout the development of wild-type and mutant imaginal discs of Drosophila. Our results show a causal relationship between the orientation of cell divisions in the imaginal disc and the adult morphology of the corresponding organs, indicating a key role of OCD in organ-shape definition. In addition, we find that a subset of planar cell polarity genes is required for the proper orientation of cell division during organ development.

The bHLH factor deadpan is a direct target of Notch signaling and regulates neuroblast self-renewal in Drosophila.

A defining feature of stem cells is their capacity to renew themselves at each division while producing differentiated progeny. How these cells balance self-renewal versus differentiation is a fundamental issue in developmental and cancer biology. The Notch signaling pathway has long been known to influence cell fate decisions during development. Indeed, there is a great deal of evidence correlating its function with the regulation of neuroblast (NB) self-renewal during larval brain development in Drosophila. However, little is known about the transcription factors regulated by this pathway during this process. Here we show that deadpan (dpn), a gene encoding a bHLH transcription factor, is a direct target of the Notch signaling pathway during type II NB development. Type II NBs undergo repeated asymmetric divisions to self-renew and to produce immature intermediate neural progenitors. These cells mature into intermediate neural progenitors (INPs) that have the capacity to undergo multiple rounds of asymmetric division to self-renew and to generate GMCs and neurons. Our results indicate that the expression of dpn at least in INPs cells depends on Notch signaling. The ectopic expression of dpn in immature INP cells can transform these cells into NBs-like cells that divide uncontrollably causing tumor over-growth. We show that in addition to dpn, Notch signaling must be regulating other genes during this process that act redundantly with dpn.

Dual role of extramacrochaetae in cell proliferation and cell differentiation during wing morphogenesis in Drosophila

The Extramacrochaetae (emc) gene encodes a transcription factor with an HLH domain without the basic region involved in interaction with DNA present in other proteins that have this domain. EMC forms heterodimers with bHLH proteins preventing their binding to DNA, acting as a negative regulator. The function of emc is required in many developmental processes during the development of Drosophila, including wing morphogenesis. Mitotic recombination clones of both null and gain-of-function alleles of emc, indicate that during wing morphogenesis, emc participates in cell proliferation within the intervein regions (vein patterning), as well as in vein differentiation. The study of relationships between emc and different genes involved in wing development reveal strong genetic interactions with genes of the Ras signalling pathway (torpedo, vein, veinlet and Gap), blistered, plexus and net, in both adult wing phenotypes and cell behaviour in genetic mosaics. These interactions are also analyzed as variations of emc expression patterns in mutant backgrounds for these genes. In addition, cell proliferation behaviour of emc mutant cells varies depending on the mutant background. The results show that genes of the Ras signalling pathway are co-operatively involved in the activity of emc during cell proliferation, and later antagonistically during cell differentiation, repressing EMC expression.

Pattern reorganization occurs independently of cell division during Drosophila wing disc regeneration in situ

One of the most intriguing problems in developmental biology is how an organism can replace missing organs or portions of its body after injury. This capacity, known as regeneration, is conserved across different phyla. The imaginal discs of Drosophila melanogaster provide a particularly well-characterized model for analyzing regeneration. We have developed a unique method to study organ regeneration under physiological conditions using the imaginal discs of Drosophila. Using this method, we revisited different aspects of organ regeneration. The results presented in this report suggest that during the initial stages of regeneration, different processes occur, including wound healing, a temporary loss of markers of cell-fate commitment, and pattern reorganization. We present evidence indicating that all of these processes occur even when cell division has been arrested. Our data also suggested that Wingless is not required during the early stages of disc regeneration.

The bHLH Factors Extramacrochaetae and Daughterless Control Cell Cycle in Drosophila Imaginal Discs through the Transcriptional Regulation of the cdc25 Phosphatase string

One of the major issues in developmental biology is about having a better understanding of the mechanisms that regulate organ growth. Identifying these mechanisms is essential to understand the development processes that occur both in physiological and pathological conditions, such as cancer. The E protein family of basic helix-loop helix (bHLH) transcription factors, and their inhibitors the Id proteins, regulate cell proliferation in metazoans. This notion is further supported because the activity of these factors is frequently deregulated in cancerous cells. The E protein orthologue Daughterless (Da) and the Id orthologue Extramacrochaetae (Emc) are the only members of these classes of bHLH proteins in Drosophila. Although these factors are involved in controlling proliferation, the mechanism underlying this regulatory activity is poorly understood. Through a genetic analysis, we show that during the development of epithelial cells in the imaginal discs, the G2/M transition, and hence cell proliferation, is controlled by Emc via Da. In eukaryotic cells, the main activator of this transition is the Cdc25 phosphatase, string. Our genetic analyses reveal that the ectopic expression of string in cells with reduced levels of Emc or high levels of Da is sufficient to rescue the proliferative defects seen in these mutant cells. Moreover, we present evidence demonstrating a role of Da as a transcriptional repressor of string. Taken together, these findings define a mechanism through which Emc controls cell proliferation by regulating the activity of Da, which transcriptionally represses string.

The bHLH factors Dpn and members of the E(spl) complex mediate the function of Notch signalling regulating cell proliferation during wing disc development.

Summary The Notch signalling pathway plays an essential role in the intricate control of cell proliferation and pattern formation in many organs during animal development. In addition, mutations in most members of this pathway are well characterized and frequently lead to tumour formation. The Drosophila imaginal wing discs have provided a suitable model system for the genetic and molecular analysis of the different pathway functions. During disc development, Notch signalling at the presumptive wing margin is necessary for the restricted activation of genes required for pattern formation control and disc proliferation. Interestingly, in different cellular contexts within the wing disc, Notch can either promote cell proliferation or can block the G1-S transition by negatively regulating the expression of dmyc and bantam micro RNA. The target genes of Notch signalling that are required for these functions have not been identified. Here, we show that the Hes vertebrate homolog, deadpan (dpn), and the Enhancer-of-split complex (E(spl)C) genes act redundantly and cooperatively to mediate the Notch signalling function regulating cell proliferation during wing disc development.

Analysis of the function of apoptosis during imaginal wing disc regeneration in Drosophila melanogaster

Regeneration is the ability that allows organisms to replace missing organs or lost tissue after injuries. This ability requires the coordinated activity of different cellular processes, including programmed cell death. Apoptosis plays a key role as a source of signals necessary for regeneration in different organisms. The imaginal discs of Drosophila melanogaster provide a particularly well-characterised model system for studying the cellular and molecular mechanisms underlying regeneration. Although it has been shown that signals produced by apoptotic cells are needed for homeostasis and regeneration of some tissues of this organism, such as the adult midgut, the contribution of apoptosis to disc regeneration remains unclear. Using a new method for studying disc regeneration in physiological conditions, we have defined the pattern of cell death in regenerating discs. Our data indicate that during disc regeneration, cell death increases first at the wound edge, but as regeneration progresses dead cells can be observed in regions far away from the site of damage. This result indicates that apoptotic signals initiated in the wound spread throughout the disc. We also present results which suggest that the partial inhibition of apoptosis does not have a major effect on disc regeneration. Finally, our results suggest that during disc regeneration distinct apoptotic signals might be acting simultaneously.

JNK and JAK/STAT signalling are required for inducing loss of cell fate specification during imaginal wing discs regeneration in Drosophila melanogaster

The regenerative process after tissue damage relies on a variety of cellular responses that includes compensatory cell proliferation and cell fate re-specification. The identification of the signalling networks regulating these cellular events is a central question in regenerative biology. Tissue regeneration models in Drosophila have shown that two of the signals that play a fundamental role during the early stages of regeneration are the c-Jun N-terminal kinase (JNK) and JAK/STAT signalling pathways. These pathways have been shown to be required for controlling regenerative proliferation, however their contribution to the processes of cellular reprogramming and cell fate re-specification that take place during regeneration are largely unknown. Here, we present evidence for a previously unrecognised function of the cooperative activities of JNK and JAK/STAT signalling pathways in inducing loss of cell fate specification in imaginal discs. We show that co-activation of these signalling pathways induces both the cell fate changes in injured areas, as well as in adjacent cells. We have also found that this function relies on the activity of the Caspase initiator encoded by the gene dronc.

Drosophila as a Model System to Study Cell Signaling in Organ Regeneration

Regeneration is a fascinating phenomenon that allows organisms to replace or repair damaged organs or tissues. This ability occurs to varying extents among metazoans. The rebuilding of the damaged structure depends on regenerative proliferation that must be accompanied by proper cell fate respecification and patterning. These cellular processes are regulated by the action of different signaling pathways that are activated in response to the damage. The imaginal discs of Drosophila melanogaster have the ability to regenerate and have been extensively used as a model system to study regeneration. Drosophila provides an opportunity to use powerful genetic tools to address fundamental problems about the genetic mechanisms involved in organ regeneration. Different studies in Drosophila have helped to elucidate the genes and signaling pathways that initiate regeneration, promote regenerative growth, and induce cell fate respecification. Here we review the signaling networks involved in regulating the variety of cellular responses that are required for discs regeneration.