scholarly journals Smoothened transduces hedgehog signals via activity-dependent sequestration of PKA catalytic subunits

PLoS Biology ◽  
2021 ◽  
Vol 19 (4) ◽  
pp. e3001191
Author(s):  
Corvin D. Arveseth ◽  
John T. Happ ◽  
Danielle S. Hedeen ◽  
Ju-Fen Zhu ◽  
Jacob L. Capener ◽  
...  
Keyword(s):  
Signal Transduction ◽  
Transcription Factors ◽  
Target Genes ◽  
Catalytic Subunits ◽  
Unknown Mechanism ◽  
Gli Transcription Factors ◽  
Intracellular Domains ◽  
G Protein Coupled ◽  
Activity Dependent ◽  
Kinase A

The Hedgehog (Hh) pathway is essential for organ development, homeostasis, and regeneration. Dysfunction of this cascade drives several cancers. To control expression of pathway target genes, the G protein–coupled receptor (GPCR) Smoothened (SMO) activates glioma-associated (GLI) transcription factors via an unknown mechanism. Here, we show that, rather than conforming to traditional GPCR signaling paradigms, SMO activates GLI by binding and sequestering protein kinase A (PKA) catalytic subunits at the membrane. This sequestration, triggered by GPCR kinase (GRK)-mediated phosphorylation of SMO intracellular domains, prevents PKA from phosphorylating soluble substrates, releasing GLI from PKA-mediated inhibition. Our work provides a mechanism directly linking Hh signal transduction at the membrane to GLI transcription in the nucleus. This process is more fundamentally similar between species than prevailing hypotheses suggest. The mechanism described here may apply broadly to other GPCR- and PKA-containing cascades in diverse areas of biology.

scholarly journals Smoothened Transduces Hedgehog Signals via Activity-Dependent Sequestration of PKA Catalytic Subunits

2020 ◽  
Author(s):  
Corvin D. Arveseth ◽  
John T. Happ ◽  
Danielle S. Hedeen ◽  
Ju-Fen Zhu ◽  
Jacob L. Capener ◽  
...  
Keyword(s):  
Signal Transduction ◽  
Transcription Factors ◽  
Target Genes ◽  
Catalytic Subunits ◽  
Unknown Mechanism ◽  
Gli Transcription Factors ◽  
Intracellular Domains ◽  
G Protein Coupled ◽  
Activity Dependent ◽  
Kinase A

ABSTRACTThe Hedgehog (Hh) pathway is essential for organ development, homeostasis, and regeneration. Dysfunction of this cascade drives several cancers. To control expression of pathway target genes, the G protein-coupled receptor (GPCR) Smoothened (SMO) activates glioma-associated (GLI) transcription factors via an unknown mechanism. Here we show that, rather than conforming to traditional GPCR signaling paradigms, SMO activates GLI by binding and sequestering protein kinase A (PKA) catalytic subunits at the membrane. This sequestration, triggered by GPCR kinase 2 (GRK2)-mediated phosphorylation of SMO intracellular domains, prevents PKA from phosphorylating soluble substrates, releasing GLI from PKA-mediated inhibition. Our work provides a mechanism directly linking Hh signal transduction at the membrane to GLI transcription in the nucleus. This process is more fundamentally similar between species than prevailing hypotheses suggest. The mechanism described here may apply broadly to other GPCR- and PKA-containing cascades in diverse areas of biology.

scholarly journals The role of the Hedgehog signaling pathway in cancer: A comprehensive review

2018 ◽  
Vol 18 (1) ◽  
pp. 8-20 ◽  
Author(s):  
Ana Marija Skoda ◽  
Dora Simovic ◽  
Valentina Karin ◽  
Vedran Kardum ◽  
Semir Vranic ◽  
...  
Keyword(s):  
Transcription Factors ◽  
Signaling Pathway ◽  
Hedgehog Signaling ◽  
Target Genes ◽  
Biological Effects ◽  
Signal Transmission ◽  
Adenosine Monophosphate ◽  
Evolutionary Pathway ◽  
Gli Transcription Factors ◽  
Hh Signaling

The Hedgehog (Hh) signaling pathway was first identified in the common fruit fly. It is a highly conserved evolutionary pathway of signal transmission from the cell membrane to the nucleus. The Hh signaling pathway plays an important role in the embryonic development. It exerts its biological effects through a signaling cascade that culminates in a change of balance between activator and repressor forms of glioma-associated oncogene (Gli) transcription factors. The components of the Hh signaling pathway involved in the signaling transfer to the Gli transcription factors include Hedgehog ligands (Sonic Hh [SHh], Indian Hh [IHh], and Desert Hh [DHh]), Patched receptor (Ptch1, Ptch2), Smoothened receptor (Smo), Suppressor of fused homolog (Sufu), kinesin protein Kif7, protein kinase A (PKA), and cyclic adenosine monophosphate (cAMP). The activator form of Gli travels to the nucleus and stimulates the transcription of the target genes by binding to their promoters. The main target genes of the Hh signaling pathway are PTCH1, PTCH2, and GLI1. Deregulation of the Hh signaling pathway is associated with developmental anomalies and cancer, including Gorlin syndrome, and sporadic cancers, such as basal cell carcinoma, medulloblastoma, pancreatic, breast, colon, ovarian, and small-cell lung carcinomas. The aberrant activation of the Hh signaling pathway is caused by mutations in the related genes (ligand-independent signaling) or by the excessive expression of the Hh signaling molecules (ligand-dependent signaling – autocrine or paracrine). Several Hh signaling pathway inhibitors, such as vismodegib and sonidegib, have been developed for cancer treatment. These drugs are regarded as promising cancer therapies, especially for patients with refractory/advanced cancers.

scholarly journals An Efficient Method to Identify Conditionally Activated Transcription Factors and their Corresponding Signal Transduction Pathway Segments

2009 ◽  
Vol 3 ◽  
pp. BBI.S3485
Author(s):  
Haiyan Hu
Keyword(s):  
Signal Transduction ◽  
Transcription Factors ◽  
Efficient Method ◽  
Molecular Mechanisms ◽  
Target Genes ◽  
Signal Transduction Pathway ◽  
Protein Interaction Data ◽  
Transduction Pathway ◽  
Binding Data ◽  
Microarray Datasets

A signal transduction pathway (STP) is a cascade composed of a series of signal transferring steps, which often activate one or more transcription factors (TFs) to control the transcription of target genes. Understanding signaling pathways is important to our understanding of the molecular mechanisms of disease. Many condition-annotated pathways have been deposited in public databases. However, condition-annotated pathways are far from complete, considering the large number of possible conditions. Computational methods to assist in the identification of conditionally activated pathways are greatly needed. In this paper, we propose an efficient method to identify conditionally activated pathway segments starting from the identification of conditionally activated TFs, by incorporating protein-DNA binding data, gene expression data and protein interaction data. Applying our methods on several microarray datasets, we have discovered many significantly activated TFs and their corresponding pathway segments, which are supported by evidence in the literature.

scholarly journals Insights into Functional Connectivity in Mammalian Signal Transduction Pathways by Pairwise Comparison of Protein Interaction Partners of Critical Signaling Hubs

2019 ◽  
Author(s):  
Chilakamarti V. Ramana
Keyword(s):  
Signal Transduction ◽  
Transcription Factors ◽  
Functional Connectivity ◽  
Protein Interaction ◽  
Target Genes ◽  
Pairwise Comparison ◽  
Signal Transduction Pathways ◽  
Critical Nodes ◽  
Interaction Partners ◽  
Signaling Modules

AbstractGrowth factors and cytokines activate signal transduction pathways and regulate gene expression in eukaryotes. Intracellular domains of activated receptors recruit several protein kinases as well as transcription factors that serve as platforms or hubs for the assembly of multi-protein complexes. The signaling hubs involved in a related biologic function often share common interaction proteins and target genes. This functional connectivity suggests that a pairwise comparison of protein interaction partners of signaling hubs and network analysis of common partners and their expression analysis might lead to the identification of critical nodes in cellular signaling. A pairwise comparison of signaling hubs across several related pathways might also reveal novel signaling modules. Analysis of Protein Interaction Connectome by Venn (PIC-VENN) of transcription factors STAT1, STAT3, NFKB1, RELA, FOS and JUN, and their common interaction network suggested that BRCA1 and TSC22D3 function as critical nodes in immune responses by connecting the signaling nodes into signaling modules. Mutations or differential expression levels of these critical nodes in pathological conditions might deregulate signaling pathways and their target genes involved in inflammation. Biological connectivity emerges from the structural connectivity of interaction networks across several signaling hubs in related pathways. Application of PIC-VENN to several signaling hubs might reveal novel nodes and modules that can be targeted to simultaneously activate or inhibit cell signaling in health and disease.

scholarly journals The unfolding stories of GPR30, a new membrane-bound estrogen receptor

2009 ◽  
Vol 204 (2) ◽  
pp. 105-114 ◽  
Author(s):  
Marcello Maggiolini ◽  
Didier Picard
Keyword(s):  
Signal Transduction ◽  
Estrogen Receptor ◽  
Transcription Factors ◽  
Steroid Hormones ◽  
Cell Types ◽  
Nuclear Receptor Superfamily ◽  
Wide Range ◽  
History Of ◽  
Membrane Bound ◽  
G Protein Coupled

Steroid hormones such as estrogens are known to signal through ligand-regulated transcription factors of the nuclear receptor superfamily. In addition, they elicit rapid nongenomic responses from membrane-associated receptors. One of these receptors belongs to an entirely different family of proteins. The G protein-coupled and seven-transmembrane receptor, GPR30, is now widely recognized as an estrogen receptor (ER), hence its official new acronym GPER. It appears to mediate a wide range of responses to estrogen in a large variety of cell types. Its functions are clearly distinct from those of the classical nuclear ERs, although these pathways may overlap and interact in some cases. Here, we review the history of the discovery of this new ER, the evidence for the claim that it is an ER, its signal transduction, and its potential functions in physiology and disease.

scholarly journals DYRK2 is a ciliary kinase involved in vertebrate Hedgehog signal transduction

2020 ◽  
Author(s):  
Nicholas Morante ◽  
Monika Abedin Sigg ◽  
Luke Strauskulage ◽  
David R. Raleigh ◽  
Jeremy F. Reiter
Keyword(s):  
Signal Transduction ◽  
Transcription Factors ◽  
Hedgehog Signaling ◽  
Primary Cilia ◽  
Skeletal Development ◽  
Skeletal Defects ◽  
Gli Transcription Factors ◽  
Hedgehog Signal Transduction ◽  
Hedgehog Signal

ABSTRACTPrimary cilia are organelles specialized for signaling. We previously defined the proteomes of sea urchin and sea anemone cilia to identify ciliary proteins that predate the origin of bilateria. This evolutionary perspective on cilia identified DYRK2, a kinase not been previously implicated in ciliary biology. We found that DYRK2 localizes to cilia and that loss of DYRK2 disrupts ciliary morphology. We also found that DYRK2 participates in ciliary Hh signal transduction, communicating between SMO and GLI transcription factors. Mutation of mouse Dyrk2 resulted in skeletal defects reminiscent of those caused by loss of Indian hedgehog (Ihh). Like Dyrk2 mutations, pharmacological inhibition of DYRK2 dysregulates ciliary length control and attenuates Hedgehog signaling. Thus, DYRK2 is required for ciliary morphology, for Hedgehog signaling in vitro, and for skeletal development. We propose that DYRK2 is part of the mechanism that transduces SMO to activate GLI transcription factors within cilia.

scholarly journals A PKA Inhibitor Motif within Smoothened Controls Hedgehog Signal Transduction

2021 ◽  
Author(s):  
John T. Happ ◽  
Corvin D. Arveseth ◽  
Jessica Bruystens ◽  
Daniela Bertinetti ◽  
Isaac B. Nelson ◽  
...  
Keyword(s):  
Signal Transduction ◽  
Active Site ◽  
Membrane Receptors ◽  
Cancer Pathways ◽  
Gli Transcription Factors ◽  
G Protein Coupled ◽  
Hedgehog Signal Transduction ◽  
Hedgehog Signal ◽  
Pka Catalytic Subunit

The Hedgehog (Hh) cascade is central to development, tissue homeostasis, and cancer. A pivotal step in Hh signal transduction is the activation of GLI transcription factors by the atypical G protein-coupled receptor (GPCR) Smoothened (SMO). How SMO activates GLI has remained unclear for decades. Here we show that SMO employs a decoy substrate sequence to physically block the active site of the PKA catalytic subunit (PKA-C) and extinguish its enzymatic activity. As a result, GLI is released from phosphorylation-induced inhibition. Using a combination of in vitro, cellular, and organismal models, we demonstrate that interfering with SMO / PKA pseudosubstrate interactions prevents Hh signal transduction. The mechanism we uncovered echoes one utilized by the Wnt cascade, revealing an unexpected similarity in how these two essential developmental and cancer pathways signal intracellularly. More broadly, our findings define a new mode of GPCR-PKA communication that may be harnessed by a range of membrane receptors and kinases.

scholarly journals ChIP-exo analysis highlights Fkh1 and Fkh2 transcription factors as hubs that integrate multi-scale networks in budding yeast

2019 ◽  
Vol 47 (15) ◽  
pp. 7825-7841 ◽  
Author(s):  
Thierry D G A Mondeel ◽  
Petter Holland ◽  
Jens Nielsen ◽  
Matteo Barberis
Keyword(s):  
Cell Cycle ◽  
Signal Transduction ◽  
Transcription Factors ◽  
Target Genes ◽  
Budding Yeast ◽  
Biochemical Networks ◽  
Molecular Networks ◽  
Forkhead Transcription Factors ◽  
Multi Scale ◽  
Cellular Environment

AbstractThe understanding of the multi-scale nature of molecular networks represents a major challenge. For example, regulation of a timely cell cycle must be coordinated with growth, during which changes in metabolism occur, and integrate information from the extracellular environment, e.g. signal transduction. Forkhead transcription factors are evolutionarily conserved among eukaryotes, and coordinate a timely cell cycle progression in budding yeast. Specifically, Fkh1 and Fkh2 are expressed during a lengthy window of the cell cycle, thus are potentially able to function as hubs in the multi-scale cellular environment that interlocks various biochemical networks. Here we report on a novel ChIP-exo dataset for Fkh1 and Fkh2 in both logarithmic and stationary phases, which is analyzed by novel and existing software tools. Our analysis confirms known Forkhead targets from available ChIP-chip studies and highlights novel ones involved in the cell cycle, metabolism and signal transduction. Target genes are analyzed with respect to their function, temporal expression during the cell cycle, correlation with Fkh1 and Fkh2 as well as signaling and metabolic pathways they occur in. Furthermore, differences in targets between Fkh1 and Fkh2 are presented. Our work highlights Forkhead transcription factors as hubs that integrate multi-scale networks to achieve proper timing of cell division in budding yeast.

scholarly journals The GLI-Code As a Potential Therapeutic Target in Myeloid Neoplasms

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 1655-1655
Author(s):  
Sayer Alharbi ◽  
Alek d Nielsen ◽  
Metis Hasipek ◽  
Bartlomiej P Przychodzen ◽  
James G Phillips ◽  
...  
Keyword(s):  
Cell Death ◽  
Transcription Factors ◽  
Tumor Cells ◽  
Cell Lines ◽  
Therapeutic Target ◽  
Target Genes ◽  
Myeloid Neoplasms ◽  
Gli1 Expression ◽  
Gli Transcription Factors ◽  
Hh Signaling

Abstract The proliferation of hematopoietic stem cells (HSCs) and progenitor cells (HPC) is tightly regulated during physiologic function of the hematopoietic system. Recent data suggest a role for activated developmental signaling pathways such as Hedgehog (Hh), Notch and Wnt in this process. The GLI genes (GLI1-4), known as GLI-code, encode Cys2-His2 zinc finger transcription factors that regulate the expression of target genes at the distal end of the canonical Hh pathway. This pathway is important during embryogenesis but mostly inactive in normal adult tissues. Activation of Hh effector SMO triggers an intracellular signaling cascade leading to activation of GLI2 and GLI3, which ultimately activate GLI1. It is well established that activation of GLI transcription factors may occur independent of canonical Hh signaling. Aberrant activation of the Hh/GLI pathway has been implicated in tumor progression and resistance to chemotherapy in MDS/AML. GLI transcription factors serve as a common node of activation through which many oncogenic signals, e.g., Hh, FLT3, PI3K-AKT, TGFβ and RAS converge. Moreover, overexpression of GLI transcription factors constitutes a negative prognostic survival marker. Higher expression of GLI transcription factors were recently reported in mutant FLT3-ITD AML cases. Extensive analysis of publicly available gene expression data sets revealed that GLI1 expression was significantly higher in complex AML karyotype (p<0.001, n=48) compared to other cytogenetic abnormalities t(15,17), inv(16), t(8,21), and t(11q23) (n=54, 47, 60 and 43 respectively). GLI1 expression was also significantly higher (p<0.05) in complex AML karyotype (n=87) compared to normal karyotype cases (n=989). The expression of GLI1 was even higher (p<0.001) in MDS (n=228) than normal AML karyotype. In addition, 8 out of 14 of NRAS mutant and 5 out of 7 KRAS mutant cases of AML in TCGA database have higher GLI1 expression. These biomarkers indicate advanced disease and constitute a major clinical problem with limited therapeutic options. Our observations strongly suggest that the GLI1 would be a bona-fide therapeutic target in MDS/AML. Thus specific and potent inhibitors of GLI1 can provide a rational approach for the treatments of these cases. While most efforts have been directed towards therapeutic targeting of upstream Hh signaling through PTCH/SMO, these measures remain ineffective in the presence of parallel signaling converging at GLI independent of canonical Hh pathway. Therefore, there is a need for the development of a GLI1-specific inhibitor that effectively blocks the nodal channel of Hh signaling. For this purpose, we used a structure guided approach to develop a highly potent and specific GLl1/GLI2 inhibitor iGli702, capable of preventing GLI1 DNA binding and thereby induction of cell death. Computational, biochemical and biophysical analysis demonstrated that iGli702 binds to the DNA binding surface of GLI1 protein between Zn-finger 2 and 3, effectively preventing Gli1-DNA interaction. Our data show that iGli702 is over 40 fold more effective than the reported GLI1/2 antagonist GANT61 in AML cell lines. iGli702 promotes apoptosis in a panel of leukemic cell lines, primary patient-derived AML samples and animal models. We observed an IC50 of 150 nM in cell viability assays. To test whether iGli702 induces cell death of progenitor tumor cells in semi-solid methylcellulose-based media, sorted CD34 positive tumor cells from AML cell lines (MOLM-13 and THP1) were treated with iGli702. This treatment significantly reduced colony formation of leukemia cells. However, iGli702 has only a minimal effect on normal PBMC and BM cells (LD50 >10µM). iGli702 efficacy was also compared in vitro to common chemotheraputic agents in AML. CytA and Ida were applied in parallel with iGli702to MOLM-13, THP-1, K562 AML cell lines. iGli702 showed superior and equivalent activity to CytA and Ida, respectively. In addition, we applied deep RNA NGS to analyze the cellular transcriptome after treatment with iGli702. Our data suggest that iGli702 directly inhibits Hedgehog pathway by reducing Hh target genes expression. In summary, iGli702 is highly effective as a single agent in killing hematopoietic malignant cells. This in turn demonstrates that targeting GLI1 Hh effector node is an excellent therapeutic approach in myeloid neoplasms. Disclosures No relevant conflicts of interest to declare.

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