Inducible expression of Pisum sativum xyloglucan fucosyltransferase in the pea root cap meristem, and effects of antisense mRNA expression on root cap cell wall structural integrity

Mitosis and cell wall synthesis in the legume root cap meristem can be induced and synchronized by the nondestructive removal of border cells from the cap periphery. Newly synthesized cells can be examined microscopically as they differentiate progressively during cap development, and ultimately detach as a new population of border cells. This system was used to demonstrate that Pisum sativum L. fucosyl transferase (PsFut1) mRNA expression is strongly expressed in root meristematic tissues, and is induced >2-fold during a 5-h period when mitosis in the root cap meristem is increased. Expression of PsFut1 antisense mRNA in pea hairy roots under the control of the CaMV35S promoter, which exhibits meristem localized expression in pea root caps, resulted in a 50–60% reduction in meristem localized endogenous PsFut1 mRNA expression measured using whole mount in situ hybridization. Changes in gross levels of cell wall fucosylated xyloglucan were not detected, but altered surface localization patterns were detected using whole mount immunolocalization with CCRC-M1, an antibody that recognizes fucosylated xyloglucan. Emerging hairy roots expressing antisense PsFut1 mRNA appeared normal macroscopically but scanning electron microscopy of tissues with altered CCRC-M1 localization patterns revealed wrinkled, collapsed cell surfaces. As individual border cells separated from the cap periphery, cell death occurred in correlation with extrusion of cellular contents through breaks in the wall

(+)-Pisatin Biosynthesis: From (-) Enantiomeric Intermediates via an Achiral Isoflavene

Pterocarpan phytoalexins are antimicrobial compounds produced by legumes when challenged by biotic stresses. Most legumes produce pterocarpan phytoalexins with (-)-stereochemistry but pea (Pisum sativum L.) produces as its major phytoalexin (+)-pisatin. Pea also occasionally produces a minor amount of (-)-maackiain as a pterocarpan phytoalexin, and studies on the biosyntheses of (+)-pisatin and (-)-maackiain have shown that up to (-)-7,2'-dihydroxy-4',5'-methylenedioxyisoflavanone [(-)-sophorol] and 7,2'-dihydroxy-4',5'-methyl-enedioxyisoflavanol [(-)-DMDI]they have common intermediates with (-)-DMDI being where the two pathways diverge. The final step in (+)-pisatin biosynthesis is the methylation of (+)-6a-hydroxymaackiain [(+)-6a-HMK] by 6a-hydroxymaackiain methyltransferase (HMM2) but the steps from (-)-DMDI to (+)-6a-HMK are unknown.The shifting of the stereochemistry from (-)-DMDI to (+)-6a-HMK has been proposed to involve the achiral isoflavene, 7, 2'-dihydroxy-4', 5'-methylene-dioxyisoflav-3-ene (DMDIF). In this dissertation, I have shown that cis-(-)-DMDI is the enzymatic product of (-)-sophorol, and is the precursor of DMDIF which is produced by the dehydration activity of "isoflavene synthase" (IFVS). IFVS activity was not observed in elicited tissues of alfalfa, chickpea, beans, pepper, and broccoli, plants that do not produce (+) pterocarpans. Partial purification of IFVS demonstrated that it is either large in size or tightly complexed with other proteins. The SDS-PAGE of the 29-fold purified product revealed 12 major bands that aggregated into 3 bands in the non-denaturing PAGE. IFVS activity was in band 3 which co-migrated with marker proteins of>100 kDa in size. Proteins identified from LC-MS/MS peptide sequences of the proteins in band 3, when compared to three protein databases, did not identify any proteins with an enzymatic activity expected for IFVS. A disease resistance-response protein (a dirigent-like protein) and two protein-binding proteins were the most abundantly detected proteins in the pea transcriptome-translated database. Also, four of the known enzymes (isoflavone reductase, HMM1, HMM2, and sophorol reductase) involved in (+)-pisatin biosynthesis were among the proteins identified. It may be that IFVS is associated with these other proteins as a complex in vitro and in vivo. The lack of detection of IFVS in the databases could be because it has not yet been sequenced as it functions in a rare biosynthetic pathway

Thermal management considerations for warm forming dies.

One approach taken by automotive manufacturers toward improving fuel economy is to reduce the overall mass of vehicles by replacing steel components with aluminum. However, the room temperature formability of aluminum is less than that of conventional drawing quality steel. One way in which researchers have sought to improve the formability of aluminum is by warm forming. Warm forming refers to sheet forming in the temperature range of 200°C to 350°C using heated, matched die sets similar to conventional stamping. This thesis work details a new approach to the design of warm forming dies based on analytical methods and finite element analyses (FEA) for accurate control of the thermal distribution within the warm forming die by strategic use of heaters and control zones. The analytical tool was written to model and simplify each boundary condition of the die system. The FEA enables optimization of heater and thermocouple locations for temperature uniformity with minimal computation time. Heat generated by the blank deformation is not considered in this approach of thermal die design. The results of this work show that the complex boundary conditions of a warm forming die can be simplified into a one-dimensional heat transfer parameter using energy conservation for each local die surface. The effective heat transfer parameters were then utilized by FEA to predict the internal temperature response of a die and proven to be consistent with the actual die thermal behavior. The steady-state temperature assumptions of the thermal finite element analyses were verified through analytical and experimental methods. These warm forming simulation tools were then applied to the development and construction of a production die and validated. In summary, through this thesis work novel simulation design tools required for handling the rapid design of large, complex, non-symmetric warm forming dies were developed to design a thermally stable warm forming die suited for high volume production.D.Eng.Applied SciencesMechanical engineeringUniversity of Michigan, Horace H. Rackham School of Graduate Studies College of Engineering Graduate Professional Programshttp://deepblue.lib.umich.edu/bitstream/2027.42/127203/2/3510590.pd

Rationalizing violence : examining discourse and school closures in Washington, D.C., public schools

[ACCESS RESTRICTED TO THE UNIVERSITY OF MISSOURI AT AUTHOR'S REQUEST.] From 2008 to 2014, the District of Columbia Public Schools (DCPS), in Washington, D.C., underwent school consolidation and reorganization that led to 38 public school closures. These closures disproportionately impacted Black students and were reiterations of a historical discourse of violence against Black livelihood. While some researchers suggest students experience minimal, if any, adverse impacts from school closure and contend no harm is inflicted, I posit the disproportionality of the impacts alone warrants further examination. The problem created by school closure and consolidation policies is a nuanced issue in that it is an extension of inequities in public education but also a new form of violence yet to be articulated in educational research. Using Freeman's (1977) theory of legitimization and Bonilla-Silva's (1997) theory of racialized social systems to expand traditional notions of violence, this study examined and (re)produced public data from a 2013 and 2014 court case, Smith et al. v. Henderson et al., as a springboard to operationalize violent discourse as it pertains to school closure. Through the application of Critical Discourse Analysis, this study examined the discursive practices that legitimize and normalize violence. The study found evidence of practices such as referencing governing authorities, framing a narrative for the court and public, and circumventing moral and ethical evaluation of legitimized violence. Furthermore, these discursive practices championed normalized violence that articulated an exclusionary, spatialized, and institutional bias against residents living east of D.C.'s Rock Creek Park. Interdisciplinary approaches to theoretical and methodological frameworks, as well as top-down and bottom-up (Sabatier, 1986) approaches, are discussed as issues for future research and K-12 education policy reform and advocacy.Includes bibliographical reference

Rhodesia agricultural journal.

Mode of access: Internet

Rhodesia agricultural journal.

v.6:1-6 (Bd w/o 2-3)(1908-1909