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Alexandra L Joyner - One of the best experts on this subject based on the ideXlab platform.

  • titration of gli3 repressor activity by sonic hedgehog signaling is critical for maintaining multiple adult neural stem cell and astrocyte functions
    The Journal of Neuroscience, 2013
    Co-Authors: Ralitsa Petrova, Alexandra L Joyner, A D R Garcia
    Abstract:

    Sonic hedgehog (SHH), a key regulator of embryonic neurogenesis, signals directly to neural stem cells (NSCs) in the subventricular zone (SVZ) and to astrocytes in the adult mouse forebrain. The specific mechanism by which the GLI2 and GLI3 transcriptional activators (GLI2A and GLI3A) and repressors (GLI2R and GLI3R) carry out SHH signaling has not been addressed. We found that the majority of slow-cycling NSCs express Gli2 and Gli3, whereas GLI1 is restricted ventrally and all three genes are downregulated when NSCs transition into proliferating progenitors. Surprisingly, whereas conditional ablation of Smo in postnatal glial fibrillary acidic protein-expressing cells results in cell-autonomous loss of NSCs and a progressive reduction in SVZ proliferation, without an increase in glial cell production, removal of Gli2 or Gli3 does not alter adult SVZ neurogenesis. Significantly, removing Gli3 in Smo conditional mutants largely rescues neurogenesis and, conversely, expression of a constitutive GLI3R in the absence of normal Gli2 and Gli3 abrogates neurogenesis. Thus unattenuated GLI3R is a primary inhibitor of adult SVZ NSC function. Ablation of Gli2 and Gli3 revealed a minor role for GLI2R and little requirement for GLIA function in stimulating SVZ neurogenesis. Moreover, we found that similar rules of GLI activity apply to SHH signaling in regulating SVZ-derived olfactory bulb interneurons and maintaining cortical astrocyte function. Namely, fewer superficial olfactory bulb interneurons are generated in the absence of Gli2 and Gli3, whereas astrocyte partial gliosis results from an increase in GLI3R. Thus precise titration of GLIR levels by SHH is critical to multiple functions of adult NSCs and astrocytes.

  • limb anterior posterior polarity integrates activator and repressor functions of gli2 as well as gli3
    Developmental Biology, 2012
    Co-Authors: Megan Bowers, Liane Eng, Zhimin Lao, Rowena Turnbull, Xiaozhong Bao, Elyn Riedel, Susan Mackem, Alexandra L Joyner
    Abstract:

    Anterior-posterior (AP) limb patterning is directed by sonic hedgehog (SHH) signaling from the posteriorly located zone of polarizing activity (ZPA). GLI3 and GLI2 are the transcriptional mediators generally utilized in SHH signaling, and each can function as an activator (A) and repressor (R). Although GLI3R has been suggested to be the primary effector of SHH signaling during limb AP patterning, a role for GLI3A or GLI2 has not been fully ruled out, nor has it been determined whether Gli3 plays distinct roles in limb development at different stages. By conditionally removing Gli3 in the limb at multiple different time points, we uncovered four Gli3-mediated functions in limb development that occur at distinct but partially over-lapping time windows: AP patterning of the proximal limb, AP patterning of the distal limb, regulation of digit number and bone differentiation. Furthermore, by removing Gli2 in Gli3 temporal conditional knock-outs, we uncovered an essential role for Gli2 in providing the remaining posterior limb patterning seen in Gli3 single mutants. To test whether GLIAs or GLIRs regulate different aspects of AP limb patterning and/or digit number, we utilized a knock-in allele in which GLI1, which functions solely as an activator, is expressed in place of the bifunctional GLI2 protein. Interestingly, we found that GLIAs contribute to AP patterning specifically in the posterior limb, whereas GLIRs predominantly regulate anterior patterning and digit number. Since GLI3 is a more effective repressor, our results explain why GLI3 is required only for anterior limb patterning and why GLI2 can compensate for GLI3A in posterior limb patterning. Taken together, our data suggest that establishment of a complete range of AP positional identities in the limb requires integration of the spatial distribution, timing, and dosage of GLI2 and GLI3 activators and repressors.

  • sonic hedgehog signaling regulates gli2 transcriptional activity by suppressing its processing and degradation
    Molecular and Cellular Biology, 2006
    Co-Authors: Yong Pan, Alexandra L Joyner, Chunyang Brian Bai, Baolin Wang
    Abstract:

    Gli2 and Gli3 are the primary transcription factors that mediate Sonic hedgehog (Shh) signals in the mouse. Gli3 mainly acts as a transcriptional repressor, because the majority of full-length Gli3 protein is proteolytically processed. Gli2 is mostly regarded as a transcriptional activator, even though it is also suggested to have a weak repressing activity. What the molecular basis for its possible dual function is and how its activity is regulated by Shh signaling are largely unknown. Here we demonstrate that unlike the results seen with Gli3 and Cubitus Interruptus, the fly homolog of Gli, only a minor fraction of Gli2 is proteolytically processed to form a transcriptional repressor in vivo and that in addition to being processed, Gli2 full-length protein is readily degraded. The degradation of Gli2 requires the phosphorylation of a cluster of numerous serine residues in its carboxyl terminus by protein kinase A and subsequently by casein kinase 1 and glycogen synthase kinase 3. The phosphorylated Gli2 interacts directly with betaTrCP in the SCF ubiquitin-ligase complex through two binding sites, which results in Gli2 ubiquitination and subsequent degradation by the proteasome. Both processing and degradation of Gli2 are suppressed by Shh signaling in vivo. Our findings provide the first demonstration of a molecular mechanism by which the Gli2 transcriptional activity is regulated by Shh signaling.

  • spatial pattern of sonic hedgehog signaling through gli genes during cerebellum development
    Development, 2004
    Co-Authors: Jomichelle D Corrales, Gina L Rocco, Sandra Blaess, Alexandra L Joyner
    Abstract:

    The cerebellum consists of a highly organized set of folia that are largely generated postnatally during expansion of the granule cell precursor (GCP) pool. Since the secreted factor sonic hedgehog (Shh) is expressed in Purkinje cells and functions as a GCP mitogen in vitro, it is possible that Shh influences foliation during cerebellum development by regulating the position and/or size of lobes. We studied how Shh and its transcriptional mediators, the Gli proteins, regulate GCP proliferation in vivo, and tested whether they influence foliation. We demonstrate that Shh expression correlates spatially and temporally with foliation. Expression of the Shh target gene GLI1 is also highest in the anterior medial cerebellum, but is restricted to proliferating GCPs and Bergmann glia. By contrast, Gli2 is expressed uniformly in all cells in the developing cerebellum except Purkinje cells and Gli3 is broadly expressed along the anteroposterior axis. Whereas Gli mutants have a normal cerebellum, Gli2 mutants have greatly reduced foliation at birth and a decrease in GCPs. In a complementary study using transgenic mice, we show that overexpressing Shh in the normal domain does not grossly alter the basic foliation pattern, but does lead to prolonged proliferation of GCPs and an increase in the overall size of the cerebellum. Taken together, these studies demonstrate that positive Shh signaling through Gli2 is required to generate a sufficient number of GCPs for proper lobe growth.

  • all mouse ventral spinal cord patterning by hedgehog is gli dependent and involves an activator function of gli3
    Developmental Cell, 2004
    Co-Authors: Brian C Bai, Alexandra L Joyner, Daniel Stephen
    Abstract:

    An important question is how the gradient of Hedgehog is interpreted by cells at the level of the Gli transcription factors. The full range of Gli activity and its dependence on Hh have not been determined, although the Gli2 activator and Gli3 repressor have been implicated. Using the spinal cord as a model system, we demonstrate that Gli3 can transduce Hedgehog signaling as an activator. All expression of the Hh target gene GLI1 is dependent on both Gli2 and Gli3. Unlike Gli2, however, Gli3 requires endogenous GLI1 for induction of floor plate and V3 interneurons. Strikingly, embryos lacking all Gli function develop motor neurons and three ventral interneuron subtypes, similar to embryos lacking Hh signaling and Gli3. Therefore, in the spinal cord all Hh signaling is Gli dependent. Furthermore, a combination of Gli2 and Gli3 is required to regulate motor neuron development and spatial patterning of ventral spinal cord progenitors.

Ruiz A I Altaba - One of the best experts on this subject based on the ideXlab platform.

  • gli2 functions in fgf signaling during antero posterior patterning
    Development, 2000
    Co-Authors: R Brewster, Jose L Mullor, Ruiz A I Altaba
    Abstract:

    Patterning along the anteroposterior (A-P) axis involves the interplay of secreted and transcription factors that specify cell fates in the mesoderm and neuroectoderm. While FGF and homeodomain proteins have been shown to play different roles in posterior specification, the network coordinating their effects remains elusive. Here we have analyzed the function of Gli zinc-finger proteins in mesodermal A-P patterning. We find that Gli2 is sufficient to induce ventroposterior development, functioning in the FGF-brachyury regulatory loop. Gli2 directly induces brachyury, a gene required and sufficient for mesodermal development, and Gli2 is in turn induced by FGF signaling. Moreover, the homeobox gene Xhox3, a critical determinant of posterior development, is also directly regulated by Gli2. Gli3, but not GLI1, has an activity similar to that of Gli2 and is expressed in ventroposterior mesoderm after Gli2. These findings uncover a novel function of Gli proteins, previously only known to mediate hedgehog signals, in the maintenance and patterning of the embryonic mesoderm. More generally, our results suggest a molecular basis for an integration of FGF and hedgehog inputs in Gli-expressing cells that respond to these signals.

  • expression of the vertebrate gli proteins in drosophila reveals a distribution of activator and repressor activities
    Development, 2000
    Co-Authors: Pedro Azablanc, Ruiz A I Altaba, Thomas B Kornberg
    Abstract:

    The Cubitus interruptus (Ci) and Gli proteins are transcription factors that mediate responses to Hedgehog proteins (Hh) in flies and vertebrates, respectively. During development of the Drosophila wing, Ci transduces the Hh signal and regulates transcription of different target genes at different locations. In vertebrates, the three Gli proteins are expressed in overlapping domains and are partially redundant. To assess how the vertebrate Glis correlate with Drosophila Ci, we expressed each in Drosophila and monitored their behaviors and activities. We found that each Gli has distinct activities that are equivalent to portions of the regulatory arsenal of Ci. Gli2 and GLI1 have activator functions that depend on Hh. Gli2 and Gli3 are proteolyzed to produce a repressor form able to inhibit hh expression. However, while Gli3 repressor activity is regulated by Hh, Gli2 repressor activity is not. These observations suggest that the separate activator and repressor functions of Ci are unevenly partitioned among the three Glis, yielding proteins with related yet distinct properties.

  • gli proteins encode context dependent positive and negative functions implications for development and disease
    Development, 1999
    Co-Authors: Ruiz A I Altaba
    Abstract:

    Several lines of evidence implicate zinc finger proteins of the Gli family in the final steps of Hedgehog signaling in normal development and disease. C-terminally truncated mutant GLI3 proteins are also associated with human syndromes, but it is not clear whether these C-terminally truncated Gli proteins fulfil the same function as full-length ones. Here, structure-function analyses of Gli proteins have been performed using floor plate and neuronal induction assays in frog embryos, as well as induction of alkaline phosphatase (AP) in SHH-responsive mouse C3H10T1/2 (10T1/2) cells. These assays show that C-terminal sequences are required for positive inducing activity and cytoplasmic localization, whereas N-terminal sequences determine dominant negative function and nuclear localization. Analyses of nuclear targeted GLI1 and Gli2 proteins suggest that both activator and dominant negative proteins are modified forms. In embryos and COS cells, tagged Gli cDNAs yield C-terminally deleted forms similar to that of Ci. These results thus provide a molecular basis for the human Polydactyly type A and Pallister-Hall Syndrome phenotypes, derived from the deregulated production of C-terminally truncated GLI3 proteins. Analyses of full-length Gli function in 10T1/2 cells suggest that nuclear localization of activating forms is a regulated event and show that only GLI1 mimics SHH in inducing AP activity. Moreover, full-length Gli3 and all C-terminally truncated forms act antagonistically whereas Gli2 is inactive in this assay. In 10T1/2 cells, protein kinase A (PKA), a known inhibitor of Hh signaling, promotes Gli3 repressor formation and inhibits GLI1 function. Together, these findings suggest a context-dependent functional divergence of Gli protein function, in which a cell represses Gli3 and activates GLI1/2 prevents the formation of repressor Gli forms to respond to Shh. Interpretation of Hh signals by Gli proteins therefore appears to involve a fine balance of divergent functions within each and among different Gli proteins, the misregulation of which has profound biological consequences.

  • combinatorial gli gene function in floor plate and neuronal inductions by sonic hedgehog
    Development, 1998
    Co-Authors: Ruiz A I Altaba
    Abstract:

    Within the developing vertebrate nervous system, it is not known how progenitor cells interpret the positional information provided by inducing signals or how the domains in which distinct groups of neural cells differentiate are defined. Gli proteins may be involved in these processes. In the frog neural plate, we have previously shown that the zinc finger transcription factor GLI1 is expressed in midline cells and mediates the effects of Shh inducing floor plate differentiation. In contrast, Gli2 and Gli3 are expressed throughout the neural plate except for the midline. Here, it is shown that Gli3 and Shh repress each other whereas Gli2, like GLI1, is a target of Shh signaling. However, only GLI1 can induce the differentiation of floor plate cells. In addition, Gli2 and Gli3 repress the ectopic induction of floor plate cells by GLI1 in co-injection assays and inhibit endogenous floor plate differentiation. The definition of the floor plate domain, therefore, appears to be defined by the antagonizing activities of Gli2 and Gli3 on GLI1 function. Because both GLI1 and Gli2 are induced by Shh, these results establish a regulatory feedback loop triggered by Shh that restricts floor plate cells to the midline. We have also previously shown that the Gli genes induce neuronal differentiation and here it is shown that there is specificity to the types of neurons the Gli proteins induce. Only GLI1 induces Nkx2.1/TTF-1(+) ventral forebrain neurons. Moreover, Gli2 and Gli3 inhibit their differentiation. In contrast, the differentiation of spinal motor neurons can be induced by the two ventrally expressed Gli genes, GLI1 and Gli2, suggesting that Gli2 directly mediates induction of motor neurons by Shh. In addition, Gli3 inhibits motor neuron differentiation by Gli2. Thus, combinatorial Gli function may pattern the neural tube, integrating positional information and cell type differentiation.

  • activation of the transcription factor GLI1 and the sonic hedgehog signalling pathway in skin tumours
    Nature, 1997
    Co-Authors: Nadia Dahmane, Jeffrey D Lee, P Robins, P Heller, Ruiz A I Altaba
    Abstract:

    Sporadic basal cell carcinoma (BCC) is the most common type of malignant cancer in fair-skinned adults. Familial BCCs and a fraction of sporadic BCCs have lost the function of Patched (Ptc), a Sonic hedgehog (Shh) receptor that acts negatively on this signalling pathway. Overexpression of Shh can induce BCCs in mice. Here we show that ectopic expression of the zinc-finger transcription factor GLI1 in the embryonic frog epidermis results in the development of tumours that express endogenous GLI1. We also show that Shh and the Gli genes are normally expressed in hair follicles, and that human sporadic BCCs consistently express GLI1 but not Shh or Gli3. Because GLI1, but not Gli3, acts as a target and mediator of Shh signalling, our results suggest that expression of GLI1 in basal cells induces BCC formation. Moreover, loss of Ptc or overexpression of Shh cannot be the sole causes of GLI1 induction and sporadic BCC formation, as they do not occur consistently. Thus any mutations leading to the expression of GLI1 in basal cells are predicted to induce BCC formation.

Fritz Aberger - One of the best experts on this subject based on the ideXlab platform.

  • gli2 specific transcriptional activation of the bone morphogenetic protein activin antagonist follistatin in human epidermal cells
    Journal of Biological Chemistry, 2008
    Co-Authors: Thomas Eichberger, Fritz Aberger, Alexandra Kaser, Claudia Pixner, Carmen Schmid, Stefan Klingler, Martina Winklmayr, Cornelia Hauserkronberger, Annamaria Frischauf
    Abstract:

    Hedgehog (HH) signaling in the epidermis is primarily mediated by the zinc finger transcription factors GLI1 and GLI2. Exquisite regulation of HH/GLI signaling is crucial for proper specification of the epidermal lineage and development of its derivatives, whereas dysregulation of HH/GLI signaling disrupts tissue homeostasis and causes basal cell carcinoma (BCC). Similarly, bone morphogenetic proteins (BMPs) and activins have been described as key signaling factors in the complex regulation of epidermal fate decisions, although their precise interplay with HH/GLI is largely elusive. Here we show that, in human epidermal cells, expression of the activin/BMP antagonist follistatin (FST) is predominantly up-regulated by the HH effector GLI2. Consistently, we found strong FST expression in the outer root sheath of human hair follicles and BCC. Detailed promoter analysis showed that two sequences with homology to the GLI consensus binding site are required for GLI2-mediated activation. Interestingly, activation of the FST promoter is highly GLI2-specific, because neither GLI1 nor GLI3 can significantly increase FST transcription. GLI2 specificity requires the presence of a 518-bp fragment in the proximal FST promoter region. On the protein level, sequences C-terminal to the zinc finger are responsible for GLI2-specific activation of FST transcription, pointing to the existence of GLI-interacting cofactors that modulate GLI target specificity. Our results reveal a key role of GLI2 in activation of the activin/BMP antagonist FST in response to HH signaling and provide new evidence for a regulatory interaction between HH and activin/BMP signaling in hair follicle development and BCC.

  • overlapping and distinct transcriptional regulator properties of the GLI1 and gli2 oncogenes
    Genomics, 2006
    Co-Authors: Thomas Eichberger, Gerhard Regl, Maria Kasper, Fritz Aberger, Alexandra Kaser, Carmen Schmid, Veronika Sander, Harald Schnidar, Sandra Plamberger, Annamaria Frischauf
    Abstract:

    The GLI transcription factors mediate the hedgehog signal in development and carcinogenesis. Basal cell carcinoma can be caused by overexpression of either GLI1 or GLI2. Though GLI1 and GLI2 have identical or very similar DNA binding specificities, some of their activities are overlapping, some are clearly distinct. We analyzed target gene specificities of GLI1 and constitutively active GLI2 (GLI2ΔN) by global expression profiling in an inducible, well-characterized HaCaT keratinocyte expression system. Four hundred fifty-six genes up- or downregulated at least twofold were identified. GLI target gene profiles correlated well with the biological activities of these transcription factors in hair follicles and basal cell carcinoma. Upregulation of largely overlapping sets of target genes was effected by both factors, repression occurred predominantly in response to GLI2. Also, significant quantitative differences in response to GLI1 and GLI2ΔN were found for a small number of activated genes. Since we have not detected a putative processed GLI2 repressor, these results point to specific but indirect target gene repression by GLI2ΔN via preferential activation of one or more negative regulators.

  • activation of the bcl2 promoter in response to hedgehog gli signal transduction is predominantly mediated by gli2
    Cancer Research, 2004
    Co-Authors: Gerhard Regl, Graham W Neill, Thomas Eichberger, Maria Kasper, Annamaria Frischauf, Cornelia Hauserkronberger, Michael P Philpott, Harald Schnidar, Harald Esterbauer, Fritz Aberger
    Abstract:

    Aberrant activation of the Hedgehog (HH)/GLI signaling pathway has been implicated in the development of basal cell carcinoma (BCC). The zinc finger transcription factors GLI1 and GLI2 are considered mediators of the HH signal in epidermal cells, although their tumorigenic nature and their relative contribution to tumorigenesis are only poorly understood. To shed light on the respective role of these transcription factors in epidermal neoplasia, we screened for genes preferentially regulated either by GLI1 or GLI2 in human epidermal cells. We show here that expression of the key antiapoptotic factor BCL2 is predominantly activated by GLI2 compared with GLI1. Detailed promoter analysis and gel shift assays identified three GLI binding sites in the human BCL2 cis-regulatory region. We found that one of these binding sites is critical for conferring GLI2-specific activation of the human BCL2 promoter and that the selective induction of BCL2 expression depends on the zinc finger DNA binding domain of GLI2. In vivo, GLI2 and BCL2 were coexpressed in the outer root sheath of hair follicles and BCC and in plasma cells that infiltrated BCC tumor islands. On the basis of the latter observation, we analyzed plasma cell-derived tumors and found strong expression of GLI2 and BCL2 in neoplastic cells of plasmacytoma patients, implicating HH/GLI signaling in the development of plasma cell-derived malignancies. The results reveal a central role for GLI2 in activating the prosurvival factor BCL2, which may represent an important mechanism in the development or maintenance of cancers associated with inappropriate HH signaling.

  • gli2 is expressed in normal human epidermis and bcc and induces GLI1 expression by binding to its promoter
    Journal of Investigative Dermatology, 2004
    Co-Authors: Mohammed S Ikram, Gerhard Regl, Graham W Neill, Thomas Eichberger, Anthony G Quinn, Annamaria Frischauf, Fritz Aberger, Michael P Philpott
    Abstract:

    Sonic hedgehog (Shh) binds to its receptor patched (PTCH), leading to the activation and repression of target genes via the GLI family of zinc-finger transcription factors. Deregulation of the Shh pathway is associated with basal cell carcinoma (BCC) due to upregulation of GLI1 and GLI2. We recently demonstrated a positive feedback loop between GLI1 and GLI2, which revealed that GLI1 may be a direct target of GLI2. Using band shift and luciferase reporter assays, we now show that GLI2 binds the GLI-binding consensus sequence in the GLI1 promoter. These data suggest that GLI2 directly activates GLI1 and that retrovirally expressed GLI2 induces expression of endogenous GLI1 in human primary keratinocytes. Finally, using in situ hybridization, we show that GLI2 is expressed in the interfollicular epidermis and the outer root sheath of hair follicles in normal skin as well as in BCC tumor islands. These results suggest an important role for GLI2 in regulating epidermal proliferation and skin tumorigenesis.

  • human gli2 and GLI1 are part of a positive feedback mechanism in basal cell carcinoma
    Oncogene, 2002
    Co-Authors: Gerhard Regl, Graham W Neill, Thomas Eichberger, Maria Kasper, Mohammed S Ikram, J Koller, Helmut Hintner, Anthony G Quinn, Annamaria Frischauf, Fritz Aberger
    Abstract:

    Transgenic mouse models have provided evidence that activation of the zinc-finger transcription factor GLI1 by Hedgehog (Hh)-signalling is a key step in the initiation of the tumorigenic programme leading to Basal Cell Carcinoma (BCC). However, the downstream events underlying Hh/GLI-induced BCC development are still obscure. Using in vitro model systems to analyse the eect of Hh/GLI-signalling in human keratinocytes, we identified a positive feedback mechanism involving the zinc finger transcription factors GLI1 and GLI2. Expression of GLI1 in human keratinocytes induced the transcriptional activator isoforms GLI2a and GLI2b. Both isoforms were also shown to be expressed at elevated levels in 21 BCCs compared to normal skin. Detailed time course experiments monitoring the transcriptional response of keratinocytes either to GLI1 or to GLI2 suggest that GLI1 is a direct target of GLI2, while activation of GLI2 by GLI1 is likely to be indirect. Furthermore, expression of either GLI2 or GLI1 led to an increase in DNA-synthesis in confluent human keratinocytes. Taken together, these results suggest an important role of the positive GLI1-GLI2 feedback loop in Hh-mediated epidermal cell proliferation. Oncogene (2002) 21, 5529‐5539. doi:10.1038/sj.onc. 1205748

Chichung Hui - One of the best experts on this subject based on the ideXlab platform.

  • kidney patterning genes disrupts renal morphogenesis
    2014
    Co-Authors: Sita Bhella, Christopher W Wilson, Paotien Chuang, Chichung Hui
    Abstract:

    Truncating mutations in Gli3, an intracellular effector in the SHH-SMO-GLI signaling pathway, cause renal aplasia/dysplasia in humans and mice. Yet, the pathogenic mechanisms are undefined. Here, we report the effect of decreased SHH-SMO signaling on renal morphogenesis, the expression of SHH target genes and GLI binding to Shh target genes. Shh deficiency or cyclopaminemediated SMO inhibition disrupted renal organogenesis, decreased expression of GLI1 and GLI2 proteins, but increased expression of GLI3 repressor relative to GLI3 activator. Shh deficiency decreased expression of kidney patterning genes (Pax2 and Sall1) and cell cycle regulators (cyclin D1 and MYCN). Elimination of Gli3 in Shh –/ – mice rescued kidney malformation and restored expression of Pax2, Sall1, cyclin D1, MYCN, GLI1 and Gli2. To define mechanisms by which SHH-SMO signaling controls gene expression, we determined the binding of GLI proteins to 5 � flanking regions containing GLI consensus binding sequences in Shh target genes using chromatin immunoprecipitation. In normal embryonic kidney tissue, GLI1 and/or GLI2 were bound to each target gene. By contrast, treatment of embryonic kidney explants with cyclopamine decreased GLI1 and/or GLI2 binding, and induced binding of GLI3. However, cyclopamine failed to decrease GLI1 and Gli2 expression and branching morphogenesis in Gli3-deficient embryonic kidney tissue. Together, these results demonstrate that SHH-SMO signaling controls renal morphogenesis via transcriptional control of Gli, renal patterning and cell cycle regulator genes in a manner that is opposed by GLI3. KEY WORDS: Kidney development, Sonic Hedgehog, GLI

  • gli3 dependent transcriptional repression of GLI1 gli2 and kidney patterning genes disrupts renal morphogenesis
    Development, 2006
    Co-Authors: Sita Bhella, Christopher W Wilson, Paotien Chuang, Chichung Hui
    Abstract:

    Truncating mutations in Gli3 , an intracellular effector in the SHH-SMO-GLI signaling pathway, cause renal aplasia/dysplasia in humans and mice. Yet, the pathogenic mechanisms are undefined. Here, we report the effect of decreased SHH-SMO signaling on renal morphogenesis, the expression of SHH target genes and GLI binding to Shh target genes. Shh deficiency or cyclopamine-mediated SMO inhibition disrupted renal organogenesis, decreased expression of GLI1 and GLI2 proteins, but increased expression of GLI3 repressor relative to GLI3 activator. Shh deficiency decreased expression of kidney patterning genes ( Pax2 and Sall1 ) and cell cycle regulators (cyclin D1 and MYCN). Elimination of Gli3 in Shh–/– mice rescued kidney malformation and restored expression of Pax2, Sall1 , cyclin D1, MYCN, GLI1 and Gli2 . To define mechanisms by which SHH-SMO signaling controls gene expression, we determined the binding of GLI proteins to 5′ flanking regions containing GLI consensus binding sequences in Shh target genes using chromatin immunoprecipitation. In normal embryonic kidney tissue, GLI1 and/or GLI2 were bound to each target gene. By contrast, treatment of embryonic kidney explants with cyclopamine decreased GLI1 and/or GLI2 binding, and induced binding of GLI3. However, cyclopamine failed to decrease GLI1 and Gli2 expression and branching morphogenesis in Gli3 -deficient embryonic kidney tissue. Together, these results demonstrate that SHH-SMO signaling controls renal morphogenesis via transcriptional control of Gli , renal patterning and cell cycle regulator genes in a manner that is opposed by GLI3.

  • negative regulation of GLI1 and gli2 activator function by suppressor of fused through multiple mechanisms
    Differentiation, 2005
    Co-Authors: Paul C Barnfield, Xiaoyun Zhang, Vijitha Thanabalasingham, Minoru Yoshida, Chichung Hui
    Abstract:

    During animal development, the Hedgehog (Hh) signal transduction pathway plays critical roles in cell fate determination and tissue patterning. In humans, aberrant Hh signaling has been linked to several genetic disorders and cancers. Binding of Hh to its receptor initiates a signaling cascade, which ultimately results in the activation of the Gli/Ci transcription factors. Suppressor of fused (Su(fu)) is a Gli/Ci-interacting protein, which acts as a negative regulator of Hh signaling in Drosophila and vertebrates. Su(fu) is also implicated as a tumor suppressor as its mutations have been found in medulloblastoma and prostate cancer. Su(fu) is thought to act by preventing the nuclear accumulation of Gli/Ci, however, mechanistic insight into its mode of action has remained elusive. We demonstrate here that Su(fu) prevents the nuclear accumulation of GLI1 and Gli2 through multiple mechanisms. While Su(fu) itself is not subject to CRM1-dependent regulation, Su(fu) sequesters GLI1 in the cytoplasm mostly through a mechanism that depends on the activity of the nuclear export protein CRM1. In contrast, CRM1-mediated export is not required for Su(fu) to sequester Gli2. Furthermore, we show that the N-terminus of Su(fu) is sufficient for Gli inactivation in the absence of cytoplasmic sequestration. Together, these observations reveal that Su(fu) regulates the activity of GLI1 and Gli2 through distinct cytoplasmic and nuclear mechanisms.

  • gli2 and gli3 have redundant and context dependent function in skeletal muscle formation
    Development, 2005
    Co-Authors: Aileen Mcdermott, Charles P Emerson, Chichung Hui, Marcus K Gustafsson, Thomas Elsam, Anne-gaëlle Borycki
    Abstract:

    The Gli family of zinc finger transcription factors are mediators of Shh signalling in vertebrates. In previous studies, we showed that Shh signalling, via an essential Gli -binding site in the Myf5 epaxial somite (ES) enhancer, is required for the specification of epaxial muscle progenitor cells. Shh signalling is also required for the normal mediolateral patterning of myogenic cells within the somite. In this study, we investigate the role and the transcriptional activities of Gli proteins during somite myogenesis in the mouse embryo. We report that Gli genes are differentially expressed in the mouse somite. Gli2 and Gli3 are essential for GLI1 expression in somites, establishing Gli2 and Gli3 as primary mediators and GLI1 as a secondary mediator of Shh signalling. Combining genetic studies with the use of a transgenic mouse line expressing a reporter gene under the control of the Myf5 epaxial somite enhancer, we show that Gli2 or Gli3 is required for Myf5 activation in the epaxial muscle progenitor cells. Furthermore, Gli3, but not Gli2 represses Myf5 transcription in a dose-dependent manner in the absence of Shh. Finally, we provide evidence that hypaxial and myotomal gene expression is mispatterned in Gli2–/–Gli3–/– and Gli3–/–Shh–/– somites. Together, our data demonstrate both positive and negative regulatory functions for Gli2 and Gli3 in the control of Myf5 activation in the epaxial muscle progenitor cells and in dorsoventral and mediolateral patterning of the somite.

  • Interplays of Gli2 and Gli3 and their requirement in mediating Shh-dependent sclerotome induction.
    Development (Cambridge England), 2003
    Co-Authors: Laura Buttitta, Chichung Hui, Chen-ming Fan
    Abstract:

    Sonic hedgehog (Shh) signaling is essential for sclerotome development in the mouse. Gli2 and Gli3 are thought to be the primary transcriptional mediators of Shh signaling; however, their roles in Shh induction of sclerotomal genes have not been investigated. Using a combination of mutant analysis and in vitro explant assays, we demonstrate that Gli2 and Gli3 are required for Shh-dependent sclerotome induction. Gli2 –/– Gli3 –/– embryos exhibit a severe loss of sclerotomal gene expression, and somitic mesoderm from these embryos cannot activate sclerotomal genes in response to exogenous Shh. We find that one copy of either Gli2 or Gli3 is required to mediate Shh induction of sclerotomal markers Pax1 and Pax9 in vivo and in vitro. Although Gli2 is generally considered an activator and Gli3 a repressor, our results also reveal a repressor function for Gli2 and an activator function for Gli3 in the developing somite. To further dissect the function of each Gli, we used adenovirus to overexpress GLI1, Gli2 and Gli3 in presomitic mesoderm explants. We find that each Gli preferentially activates a distinct set of Shh target genes, suggesting that the functions of Shh in patterning, growth and negative feedback are divided preferentially between different Gli proteins in the somite.

Gerhard Regl - One of the best experts on this subject based on the ideXlab platform.

  • overlapping and distinct transcriptional regulator properties of the GLI1 and gli2 oncogenes
    Genomics, 2006
    Co-Authors: Thomas Eichberger, Gerhard Regl, Maria Kasper, Fritz Aberger, Alexandra Kaser, Carmen Schmid, Veronika Sander, Harald Schnidar, Sandra Plamberger, Annamaria Frischauf
    Abstract:

    The GLI transcription factors mediate the hedgehog signal in development and carcinogenesis. Basal cell carcinoma can be caused by overexpression of either GLI1 or GLI2. Though GLI1 and GLI2 have identical or very similar DNA binding specificities, some of their activities are overlapping, some are clearly distinct. We analyzed target gene specificities of GLI1 and constitutively active GLI2 (GLI2ΔN) by global expression profiling in an inducible, well-characterized HaCaT keratinocyte expression system. Four hundred fifty-six genes up- or downregulated at least twofold were identified. GLI target gene profiles correlated well with the biological activities of these transcription factors in hair follicles and basal cell carcinoma. Upregulation of largely overlapping sets of target genes was effected by both factors, repression occurred predominantly in response to GLI2. Also, significant quantitative differences in response to GLI1 and GLI2ΔN were found for a small number of activated genes. Since we have not detected a putative processed GLI2 repressor, these results point to specific but indirect target gene repression by GLI2ΔN via preferential activation of one or more negative regulators.

  • activation of the bcl2 promoter in response to hedgehog gli signal transduction is predominantly mediated by gli2
    Cancer Research, 2004
    Co-Authors: Gerhard Regl, Graham W Neill, Thomas Eichberger, Maria Kasper, Annamaria Frischauf, Cornelia Hauserkronberger, Michael P Philpott, Harald Schnidar, Harald Esterbauer, Fritz Aberger
    Abstract:

    Aberrant activation of the Hedgehog (HH)/GLI signaling pathway has been implicated in the development of basal cell carcinoma (BCC). The zinc finger transcription factors GLI1 and GLI2 are considered mediators of the HH signal in epidermal cells, although their tumorigenic nature and their relative contribution to tumorigenesis are only poorly understood. To shed light on the respective role of these transcription factors in epidermal neoplasia, we screened for genes preferentially regulated either by GLI1 or GLI2 in human epidermal cells. We show here that expression of the key antiapoptotic factor BCL2 is predominantly activated by GLI2 compared with GLI1. Detailed promoter analysis and gel shift assays identified three GLI binding sites in the human BCL2 cis-regulatory region. We found that one of these binding sites is critical for conferring GLI2-specific activation of the human BCL2 promoter and that the selective induction of BCL2 expression depends on the zinc finger DNA binding domain of GLI2. In vivo, GLI2 and BCL2 were coexpressed in the outer root sheath of hair follicles and BCC and in plasma cells that infiltrated BCC tumor islands. On the basis of the latter observation, we analyzed plasma cell-derived tumors and found strong expression of GLI2 and BCL2 in neoplastic cells of plasmacytoma patients, implicating HH/GLI signaling in the development of plasma cell-derived malignancies. The results reveal a central role for GLI2 in activating the prosurvival factor BCL2, which may represent an important mechanism in the development or maintenance of cancers associated with inappropriate HH signaling.

  • gli2 is expressed in normal human epidermis and bcc and induces GLI1 expression by binding to its promoter
    Journal of Investigative Dermatology, 2004
    Co-Authors: Mohammed S Ikram, Gerhard Regl, Graham W Neill, Thomas Eichberger, Anthony G Quinn, Annamaria Frischauf, Fritz Aberger, Michael P Philpott
    Abstract:

    Sonic hedgehog (Shh) binds to its receptor patched (PTCH), leading to the activation and repression of target genes via the GLI family of zinc-finger transcription factors. Deregulation of the Shh pathway is associated with basal cell carcinoma (BCC) due to upregulation of GLI1 and GLI2. We recently demonstrated a positive feedback loop between GLI1 and GLI2, which revealed that GLI1 may be a direct target of GLI2. Using band shift and luciferase reporter assays, we now show that GLI2 binds the GLI-binding consensus sequence in the GLI1 promoter. These data suggest that GLI2 directly activates GLI1 and that retrovirally expressed GLI2 induces expression of endogenous GLI1 in human primary keratinocytes. Finally, using in situ hybridization, we show that GLI2 is expressed in the interfollicular epidermis and the outer root sheath of hair follicles in normal skin as well as in BCC tumor islands. These results suggest an important role for GLI2 in regulating epidermal proliferation and skin tumorigenesis.

  • human gli2 and GLI1 are part of a positive feedback mechanism in basal cell carcinoma
    Oncogene, 2002
    Co-Authors: Gerhard Regl, Graham W Neill, Thomas Eichberger, Maria Kasper, Mohammed S Ikram, J Koller, Helmut Hintner, Anthony G Quinn, Annamaria Frischauf, Fritz Aberger
    Abstract:

    Transgenic mouse models have provided evidence that activation of the zinc-finger transcription factor GLI1 by Hedgehog (Hh)-signalling is a key step in the initiation of the tumorigenic programme leading to Basal Cell Carcinoma (BCC). However, the downstream events underlying Hh/GLI-induced BCC development are still obscure. Using in vitro model systems to analyse the eect of Hh/GLI-signalling in human keratinocytes, we identified a positive feedback mechanism involving the zinc finger transcription factors GLI1 and GLI2. Expression of GLI1 in human keratinocytes induced the transcriptional activator isoforms GLI2a and GLI2b. Both isoforms were also shown to be expressed at elevated levels in 21 BCCs compared to normal skin. Detailed time course experiments monitoring the transcriptional response of keratinocytes either to GLI1 or to GLI2 suggest that GLI1 is a direct target of GLI2, while activation of GLI2 by GLI1 is likely to be indirect. Furthermore, expression of either GLI2 or GLI1 led to an increase in DNA-synthesis in confluent human keratinocytes. Taken together, these results suggest an important role of the positive GLI1-GLI2 feedback loop in Hh-mediated epidermal cell proliferation. Oncogene (2002) 21, 5529‐5539. doi:10.1038/sj.onc. 1205748

  • Human GLI2 and GLI1 are part of a positive feedback mechanism in Basal Cell Carcinoma
    Oncogene, 2002
    Co-Authors: Gerhard Regl, Graham W Neill, Thomas Eichberger, Maria Kasper, Mohammed S Ikram, J Koller, Helmut Hintner, Anthony G Quinn, Annamaria Frischauf, Fritz Aberger
    Abstract:

    Transgenic mouse models have provided evidence that activation of the zinc-finger transcription factor GLI1 by Hedgehog (Hh)-signalling is a key step in the initiation of the tumorigenic programme leading to Basal Cell Carcinoma (BCC). However, the downstream events underlying Hh/GLI-induced BCC development are still obscure. Using in vitro model systems to analyse the effect of Hh/GLI-signalling in human keratinocytes, we identified a positive feedback mechanism involving the zinc finger transcription factors GLI1 and GLI2. Expression of GLI1 in human keratinocytes induced the transcriptional activator isoforms GLI2α and GLI2β. Both isoforms were also shown to be expressed at elevated levels in 21 BCCs compared to normal skin. Detailed time course experiments monitoring the transcriptional response of keratinocytes either to GLI1 or to GLI2 suggest that GLI1 is a direct target of GLI2, while activation of GLI2 by GLI1 is likely to be indirect. Furthermore, expression of either GLI2 or GLI1 led to an increase in DNA-synthesis in confluent human keratinocytes. Taken together, these results suggest an important role of the positive GLI1-GLI2 feedback loop in Hh-mediated epidermal cell proliferation.