Our long-term goal is to understand the structural and functional roles of capping protein (CapZ) in regulating the actin cytoskeleton and thereby cell shape and movement. A. Cloning and Analysis of CapZ Isoforms. We will determine whether CapZ exists as structurally and functionally distinct sarcomeric and non- sarcomeric isoforms and characterize the properties of those isoforms. cDNA's will be cloned and analyzed. The subcellular localization of the isoforms and their interaction with actin and actin-related protein in vitro will be studied. B. Structure-Function Analysis of CapZ. We identified an actin-binding site in the C-terminal region of the beta subunit of CapZ. To define the site more precisely, the actin-binding properties of point mutants in the region and of peptides from the region will be tested. Since the C- terminal region on its own binds actin monomers but does not cap filament, we will characterize the interaction of CapZ with actin monomers. Phosphoinositides bind to and inhibit CapZ, the only known regulatory mechanism for CapZ. We will test whether the C-terminal region of the beta subunit also includes the phosphoinositide-binding site. C. Analysis of CapZ and Actin-related Protein in the Dynactin Complex. Dynactin complex contains a short actin-like filament and both actin- related protein and the non-sarcomeric form of CapZ. We will investigate the structural and functional properties of the complex, especially how the properties of the actin-like filament compare to those of actin and how the assembly of the actin-like filament is regulated. D. Analysis of In Vivo Function of Non-sarcomeric CapZ. Our long-term goal is to investigate the in vivo function of non-sarcomeric CapZ by altering its activity in cells. We will study the localization and biochemical properties of non-sarcomeric CapZ, in order to generate specific testable hypotheses for future experiments and to design specific inhibitors for such experiments. We will analyze its role in cell junctions, microvilli, and fibroblasts.
| Kim, Joanna; Cooper, John A (2018) Septins regulate junctional integrity of endothelial monolayers. Mol Biol Cell 29:1693-1703 |
| Yen, Matthew; Qi, Zongtai; Chen, Xuhua et al. (2018) Transposase mapping identifies the genomic targets of BAP1 in uveal melanoma. BMC Med Genomics 11:97 |
| Santiago-Tirado, Felipe H; Onken, Michael D; Cooper, John A et al. (2017) Trojan Horse Transit Contributes to Blood-Brain Barrier Crossing of a Eukaryotic Pathogen. MBio 8: |
| Zhou, Julie Y; Szasz, Taylor P; Stewart-Hutchinson, Phillip J et al. (2016) L-Plastin promotes podosome longevity and supports macrophage motility. Mol Immunol 78:79-88 |
| Wang, Xinxin; Galletta, Brian J; Cooper, John A et al. (2016) Actin-Regulator Feedback Interactions during Endocytosis. Biophys J 110:1430-43 |
| Xu, Xiaolu; Wang, Xinxin; Todd, Elizabeth M et al. (2016) Mst1 Kinase Regulates the Actin-Bundling Protein L-Plastin To Promote T Cell Migration. J Immunol 197:1683-91 |
| Mooren, Olivia L; Kim, Joanna; Li, Jinmei et al. (2015) Role of N-WASP in Endothelial Monolayer Formation and Integrity. J Biol Chem 290:18796-805 |
| Mukherjee, Suranjana; Kim, Joanna; Mooren, Olivia L et al. (2015) Role of cortactin homolog HS1 in transendothelial migration of natural killer cells. PLoS One 10:e0118153 |
| Sherman, Marc S; Lorenz, Kim; Lanier, M Hunter et al. (2015) Cell-to-cell variability in the propensity to transcribe explains correlated fluctuations in gene expression. Cell Syst 1:315-325 |
| Onken, Michael D; Mooren, Olivia L; Mukherjee, Suranjana et al. (2014) Endothelial monolayers and transendothelial migration depend on mechanical properties of the substrate. Cytoskeleton (Hoboken) 71:695-706 |
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