Research Interests for Daniel P. Kiehart

Our intellectual focus is on identifying determinants of cell shape that function during development and wound healing. We utilize novel biophysical strategies (in collaboration with Glenn Edwards' group in Physics and with Stephanos Venakide's and John Harer's groups in Mathematics) in concert with modern molecular genetic and reverse genetic approaches in Drosophila to explore the forces that are responsible for cell shape change and movements. We show that both the amnioserosa and a "supracellular purse string" in the leading edge of the lateral epidermis contribute to the movements of dorsal closure. Dorsal closure proceeds even if we ablate one (but NOT both!)of the tissues responsible for closure, indicating that this model cell (epithelial) sheet movement depends on redundant forces that in concert drive morphogenesis. We show that the magnitude of each force is significantly larger than their vector sum indicating that there is both potential for generating large forces and that successful morphogenesis requires that the forces applied be precisely balanced. We have also explored the molecules responsible for generating those movements. We showed that conventional nonmuscle myosin (myosin II) provides key contractile forces in different tissues where the supramolecular complexes that incorporate this motor protein are distinct. How molecular events are regulated such that large, opposing forces efficiently drive morphogenesis remains a mystery, but we are pursuing leads that point to two distinct pathways: the bidirectionally signaling integrin cell surface receptors and mechanically gated channels.

We are also pursuing the morphogenesis of actin-cytoskeleton based projections that are a key feature of a variety of cells, including those that are specialized for sensory reception in human vision and hearing. We have again turned to Drosophila as a model system where we study the morphogenesis of epidermal hairs and sensory bristles. Our work centers on an unconventional myosin (myosin VIIA) encoded by crinkled a gene that is required for the formation of epidermal hairs and bristles. We show that myosin VIIA is required for the coallescence of actin pre-hairs into the robust actin bundles that form the skeleton on which hairs and bristles can be built. In collaboration with Dan Eberl's lab (University of Iowa) we showed that myosin VIIA is also essential for fly hearing -- remarkably, its human homolog is also required for human hearing, even though the mechanisms of auditory sensory reception in these phylogenetically diverged systems are very different. We have begun to characterize myosin VIIA structurally using NMR of purified protein domains. With Jim Seller's lab at the NIH we have used fast time course kinetics and single molecule assays to analyze molecular function and show that this myosin VIIA is a processive motor. We are beginning to characterize the proteins that collaborate with both myosin II and myosin VIIA using biochemical strategies in vitro, yeast two hybrid approaches in vivo and genetic interaction strategies in fly.

Together, our experiments promise to reveal the nature of cytoskeletal function in cell shape determination for cell division and morphogenesis throughout development and organismal homeostasis.

Keywords:

3T3 Cells, Acanthamoeba, Acrosome, actin, Actin Cytoskeleton, Actin Depolymerizing Factors, Actin-Related Protein 2, Actin-Related Protein 3, Actinin, Actins, Actomyosin, Adenosine Triphosphatases, Adenosine Triphosphate, Adhesion, Alleles, Alternative Splicing, Amino Acid Motifs, Amino Acid Sequence, Amoeba, Anaphase, Animal Structures, Animals, Animals, Genetically Modified, Antibodies, Antibodies, Monoclonal, Antibody Specificity, Antigen-Antibody Complex, Apoptosis, Auditory Perception, Avian Proteins, Base Sequence, beta-Galactosidase, Binding Sites, Biomechanics, Biophysics, Birefringence, Blastoderm, Blastomeres, Blood Platelet Disorders, Blotting, Northern, Blotting, Southern, Body Patterning, Brain, Buffers, Ca(2+) Mg(2+)-ATPase, Cadherins, Caenorhabditis elegans, Caenorhabditis elegans Proteins, Caffeine, Calcium, Calcium-Transporting ATPases, Calmodulin, Carrier Proteins, Catalytic Domain, cdc42 GTP-Binding Protein, cdc42 GTP-Binding Protein, Saccharomyces cerevisiae, Cell Adhesion, Cell Adhesion Molecules, Cell adhesion--Molecular aspects, Cell Communication, Cell Compartmentation, Cell Cycle Proteins, Cell Differentiation, Cell Division, Cell Line, Cell Membrane, Cell Membrane Permeability, cell motility, Cell Movement, Cell Nucleus, Cell Polarity, Cell Proliferation, Cell Shape, Cell Size, Cells, Cultured, Chelating Agents, CHO Cells, Chromosome Aberrations, Chromosome Mapping, Chromosomes, Circular Dichroism, Cleavage Stage, Ovum, Cloning, Molecular, Colchicine, Cold Temperature, Computer Simulation, Conserved Sequence, Contractile Proteins, Cricetinae, Crosses, Genetic, Cytochalasins, Cytology, Cytoplasm, Cytoskeletal Proteins, cytoskeleton, Cytoskeleton, Cytosol, Deafness, Demecolcine, Dictyostelium, Dimerization, Disease Models, Animal, DNA, DNA Glycosylases, DNA Mutational Analysis, DNA Restriction Enzymes, DNA, Complementary, DNA-Binding Proteins, dorsal closure, Drosophila, Drosophila melanogaster, Drosophila Proteins, Drosophila--Genetics, Drug Stability, Dyneins, Dystrophin, Ear, Inner, Echinodermata, Elasticity, Electrophoresis, Polyacrylamide Gel, Electrophysiology, Embryo, Nonmammalian, Embryology, Embryonic Development, Enhancer Elements, Genetic, Enzyme Activation, Enzyme Inhibitors, Enzyme-Linked Immunosorbent Assay, Epidermis, Epithelial Cells, Epithelium, Epitopes, Erythrocytes, Evaluation Studies as Topic, Evoked Potentials, Auditory, Evolution, Molecular, Exons, Extracellular Matrix, Extremities, Eye, Eye Abnormalities, Female, Fertilization, Fibroblasts, Fibronectins, Fluorescence, Fluorescence Recovery After Photobleaching, Fluorescent Antibody Technique, Fungal Proteins, gamma-Globulins, Gastrula, Gastrulation, Gene Dosage, Gene Expression, Gene Expression Regulation, Gene Expression Regulation, Developmental, Gene Library, Genes, Genes, Dominant, Genes, Homeobox, Genes, Insect, Genes, Lethal, Genetic Complementation Test, Genetic Vectors, Genetics, Green Fluorescent Proteins, GTP Phosphohydrolases, GTP-Binding Proteins, Guanine Nucleotide Exchange Factors, Guanosine Triphosphate, Hair Cells, Auditory, hearing, Heat-Shock Response, Helminth Proteins, Heterotrimeric GTP-Binding Proteins, Hippocampus, Homeodomain Proteins, HSP70 Heat-Shock Proteins, Humans, Hydrogen, Hydrogen-Ion Concentration, Hypertonic Solutions, Hypotonic Solutions, Image processing, Image Processing, Computer-Assisted, Immunoblotting, Immunoglobulin G, Immunoglobulin M, Immunoglobulins, Immunohistochemistry, Immunoprecipitation, In Situ Hybridization, Indicators and Reagents, Infertility, Insect Hormones, Insect Proteins, Insects, Integrin alpha Chains, Integrins, Intercellular Junctions, Ion Channels, Ionophores, Isoelectric Focusing, Isoenzymes, JNK Mitogen-Activated Protein Kinases, Kinetics, Larva, Lasers, Luminescent Measurements, Luminescent Proteins, Macromolecular Substances, Magnesium Chloride, Male, Mannitol, Mass Spectrometry, Mathematics, Mechanical Phenomena, Mechanotransduction, Cellular, Meiosis, Membrane Potentials, Membrane Proteins, Metabolism, Methods, Mice, Microfilament Proteins, Microinjections, Microscopy, Microscopy, Confocal, Microscopy, Electron, Microscopy, Electron, Scanning, Microscopy, Electron, Transmission, Microscopy, Fluorescence, Microscopy, Immunoelectron, Microscopy, Phase-Contrast, Microscopy, Video, Microsurgery, Microtubules, Microvilli, Mitogen-Activated Protein Kinases, Mitosis, Models, Biological, Models, Genetic, Models, Molecular, Models, Structural, Molecular Biology, Molecular Motor Proteins, Molecular Sequence Data, Molecular Weight, morphogenesis, Morphogenesis, Morphogenesis--Molecular aspects, motor protein, Multigene Family, Muscle Proteins, Muscle, Skeletal, Muscles, Mutagenesis, Mutagenesis, Insertional, Mutagenesis, Site-Directed, Mutation, Mutation, Missense, myosin, Myosin, Myosin antibodies, Myosin Heavy Chains, Myosin Light Chains, Myosin Subfragments, Myosin Type II, Myosin Type V, Myosin-Light-Chain Kinase, Myosin-Light-Chain Phosphatase, Myosins, NAD, NADP, Nerve Tissue Proteins, Nervous System, Neurons, Nonmuscle Myosin Type IIA, Nucleic Acid Hybridization, Nucleoproteins, Oocytes, Oogenesis, Osmolar Concentration, Ovarian Follicle, Ovary, Ovum, Oxalates, Oxidation-Reduction, Peptide Fragments, Peptides, Phenotype, Phosphorylation, Photoreceptor Cells, Invertebrate, Phylogeny, Physical Chromosome Mapping, Physiology, Poly A, Polymerase Chain Reaction, Polymerization, Polymers, Profilins, Promoter Regions, Genetic, Protein Binding, Protein Biosynthesis, Protein Conformation, Protein Kinase C, Protein Kinases, Protein Precursors, Protein Structure, Secondary, Protein Structure, Tertiary, Protein-Serine-Threonine Kinases, Proteins, Pseudopodia, Pupa, Rabbits, Rats, Recombinant Fusion Proteins, Recombinant Proteins, Regulatory Sequences, Nucleic Acid, Repetitive Sequences, Amino Acid, Repetitive Sequences, Nucleic Acid, Repressor Proteins, Restriction Mapping, Retinal Rod Photoreceptor Cells, rhoA GTP-Binding Protein, rhoB GTP-Binding Protein, RNA Splicing, RNA, Messenger, Saccharomyces cerevisiae Proteins, Salts, Sarcomeres, Schizosaccharomyces, Schizosaccharomyces pombe Proteins, Scyphozoa, Sea Urchins, sensory reception, Sensory Receptor Cells, Sequence Alignment, Sequence Analysis, DNA, Sequence Analysis, Protein, Sequence Homology, Amino Acid, Sequence Homology, Nucleic Acid, Serine, Serine Endopeptidases, Serous Membrane, Signal Transduction, Sodium Channels, Sodium Chloride, Species Specificity, Spectrin, Spectrometry, Fluorescence, Spermatozoa, Spider Venoms, Staining and Labeling, Starfish, Stress, Mechanical, Swine, Temperature, Time Factors, Tissue Distribution, Transcription Factors, Transcription, Genetic, Transfection, Transforming Growth Factor beta, Transgenes, Tropomyosin, TRPC Cation Channels, Trypsin, Ultraviolet Rays, Up-Regulation, Vertebrates, Wing, wound healing, Wound Healing, Wounds and Injuries, X Chromosome, Xenopus, Zygote

Current projects:

Cytoskeleton and motor protein function in morphogenesis and wound healing

Light activated gene expression

Filopodia function in morphogenesis and wound healing

Protein complex function in hearing

Areas of Interest:

morphogenesis
wound healing
motor protein structure and function
cytoskeleton
molecular structure
phylogeny of gene families

Recent Publications   (search)

1. Allen, RL; George, AN; Miranda, E; Phillips, TM; Crawford, JM; Kiehart, DP; McClay, DR, Wound repair in sea urchin larvae involves pigment cells and blastocoelar cells., Developmental biology, vol. 491 (November, 2022), pp. 56-65 [doi] [abs]
2. Haertter, D; Wang, X; Fogerson, SM; Ramkumar, N; Crawford, JM; Poss, KD; Di Talia, S; Kiehart, DP; Schmidt, CF, DeepProjection: specific and robust projection of curved 2D tissue sheets from 3D microscopy using deep learning., Development, vol. 149 no. 21 (November, 2022) [doi] [abs]
3. Moore, RP; Fogerson, SM; Tulu, US; Yu, JW; Cox, AH; Sican, MA; Li, D; Legant, WR; Weigel, AV; Crawford, JM; Betzig, E; Kiehart, DP, Superresolution microscopy reveals actomyosin dynamics in medioapical arrays., Molecular biology of the cell, vol. 33 no. 11 (September, 2022), pp. ar94 [doi] [abs]
4. Sallee, JL; Crawford, JM; Singh, V; Kiehart, DP, Mutations in Drosophila crinkled/Myosin VIIA disrupt denticle morphogenesis., Developmental biology, vol. 470 (February, 2021), pp. 121-135 [doi] [abs]
5. Fogerson, SM; Mortensen, RD; Moore, RP; Chiou, HY; Prabhu, NK; Wei, AH; Tsai, D; Jadi, O; Andoh-Baidoo, K; Crawford, J; Mudziviri, M; Kiehart, DP, Identifying Key Genetic Regions for Cell Sheet Morphogenesis on Chromosome 2L Using a Drosophila Deficiency Screen in Dorsal Closure., G3 (Bethesda, Md.), vol. 10 no. 11 (November, 2020), pp. 4249-4269 [doi] [abs]