Protein dynamics underlying cilium-dependent Hedgehog signaling

NIH Pandemic-Era Grants

Pandemic Era Grants

2024

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Principal Investigator: Radhika  Subramanian
Organization: MASSACHUSETTS GENERAL HOSPITAL
Fiscal Year: 2024
Award: $336,000
Funding agency: National Institute of General Medical Sciences

ABSTRACT
The Hedgehog (Hh) signaling pathway is a major intercellular signaling pathway important for embryonic
development and adult tissue homeostasis. Errors in Hh signaling are linked to several newborn birth defects
such as skeletal malformations and craniofacial defects, and associated with multiple tumors including basal
cell carcinoma and medulloblastoma. An important but poorly understood aspect of vertebrate Hh signaling is
the strict requirement of a microtubule-based organelle known as the primary cilium. While we now have a
wealth of data on the “parts-list” of proteins involved in the Hh signaling, the molecular mechanisms underlying
cilia-mediated signal transduction remains poorly understood. Our overall goal is to fill this major knowledge
gap and provide a biochemical framework for the Hh signaling pathway by reconstituting key reaction of this
pathway from its components. For this research, we will build on our experience in integrating single-molecule
imaging methods with biochemical assays and cell biological readouts to connect the biochemical properties of
the protein components to their cellular function. In this proposal, we focus a key step of the Hh signal
transduction pathway which is the establishment of signaling complexes at the base and the tip of the cilia, as
is needed for the proper activation or repression of the transcription factor Gli, the major effector of the Hh
pathway. Here, we will: (1) define these protein-protein interactions through a series of reconstitution studies
and determine how they restrict the Gli binding to the nuclear import machinery and (2) analyze the dynamics
of key pathway proteins in the cilium using high-resolution real-time imaging. Together, these studies will
define how Gli is regulated through dynamic transit between protein complexes at defined cytoplasmic
locations. We expect that our findings will not only advance our understanding of the basic biology of this
important signal transduction pathway but also shed light on how mutations in pathway components contribute
to developmental disorders and human cancers.

Terms: <0-4 weeks old><21+ years old><Acceleration><Adenosine Cyclic Monophosphate-Dependent Protein Kinases><Adult><Adult Human><Assay><Autoregulation><Basal Cell Epithelioma><Basal Transcription Factor><Basal cell carcinoma><Basal transcription factor genes><Basiloma><Binding><Binding Proteins><Binding Sites><Bioassay><Biochemical><Biological><Biological Assay><Biology><Birth Defects><Body Tissues><Cancer Genes><Cancer-Promoting Gene><Cancers><Catalytic Core><Catalytic Domain><Catalytic Region><Catalytic Site><Catalytic Subunit><Cell Body><Cell Communication and Signaling><Cell Function><Cell Nucleus><Cell Physiology><Cell Process><Cell Signaling><Cell surface><Cells><Cellular Function><Cellular Physiology><Cellular Process><Cilia><Cilium Microtubule><Combining Site><Complex><Congenital Abnormality><Congenital Anatomical Abnormality><Congenital Defects><Congenital Deformity><Congenital Malformation><Craniofacial Abnormalities><Cyclic AMP-Dependent Protein Kinases><Cytoplasm><Data><Distal><Embryo Development><Embryogenesis><Embryonic Development><Erinaceidae><Gene Down-Regulation><Gene Transcription><General Transcription Factor Gene><General Transcription Factors><Genetic Alteration><Genetic Change><Genetic Transcription><Genetic defect><Genetics-Mutagenesis><Glial Cell Tumors><Glial Neoplasm><Glial Tumor><Glioma><Goals><Growing End of the Microtubule><Hedgehog (Hh) signal transduction pathway><Hedgehog signaling complex><Hedgehog signaling complex location><Hedgehogs><Homeostasis><Human><Image><Impairment><Importins><In Vitro><Intracellular Communication and Signaling><Kinases><Kinesin><Knowledge><Ligand Binding Protein><Ligand Binding Protein Gene><Link><Location><Malignant Neoplasms><Malignant Tumor><Mediating><Medulloblastoma><Micro-tubule><Microtubules><Modern Man><Molecular><Molecular Interaction><Mutagenesis><Mutagenesis Molecular Biology><Mutation><NLS Peptide><Neuroglial Neoplasm><Neuroglial Tumor><Newborn Infant><Newborns><Nuclear><Nuclear Import><Nuclear Localization Signal><Nuclear Localization Signal Peptide><Nuclear Proteins><Nuclear Translocation><Nucleus><Oncogene Products><Oncogene Proteins><Oncogenes><Oncoproteins><Organelles><Output><PKA><Pathway interactions><Phosphotransferase Gene><Phosphotransferases><Physiological Homeostasis><Plus End of the Microtubule><Position><Positioning Attribute><Process><Property><Protein Binding><Protein Dynamics><Protein Kinase A><Proteins><Protocol><Protocols documentation><RNA Expression><Reaction><Reactive Site><Recombinant Proteins><Regulation><Research><Resolution><Rodent Ulcer><Series><Signal Pathway><Signal Transduction><Signal Transduction Pathway><Signal Transduction Systems><Signaling><Signaling Factor Proto-Oncogene><Signaling Pathway Gene><Signaling Protein><Site><Structure><Subcellular Process><TIRF Microscopy><Techniques><Therapeutic><Time><Tissues><Total Internal Reflection Fluorescent><Total Internal Reflection Fluorescent Microscopy><Transcription><Transcription Activator><Transcription Coactivator><Transcription Factor Coactivator><Transcription Factor Oncogene><Transcription Factor Proto-Oncogene><Transcription Repression><Transcription factor genes><Transcriptional Activator><Transcriptional Activator/Coactivator><Transcriptional Coactivator><Transcriptional Repression><Transforming Genes><Transphosphorylases><Visualization><Work><adulthood><base><bases><biologic><biological signal transduction><bound protein><cAMP-Dependent Protein Kinases><combat><craniofacial anomalies><craniofacial defects><craniofacial malformation><developmental disease><developmental disorder><experience><gene repression><genome mutation><glial-derived tumor><hedgehog signal transduction><hedgehog signaling><hedgehog signaling pathway><hh signal transduction><hh signaling pathway><image-based method><imaging><imaging method><imaging modality><insight><malformation><malignancy><neoplasm/cancer><neuroglia neoplasm><neuroglia tumor><newborn child><newborn children><pathway><prevent><preventing><programs><protein complex><protein protein interaction><real-time images><realtime image><reconstitute><reconstitution><recruit><resolutions><scaffold><scaffolding><single molecule><skeletal><smoothened signaling pathway><therapeutic agent development><therapeutic development><transcription co-activator><transcription factor><transcriptional co-activator><tumor>