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Principal Investigator: Clare Michal Waterman
Organization: NATIONAL HEART, LUNG, AND BLOOD INSTITUTE
Fiscal Year: 2024
Award: $545,678
Funding agency: National Heart Lung and Blood Institute
Project: CDK-1-mediated phase dissolution of Tensin1 condensates contributes to focal adhesion disassembly at mitosis to relieve an integrin inactivation G2-M checkpoint.
Personnel: Ana Pasapera, Hrishika Gogeni
Goal: Test the hypothesis that mitotic cell rounding is mediated by reducing cell-ECM adhesion through timely regulation of focal adhesion (FA) proteins by mitotic promoting factors.
We found that cell de-adhesion is an early prophase event that occurs perfectly coincidentally with mitotic cell rounding and cyclin B nuclear translocation, and is accompanied by a rapid drop in traction forces with no apparent pre-de-adhesion peak. Taken together, our data suggest that the mitotic-promoting factor CDK1 regulates cell de-adhesion at mitosis by mediating massive phosphorylation of the integrin-actin linker, tensin 1 in its IDR. This phosphorylation potentially drives the dissolution of tensin 1-containing biomolecular condensates and tensin 1 early removal from FA, leading to integrin inactivation and FA disassembly. Our data further suggest that cell de-adhesion at mitosis is a checkpoint required for cell cycle progression in adherent cells. We are right this very second making non-phosphorylatable and phosphomimetic tensin 1 variants with all the Ss and Ts in the IDR mutated to see if the phosphorylation is required for dissolution of the droplets, and if dissolution of droplets is required for FA disassembly at mitosis and for cell cycle progression.
Project: Role of ezrin in ECM stiffness sensing. Zbyszek Baster is working on the role of ezrin, radixin and moesin family (ERM) proteins in cell mechnosensation. ERMs are the main proteins linking the actin cortex to the cell plasma membrane118. ERM protein activity is regulated through their N-terminal binding to PIP2 rich plasma membranes followed by their phosphorylation at T567/T564/T558 (ezrin/radixin/moesin, respectively) by the homologous kinases, LOK or SLK, or by the Rho/ROCK pathway, to promote binding of the ERM C-terminal domains to actin119121. Activation of ERMs thus drives cortex-membrane tethering, which serves as one means of increasing cell stiffness and surface tension122. Since it is well established that mechanosensing of stiff ECM causes cell cortical stiffening89,122,123, it seems sort of shocking that little is known about whether ERM proteins are regulated in response to different mechanical stimuli. Zbyszek will thus test the hypothesis that cells respond to mechanical parameters of their environment in an ERM-dependent manner, and will dissect the signaling pathway mediating that response. He will use polyacrylamide or silicone cell culture substrates of a defined Young modulus and the Strex cell stretcher we plan to buy. ERM expression, activation and distribution will be analyzed in fibroblasts using molecular biology- and light microscopy-based techniques. ERM proteins and known ERM regulation or mechanotransduction pathways will be inhibited or stimulated using drug and gene expression manipulations to find molecular mechanisms by which ERM mechanosensation and mechanoresponse are mediated. Cell mechanical properties will be measured by AFM. This study will elucidate the mechanism of stiffness-dependent control of cell mechanical properties.
Terms: <3-D><3-Dimensional><3D><Abscission><Actin Filaments><Actins><Adhesion Plaques><Adhesions><Adhesives><Adopted><Binding><Biochemical><Blood Plasma Cell><C-terminal><CDC2><CDC2 Protein Kinase><CDC2 gene><CDK1><Cadherins><Cancers><Cell Adhesion><Cell Body><Cell Culture Techniques><Cell Cycle Controller CDC2 Gene><Cell Cycle Controller cdc2><Cell Cycle Progression><Cell Division Control Protein 2 Homolog><Cell Division Cycle 2><Cell Division Cycle 2 Protein><Cell Locomotion><Cell Migration><Cell Movement><Cell Nucleus><Cell Shape><Cell membrane><Cell-Extracellular Matrix><Cell-Matrix Adherens Junctions><Cells><Cellular Adhesion><Cellular Matrix><Cellular Mechanotransduction><Cellular Migration><Cellular Motility><Cessation of life><Collaborations><Cyclin B><Cyclin-Dependent Kinase 1><Cytoplasmic Membrane><Cytoskeletal System><Cytoskeleton><DNA Molecular Biology><Data><Death><Decision Making><Development><Dissociation><Drops><Drug usage><ECM><Embryo Development><Embryogenesis><Embryonic Development><Environment><Event><Excision><Extirpation><Extracellular Matrix><Family><Fibroblasts><Focal Adhesions><Focal Contacts><Friction><Gene Expression><Generations><Goals><Human Resources><Immune Diseases><Immune Disorders><Immune Dysfunction><Immune System Diseases><Immune System Disorder><Immune System Dysfunction><Immune System and Related Disorders><Immune response><Immunodeficiency and Immunosuppression Disorders><Immunologic Diseases><Immunological Diseases><Immunological Dysfunction><Immunological System Dysfunction><Immunological response><Integrins><Integrins Extracellular Matrix><Kinases><M Phase><Malignant Neoplasms><Malignant Tumor><Manpower><Measurement><Measures><Mechanical Signal Transduction><Mechanics><Mechanosensory Transduction><Mediating><Membrane><Micro-tubule><Microfilaments><Microtubules><Mitosis><Mitosis Stage><Mitotic><Modulus><Molecular><Molecular Biology><Molecular Interaction><Morphogenesis><Morphology><Mutate><Myofilaments><N-terminal><NH2-terminal><Nervous System><Neurologic Body System><Neurologic Organ System><Nuclear><Nuclear Translocation><Nucleus><Organelles><Output><PIP2><Pathway interactions><Phagocytosis><Phase><Phosphatidylinositol 4,5-Biphosphate><Phosphatidylinositol 4,5-Diphosphate><Phosphatidylinositol-4,5-Bisphosphate><Phosphorylation><Phosphotransferase Gene><Phosphotransferases><Physical Phenomena or Properties><Physical condensation><Physiologic><Physiological><Plasma Cells><Plasma Membrane><Plasmacytes><Polymers><Process><Proliferating><Prophase><Protein Family><Protein Phosphorylation><Proteins><PtIns 4,5-P2><PtdInsP2><Regulation><Removal><Research><Resolution><Role><Signal Pathway><Silicones><Site><Sorting><Stimulus><Surface><Surgical Removal><Techniques><Testing><Time><Traction><Translating><Transmission><Transphosphorylases><Variant><Variation><Vascular Diseases><Vascular Disorder><Vascular System><Wound Repair><biophysical approaches><biophysical methodology><biophysical methods><biophysical techniques><blood vessel disorder><cartilage link protein><cdc13 Protein><cdc2 gene product><cdc2+ Protein><cdk1 Kinase><cell behavior><cell cortex><cell culture><cell cultures><cell motility><cellular behavior><condensation><developmental><drug use><ezrin><host response><immune system response><immunoresponse><innovative technologies><insight><intracellular skeleton><light microscopy><link protein><malignancy><mechanic><mechanical><mechanical properties><mechanical stimulus><mechanosensing><mechanotransduction><membrane structure><membrane-organizing extension spike protein><migration><mimetics><moesin><morphogenetic process><nano meter scale><nano meter sized><nanometer scale><nanometer sized><nanoscale><neoplasm/cancer><p34 Protein Kinase><p34 Protein Kinase Gene><p34(CDC2) Gene><p34CDC2><p56cdc13><pathway><personnel><phosphoprotein p81><physical process><plasmalemma><plasmocyte><polarized cell><polyacrylamide><polymer><polymeric><programs><protein protein interaction><radixin><radixin protein><resection><resolutions><response><rho><self organization><social role><temporal measurement><temporal resolution><tensin><tensin1><three dimensional><time measurement><transmission process><vascular dysfunction><vasculopathy><wound healing><wound recovery><wound resolution>