Browsing by Subject "Actin"
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Item Assessing the Roles of Striatin Orthologs in Fungal Morphogenesis, Sexual Development and Pathogenicity(2012-10-19) Wang, Chih-LiStriatin family proteins contain a caveolin binding domain, a coiled-coil motif, a calmodulin binding domain, and a WD-repeat domain. Homologs of striatin protein have been However, our knowledge of the function and the molecular mechanism of fungal striatin homologs is limited. Based on the conserved sequences of functional domains, I hypothesized that the fungal striatin orthologs also act as scaffolding proteins that are functionally conserved among fungal species and involved in multiple types of development in the diverse kingdom Mycota. I used reverse genetic strategies to study the function of the Aspergillus nidulans striatin ortholog (strA) and the Colletotrichum graminicola striatin ortholog (str1). In assays of sexual development, the strA deletion strain (?strA) produces fewer ascospores with smaller cleistothecia, while the str1 deletion strain (?str1) is defective in perithecia development. The ?strA phenotypes indicate that StrA is associated with ascosporogenesis in cleistothecia. Both ?strA and ?str1 are reduced in radial growth and in conidia production. The ?str1 strain is also altered in its spiral growth pattern and morphology of conidia and hyphopodia, but it produces appressoria similar to wild type. The pairing of nitrate non-utilizing mutants demonstrates that Str1 is required for hyphal fusion. In pathogenicity, ?str1 is less virulent in maize anthracnose leaf blight and stalk rot. The phenotypes of ?str1 are complemented by the Fusarium verticillioides striatin ortholog (fsr1), indicating that Fsr1 and Str1 are functionally conserved. Over-expression of StrA reveals its positive role in conidiation and the sexual production. StrA::eGFP localizes mainly to the endoplasmic reticulum. After comparing the results from these two species and other studied fungal species, I suggest that fungal striatins are involved in five types of development including hyphal growth, hyphal fusion, conidiation, sexual development, and virulence, and propose a model of fungal striatin protein interactions to account for these diverse phenotypes.Item Development of methodology for freeze-substitution and immunolocalization in differentiating tracheary elements of Zinnia elegans(Texas Tech University, 1998-05) Muehring, Trina CelesteIn higher plants, redifferentiation occurs when one cell type transforms into another with an associated change in cell function. Xylem tissue is formed either in this manner, from parenchyma cells, or by de novo differentiation from procambium/cambium initials. Tracheary elements (TEs) are water-conducting cells in xylem tissue. Dead at maturity, these elements connect end to end to form a transport system In culture, parenchyma cells have been shown to redifferentiate into tracheary elements, forming secondary wall patterns by locahzed thickenings (Kohlenbach et al., 1981) (see Figs. 1.1 and 1.2). Zinnia elegans has become a model system by which to study TE differentiation because single cells can be induced to form TEs in culture. Present in only specialized cell types such as TEs, the secondary cell wall is laid down inside the primary ceU waU under cytoskeletal control. The control of secondary wall development is not well defined. Actin and microtubules are two cytoskeletal elements that theory and observations predict will be involved in this process. Research with antibodies in the Zinnia system can reveal whether or not any pattem is recognizeable before the secondary wall thickenings appear and the morphological stages can then be identified that represent the estabhshment of that pattem. "The possible role of cytoskeletal elements in regulating microfibril organization remains one of the central unresolved questions in plant cell biology... diflSculty arises in proving that the cytoskeleton is essential for the organized deposition of wall microfibrils" (Seagull 1989. p. 145). The focus of this research is to optimize uhra rapid freezing methods so that routine experiments performed on Zinnia TEs will resuh in state-of-the-art preservation of cytoplasm and immuno-gold labeUng by antibodies.Item Elastic network & finite element model to study actin protein mechanics & its molecular elasticity(2010-12) Marquez, Joel David; Moon, T. J. (Tess J.); Ren, PengyuWhile there have been many recently developed Elastic Network Models (ENM) to calculate the fluctuation dynamics of proteins, e.g., Gaussian Network Model (GNM), Anisotropic Network Model (ANM), Distance Network Model (DNM), the concept of loading these models to study the molecular mechanics and constitutive behavior of structural proteins has remained relatively untouched, until very recently. This work entails using the ANM as the framework for developing a finite element model of a 9–monomer strand of actin. Critical input parameters to the model, such as the cutoff radius, r[subscript c], and spring constant, k, are generated by matching the all-atom steered molecular dynamics (SMD) residue displacements to that of the ANM. The parameters yielding the best match between the SMD and structural ENM (SENM) simulations will then be input into the finite element model (FEM) for a more in depth analysis. The finite element model incorporates a 9–monomer strand of actin. The F–actin strand is subjected axial and torsional loads comparable to those seen in vivo. Key areas of interest in the protein are examined, such as the nucleotide binding pocket (NBP) and the DNase I binding loop, to demonstrate how loading affects the protein’s conformation. Local residue displacements are tracked in an effort to garner a better understanding of how various loads are transmitted through F–actin during key events. Insights and conclusions are discussed along with the implications of this work.Item Myosin II alters the viscoelasticity and self-assembly properties of actin networks(2002-05) Humphrey, David Harold; Käs, Joseph A.; Molineux, Ian