Biomedical subjects

Ilaria Rebay

Publications and source records attributed to Ilaria Rebay.

At least 19 recordsLinked to original sources

PubMed2006

Characterization of the split ends-like gene spenito reveals functional antagonism between SPOC family members during Drosophila eye development.

The novel family of SPOC domain proteins is composed of broadly conserved nuclear factors that fall into two subclasses, termed large and small, based on protein size. Members of the large subgroup, which includes Drosophila SPEN and human SHARP, have been characterized as transcriptional corepressors acting downstream of a variety of essential cell signaling pathways, while those of the small subclass have remained largely unstudied. Since SPEN has been implicated in Drosophila eye development, and the small SPOC protein NITO is also expressed in the developing eye, we have used this context to perform a structure-function analysis of NITO and to examine the relationship between the two SPOC family subclasses. Our results demonstrate that the phenotypes obtained from overexpressing NITO share striking similarity to those associated with loss of spen. Dosage-sensitive genetic interactions further support a model of functional antagonism between NITO and SPEN during Drosophila eye development. These results suggest that large and small SPOC family proteins may have opposing functions in certain developmental contexts.

Amino Acid Sequence↗

PubMed2006

Intersection of signal transduction pathways and development.

One of the challenges of modern biology is to understand how cells within a developing organism generate, integrate, and respond to dynamic informational cues. Based on over two decades of intensive research, many parts and subroutines of the responsible signal transduction networks have been identified and functionally characterized. From this work, it has become evident that a complicated interplay between signaling pathways, involving extensive feedback regulation and multiple levels of cross-talk, underlies even the "simplest" developmental decision. Thus a signaling pathway can no longer be thought of as a rigid linear process, but rather must be considered a dynamic, self-interacting, and self-adjusting network. The Epidermal Growth Factor Receptor tyrosine kinase signaling pathway provides a prime vantage point from which to explore emerging principles in developmental signal transduction.

Animals↗

PubMed2006

Targeting Drosophila eye development.

In order to understand the role of transcription factors in particular developmental processes it is necessary to know their target genes. A combination of bioinformatics, comparative expression profiling and microarray-based epistasis experiments has recently identified new targets of Eyeless, a key transcription factor in Drosophila retinal determination.

Animals↗

PubMed2005

The Drosophila nucleosome remodeling factor NURF is required for Ecdysteroid signaling and metamorphosis.

Drosophila NURF is an ISWI-containing ATP-dependent chromatin remodeling complex that regulates transcription by catalyzing nucleosome sliding. To determine in vivo gene targets of NURF, we performed whole genome expression analysis on mutants lacking the NURF-specific subunit NURF301. Strikingly, a large set of ecdysone-responsive targets is included among several hundred NURF-regulated genes. Null Nurf301 mutants do not undergo larval to pupal metamorphosis, and also enhance dominant-negative mutations in ecdysone receptor. Moreover, purified NURF binds EcR in an ecdysone-dependent manner, suggesting it is a direct effector of nuclear receptor activity. The conservation of NURF in mammals has broad implications for steroid signaling.

Animals↗

PubMed2005

Molecular genetics of spinocerebellar ataxia type 8 (SCA8).

Spinocerebellar ataxias (SCAs) belong to a group of autosomal dominant, late-onset neurodegenerative disorders characterized by slowly progressive ataxia that eventually leads to severe gait, speech, coordination and sensory loss. The majority of these diseases result from expanded polyglutamine tracts in the encoded protein as seen in SCA1, SCA2, SCA3, SCA6, SCA7 and Dentatorubral-Pallidoluysian Atrophy (DRPLA). However, two novel forms of SCAs, SCA8 and SCA12, are associated with trinucleotide repeat expansions in non-translated regions of the genes. In the case of SCA8, the CUG expansion occurs at the 3' end of a processed non-coding RNA. While understanding of how expanded polyglutamine tracts compromise or alter protein function has advanced rapidly in the last five years, understanding of how trinucleotide repeat expansions alter the function of the non-coding SCA8 RNA and lead to human disease remains quite limited. Encouragingly, as discussed in this review, recent studies from murine and Drosophila models have provided new insights into both the cellular context in which SCA8 normally operates and the potential role of CTG expansion in the disease. Continued exploration of these genetically tractable model systems will further illuminate the biology underlying SCA8 disease, ultimately providing the necessary foundation on which to develop effective therapeutic interventions.

Animals↗

PubMed2005

Using Drosophila to decipher how mutations associated with human branchio-oto-renal syndrome and optical defects compromise the protein tyrosine phosphatase and transcriptional functions of eyes absent.

Eyes absent (EYA) proteins are defined by a conserved C-terminal EYA domain (ED) that both contributes to its function as a transcriptional coactivator by mediating protein-protein interactions and possesses intrinsic protein tyrosine phosphatase activity. Mutations in human EYA1 result in an autosomal dominant disorder called branchio-oto-renal (BOR) syndrome as well as congenital cataracts and ocular defects (OD). Both BOR- and OD-associated missense mutations alter residues in the conserved ED as do three missense mutations identified from Drosophila eya alleles. To investigate the molecular mechanisms whereby these mutations disrupt EYA function, we tested their activity in a series of assays that measured in vivo function, phosphatase activity, transcriptional capability, and protein-protein interactions. We find that the OD-associated mutations retain significant in vivo activity whereas those derived from BOR patients show a striking decrease or loss of in vivo functionality. Protein-protein interactions, either with its partner transcription factor Sine oculis or with EYA itself, were not significantly compromised. Finally, the results of the biochemical assays suggest that both loss of protein tyrosine phosphatase activity and reduced transcriptional capability contribute to the impaired EYA function associated with BOR/OD syndrome, thus shedding new light into the molecular mechanisms underlying this disease.

Alleles↗

PubMed2005

Post-translational modifications influence transcription factor activity: a view from the ETS superfamily.

Transcription factors provide nodes of information integration by serving as nuclear effectors of multiple signaling cascades, and thus elaborate layers of regulation, often involving post-translational modifications, modulating and coordinate activities. Such modifications can rapidly and reversibly regulate virtually all transcription factor functions, including subcellular localization, stability, interactions with cofactors, other post-translational modifications and transcriptional activities. Aside from analyses of the effects of serine/threonine phosphorylation, studies on post-translational modifications of transcription factors are only in the initial stages. In particular, the regulatory possibilities afforded by combinatorial usage of and competition between distinct modifications on an individual protein are immense, and with respect to large families of closely related transcription factors, offer the potential of conferring critical specificity. Here we will review the post-translational modifications known to regulate ETS transcriptional effectors and will discuss specific examples of how such modifications influence their activities to highlight emerging paradigms in transcriptional regulation.

Animals↗

PubMed2005

New vision from Eyes absent: transcription factors as enzymes.

Post-translational modifications such as phosphorylation provide versatile context-specific strategies for modulating transcription factor activity. In the prevailing view of this process, modifying enzymes indirectly influence gene expression by shuttling to and from the nucleus where they alter the activity of their target transcription factors. However, a new paradigm has recently been suggested from studies of Eyes absent (EYA), a member of a conserved network of transcriptional regulators implicated in the development of numerous tissues and organs including the eye, ear, muscle and kidney. These findings indicate that EYA operates both as a transcriptional coactivator and as the prototype of a novel class of protein tyrosine phosphatases. The regulatory potential of such a bifunctional transcription factor is enormous and suggests a new layer of dynamic regulation in which transcription factors themselves might provide intrinsic enzymatic activities to fine-tune nuclear output.

Animals↗

PubMed2005

Signaling circuitries in development: insights from the retinal determination gene network.

Context-specific integration of information received from the Notch, Transforming growth factor beta, Wingless/Wnt, Hedgehog and Epidermal growth factor receptor signaling pathways sets the stage for deployment of the retinal determination gene network (RDGN), a group of transcription factors that collectively directs the formation of the eye and other tissues. Recent investigations have revealed how these transcription factors are regulated by their interactions with each other and with effectors of the above signaling pathways. Further study of the RDGN may provide insights into how common cues can generate context-specific responses, a key aspect of developmental regulation that remains poorly understood.

Animals↗

PubMed2005

Function of the ETS transcription factor Yan in border cell migration.

Invasive cell migration in both normal development and metastatic cancer is regulated by various signaling pathways, transcription factors and cell-adhesion molecules. The coordination between these activities in the context of cell migration is poorly understood. During Drosophila oogenesis, a small group of cells called border cells exit the follicular epithelium to perform a stereotypic, invasive migration. We find that the ETS transcription factor Yan is required for border cell migration and that Yan expression is spatiotemporally regulated as border cells migrate from the anterior pole of the egg chamber towards the nurse cell-oocyte boundary. Yan expression is dependent on inputs from the JAK/STAT, Notch and Receptor Tyrosine Kinase pathways in border cells. Mechanistically, Yan functions to modulate the turnover of DE-Cadherin-dependent adhesive complexes to facilitate border cell migration. Our results suggest that Yan acts as a pivotal link between signal transduction, cell adhesion and invasive cell migration in Drosophila border cells.

Animals↗

PubMed2004

Derepression by depolymerization; structural insights into the regulation of Yan by Mae.

Yan, an ETS family transcriptional repressor, is regulated by receptor tyrosine kinase signaling via the Ras/MAPK pathway. Phosphorylation and downregulation of Yan is facilitated by a protein called Mae. Yan and Mae interact through their SAM domains. We find that repression by Yan requires the formation of a higher order structure mediated by Yan-SAM polymerization. Moreover, a crystal structure of the Yan-SAM/Mae-SAM complex shows that Mae-SAM specifically recognizes a surface on Yan-SAM that is also required for Yan-SAM polymerization. Mae-SAM binds to Yan-SAM with approximately 1000-fold higher affinity than Yan-SAM binds to itself and can effectively depolymerize Yan-SAM. Mutations on Mae that specifically disrupt its SAM domain-dependent interactions with Yan disable the derepression function of Mae in vivo. Depolymerization of Yan by Mae represents a novel mechanism of transcriptional control that sensitizes Yan for regulation by receptor tyrosine kinases.

Amino Acid Sequence↗

PubMed2004

The spinocerebellar ataxia 8 noncoding RNA causes neurodegeneration and associates with staufen in Drosophila.

Spinocerebellar Ataxia 8 (SCA8) appears unique among triplet repeat expansion-induced neurodegenerative diseases because the predicted gene product is a noncoding RNA. Little is currently known about the normal function of SCA8 in neuronal survival or how repeat expansion contributes to neurodegeneration. To investigate the molecular context in which SCA8 operates, we have expressed the human SCA8 noncoding RNA in Drosophila. SCA8 induces late-onset, progressive neurodegeneration in the Drosophila retina. Using this neurodegenerative phenotype as a sensitized background for a genetic modifier screen, we have identified mutations in four genes: staufen, muscle-blind, split ends, and CG3249. All four encode neuronally expressed RNA binding proteins conserved in Drosophila and humans. Although expression of both wild-type and repeat-expanded SCA8 induce neurodegeneration, the strength of interaction with certain modifiers differs between the two SCA8 backgrounds, suggesting that CUG expansions alter associations with specific RNA binding proteins. Our demonstration that SCA8 can recruit Staufen and that the interaction domain maps to the portion of the SCA8 RNA that undergoes repeat expansion in the human disease suggests a specific mechanism for SCA8 function and disease. Genetic modifiers identified in our SCA8-based screens may provide candidates for designing therapeutic interventions to treat this disease.

Animals↗

PubMed2004

Signal integration during development: insights from the Drosophila eye.

The Drosophila eye is a highly ordered epithelial tissue composed of approximately 750 subunits called ommatidia arranged in a reiterated hexagonal pattern. At higher resolution, observation of the constituent photoreceptors, cone cells, and pigment cells of the eye reveals a highly ordered mosaic of amazing regularity. This relatively simple organization belies the repeated requirement for spatially and temporally coordinated inputs from the Hedgehog (Hh), Wingless (Wg), Decapentaplegic (Dpp), JAK-STAT, Notch, and receptor tyrosine kinase (RTK) signaling pathways. This review will discuss how signaling inputs from the Notch and RTK pathways, superimposed on the developmental history of a cell, facilitate context-specific and appropriate cell fate specification decisions in the developing fly eye. Lessons learned from investigating the combinatorial signal integration strategies underlying Drosophila eye development will likely reveal cell-cell communication paradigms relevant to many aspects of invertebrate and mammalian development. Developmental Dynamics 229:162-175, 2004.

Animals↗

PubMed2004

New class of Son-of-sevenless (Sos) alleles highlights the complexities of Sos function.

The guanine nucleotide exchange factor (GEF) Son-of-sevenless (Sos) encodes a complex multidomain protein best known for its role in activating the small GTPase RAS in response to receptor tyrosine kinase (RTK) stimulation. Much less well understood is SOS's role in modulating RAC activity via a separate GEF domain. In the course of a genetic modifier screen designed to investigate the complexities of RTK/RAS signal transduction, a complementation group of 11 alleles was isolated and mapped to the Sos locus. Molecular characterization of these alleles indicates that they specifically affect individual domains of the protein. One of these alleles, SosM98, which contains a single amino acid substitution in the RacGEF motif, functions as a dominant negative in vivo to downregulate RTK signaling. These alleles provide new tools for future investigations of SOS-mediated activation of both RAS and RAC and how these dual roles are coordinated and coregulated during development.

Alleles↗

PubMed2004

MAE, a dual regulator of the EGFR signaling pathway, is a target of the Ets transcription factors PNT and YAN.

Ets transcription factors play crucial roles in regulating diverse cellular processes including cell proliferation, differentiation and survival. Coordinated regulation of the Drosophila Ets transcription factors YAN and POINTED is required for eliciting appropriate responses to Receptor Tyrosine Kinase (RTK) signaling. YAN, a transcriptional repressor, and POINTED, a transcriptional activator, compete for regulatory regions of common target genes, with the ultimate outcome likely influenced by context-specific interactions with binding partners such as MAE. Previous work in cultured cells has led us to propose that MAE attenuates the transcriptional activity of both YAN and POINTED, although its effects on POINTED remain controversial. Here we describe a new layer of complexity to this regulatory hierarchy whereby mae expression is itself directly regulated by the opposing action of YAN and POINTED. In addition, we report that MAE can antagonize POINTED function during eye development; a finding that suggests MAE operates as a dual positive and negative regulator of RTK-mediated signaling in vivo. Together our results lead us to propose that a combination of protein-protein and transcriptional interactions between MAE, YAN and POINTED establishes a complex regulatory circuit that ensures that both down-regulation and activation of the RTK pathway occur appropriately according to specific developmental context.

DNA-Binding Proteins↗

PubMed2003

The transcription factor Eyes absent is a protein tyrosine phosphatase.

Post-translational modifications provide sensitive and flexible mechanisms to dynamically modulate protein function in response to specific signalling inputs. In the case of transcription factors, changes in phosphorylation state can influence protein stability, conformation, subcellular localization, cofactor interactions, transactivation potential and transcriptional output. Here we show that the evolutionarily conserved transcription factor Eyes absent (Eya) belongs to the phosphatase subgroup of the haloacid dehalogenase (HAD) superfamily, and propose a function for it as a non-thiol-based protein tyrosine phosphatase. Experiments performed in cultured Drosophila cells and in vitro indicate that Eyes absent has intrinsic protein tyrosine phosphatase activity and can autocatalytically dephosphorylate itself. Confirming the biological significance of this function, mutations that disrupt the phosphatase active site severely compromise the ability of Eyes absent to promote eye specification and development in Drosophila. Given the functional importance of phosphorylation-dependent modulation of transcription factor activity, this evidence for a nuclear transcriptional coactivator with intrinsic phosphatase activity suggests an unanticipated method of fine-tuning transcriptional regulation.

Amino Acid Motifs↗

PubMed2003

The novel plant homeodomain protein rhinoceros antagonizes Ras signaling in the Drosophila eye.

The sequential specification of cell fates in the Drosophila eye requires repeated activation of the epidermal growth factor receptor (EGFR)/Ras/MAP kinase (MAPK) pathway. Equally important are the multiple layers of inhibitory regulation that prevent excessive or inappropriate signaling. Here we describe the molecular and genetic analysis of a previously uncharacterized gene, rhinoceros (rno), that we propose functions to restrict EGFR signaling in the eye. Loss of rno results in the overproduction of photoreceptors, cone cells, and pigment cells and a corresponding reduction in programmed cell death, all phenotypes characteristic of hyperactivated EGFR signaling. Genetic interactions between rno and multiple EGFR pathway components support this hypothesis. rno encodes a novel but evolutionarily conserved nuclear protein with a PHD zinc-finger domain, a motif commonly found in chromatin-remodeling factors. Future analyses of rno will help to elucidate the regulatory strategies that modulate EGFR signaling in the fly eye.

Amino Acid Sequence↗

PubMed2003

Functional dissection of eyes absent reveals new modes of regulation within the retinal determination gene network.

The retinal determination (RD) gene network encodes a group of transcription factors and cofactors necessary for eye development. Transcriptional and posttranslational regulation of RD family members is achieved through interactions within the network and with extracellular signaling pathways, including epidermal growth factor receptor/RAS/mitogen-activated protein kinase (MAPK), transforming growth factor beta/DPP, Wingless, Hedgehog, and Notch. Here we present the results of structure-function analyses that reveal novel aspects of Eyes absent (EYA) function and regulation. We find that the conserved C-terminal EYA domain negatively regulates EYA transactivation potential, and that GROUCHO-SINE OCULIS (SO) interactions provide another mechanism for negative regulation of EYA-SO target genes. We have mapped the transactivation potential of EYA to an internal proline-, serine-, and threonine-rich region that includes the EYA domain 2 (ED2) and two MAPK phosphorylation consensus sites and demonstrate that activation of the RAS/MAPK pathway potentiates transcriptional output of EYA and the EYA-SO complex in certain contexts. Drosophila S2 cell two-hybrid assays were used to describe a novel homotypic interaction that is mediated by EYA's N terminus. Our data suggest that EYA requires homo- and heterotypic interactions and RAS/MAPK signaling responsiveness to ensure context-appropriate RD gene network activity.

Animals↗