Browsing by Subject "Integrins"

Now showing 1 - 4 of 4
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    Cellular adhesion dynamics: investigation of molecular clutch attachment and force transmission.
    (2008-12) Chan, Clarence Elvin
    As the major structural element of the cell, the cytoskeleton plays a vital role in response and transmission of forces in both extracellular and intracellular environments. For instance, in cell motility, the cell utilizes a host of proteins to physically link F-actin to the extracellular substrate, allowing the cell to exert traction forces as well as probe the mechanics of its local environment. During mitosis, the cell constructs a mitotic spindle, using microtubules and kinetochores to exert forces that segregate sister chromatids. Ultimately, understanding how cells build these robust molecular machines for unique tasks could one day lead to therapeutics that treat disease causing dysfunctions in these vital cellular processes. In order to explore how molecular clutches work in concert with the cytoskeleton to exert forces and maintain attachment under load, we developed a mechano-chemical cellular adhesion dynamics framework to simulate these processes. In the case of cellular motility, we find that a "motor-clutch" mechanism exhibits substrate-stiffness sensitive dynamics. On soft substrates, motor-clutch motility exhibits "load-and-fail" dynamics that lead to higher rates of retrograde flow and lower traction force transmission compared to stiff substrates. We confirm these predictions experimentally using embryonic chick forebrain neurons (ECFNs) plated on compliant polyacrylamide gels (PAGs) demonstrating that a motor clutch system could be the basis of cellular mechanosensing. We also use cellular adhesion dynamics to explore kinetochore-microtubule attachment during mitosis to identify what properties might be important in maintaining attachment during mitosis. We show that molecular clutch microtubule-lattice diffusion is important for relieving clutch stresses, prolonging bond life-times and minimizing detachment forces. Furthermore, molecular clutches that preferentially associate with interdimer interfaces, rather than with intradimer interfaces, promote robust kinetochore attachment by preventing the more distal, attachment-promoting linkers from becoming nonproductive. These findings help further our understanding of the mechanochemical basis of kinetochore attachment and mitosis, a process essential throughout development.
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    Crosstalk Between Adhesion Molecules Influences Cell Traction and Migration
    (2023-07) Kelly, Marcus
    Cell migration is the major driver of invasion and metastasis during cancer progression. For cells to migrate, they utilize the actin-myosin cytoskeleton and adhesion molecules, such as integrins and CD44, to generate traction forces in their environment. Whereas CD44 primarily binds to hyaluronic acid (HA), integrins primarily bind to extracellular matrix proteins (ECM) such as collagen. However, the role of CD44 under integrin-mediated conditions, and vice versa, is not well known. Here we used TFM to assess the functional mechanical relationship between integrins and CD44. Performing TFM on integrin-mediated adhesion conditions, i.e., collagen, we found that CD44KO U251 cells exerted more traction force than wild-type (WT) U251 cells. When using untreated WT and CD44-blocked WT, we observed comparable results with CD44KO cells again showing an increase in traction force on collagen gels. Conversely, in CD44-mediated adhesive conditions, integrin-blocked WT cells exerted higher traction force than untreated WT cells. Our data suggests that CD44 and integrins have a mutually antagonistic relationship where one receptor represses the other’s ability to generate traction force on its cognate substrate.
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    Effect of mutant Endostatin and Kringle 5 fusion protein on Tumor angiogenesis.
    (2016-05) Venkatachalam, Annapoorna
    Angiogenesis is important for the growth and metastasis of tumors. Endostatin is an endogenous inhibitor of angiogenesis and is shown to inhibit bFGF and VEGF induced signaling in endothelial cells. Recent evidence also indicates that it binds to αν and α5 group of integrins expressed endothelial cells and inhibits downstream signaling. Kringle 5, a potent anti-angiogenic molecule causes apoptosis of endothelial cells by associating with glucose-regulated protein 78 (GRP78). Previous studies from our laboratory have shown that a mutant (P125A) form of Endostatin (derived from collagen type XVIII) and Kringle 5 fragment of plasminogen can inhibit angiogenesis by inducing autophagy. P125A-endostatin and Kringle 5 interfere with distinct signaling pathways in endothelial cells. We hypothesized that a chimeric protein made of P125A-endostatin and Kringle 5 will have better anti-angiogenic activities. To test this hypothesis, we constructed a fusion protein consisting of P125A-endostatin and Kringle 5 (E-K5). Recombinant fusion protein was expressed in yeast and purified. E-K5 was found to inhibit proliferating endothelial cells and effectively blocked tumor induced angiogenesis. Anti-proliferative activity of E-K5 was linked to VEGF receptor and ανβ3 integrin-mediated signaling pathways. These studies establish the therapeutic potential of E-K5 as a potent anti-angiogenic molecule.
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    Involvement of Histone Deacetylase 4 (HDAC4) in Osteoclast Function
    (2019-08) Faulkner, Bora
    Bone modeling and remodeling during development and bone integrity throughout life are normally regulated via complex coupled actions of bone-resorbing osteoclasts and bone-forming osteoblasts. Loss of coupling between bone formation and bone resorption leads to pathogenesis of bone metabolic diseases including osteoporosis or osteopetrosis. Defects in osteoclast activity, whether increased activity or deficiency is responsible for bone destruction in many bone diseases such as osteoporosis, osteopetrosis and rheumatoid arthritis rather than impaired osteoblastic bone formation. Despite our growing knowledge in the mechanisms involved in the regulation of osteoclast differentiation and function, there is still a lot unknown. For this reason, it is important to understand the molecular mechanisms underlying how the activity of these bone-resorbing cells are regulated in order to develop effective therapies for bone disorders. Histone deacetylation is one such potential mechanism. It has been reported that class IIa histone deacetylase (HDAC), which include HDAC4, 5, 7 and 9 are regulators of osteoclastogenesis. Evidence from our lab and other labs using in vitro cell culture and in vivo mouse model systems indicated that HDAC7 and 9 are negative regulators of osteoclast differentiation and activity. However, whether the other class IIa members are functionally significant in osteoclasts is largely unknown. The aim of this research was to use in vitro osteoclast cell culture assays in conjunction with an in vivo mouse model to investigate the role(s) of HDAC4 in osteoclasts. HDAC4 conditional knockout (4cKO) mice exhibited increased bone mass phenotype (osteopetrosis) cause by decreased bone-resorbing activity of osteoclasts. HDAC4-deficient osteoclasts show reduced resorptive activity resulting from impaired signaling downstream of the M-CSF and v3 integrin and diminished M-CSF mediated adhesion and migration. Moreover, I demonstrated that c-Src activation in osteoclasts is regulated by HDAC4. The results of this thesis have identified HDAC4 as an essential regulator of osteoclast bone resorption activity both in vivo and in vitro.