Determination of AtSCD1 mRNA and protein levels in the temperature sensitive mutant scd1-1

Abstract only availablePlant cells defend themselves against bacterial infection by first detecting bacterial flagellin outside the cell and then initiating defense responses within the cell. Our lab is interested in identifying and characterizing vesicular trafficking components that play a role in defense responses to bacterial flagellin and its active peptide derivative, flg22, in Arabidopsis thaliana. Our current research focuses on SCD1, a protein implicated in polarized vesicle secretion. Recent results in our lab also indicate that scd1-1 plants, which contain a point mutation in the SCD1 gene, exhibit reduced flg22-responses, notably in the production of reactive oxygen species (ROS). To gain a better understanding of the role of SCD1 in flg22-responses, we analyzed SCD1 mRNA and protein levels in scd1-1 plants compared to the wild-type (WT), Colgl1, plants using qRT-PCR and protein blot analysis, respectively. SCD1 levels were compared in plants grown continuously at 22°C, the non-permissive temperature, to plants grown initially at 22°C and then shifted for 9 days to the permissive temperature, 17°C. Data indicate that at both 22°C and 17°C, SCD1 mRNA levels were statistically similar in scd1-1 compared to WT plants. Conversely, SCD1 protein levels were significantly higher in scd1-1 plants shifted to 17°C compared to those grown at 22°C. An increase in SCD1 protein level also correlated with an increase in flg22-induced ROS production at 17°C. To further confirm a requirement for SCD1 in flg22-repsonses, we showed that in an scd1-1 mutant plant transformed with the SCD1 gene (scd1-1/pscd1::SCD1), SCD1 protein levels were similar to those in WT grown at 22°C. These results are consistent with the complemented line exhibiting flg22-induced ROS production similar to WT levels. Given these results, both the temperature shift and the complementation experiments support our hypothesis that SCD1 protein is required for full flg22-induced responses.Life Sciences Undergraduate Research Opportunity Progra

Women without children : identity, choice, responsibility

Title from PDF of title page (University of Missouri--Columbia, viewed on May 26, 2010).The entire thesis text is included in the research.pdf file; the official abstract appears in the short.pdf file; a non-technical public abstract appears in the public.pdf file.Dissertation advisor: Dr. Barbara J. Bank.Vita.Ph. D. University of Missouri--Columbia 2010.Research about childless women tends to reinforce two widely-held assumptions. The first is that being childless is a central component informing non-mothers' identities. The second is that choice is the best concept for understanding how women experience having no children. Guided by the principles of social constructionism and utilizing the Twenty-Statements Test and semi-structured interviews, this research calls both of those assumptions into question. This research also proposes recruitment of naive subjects in identity research to reduce the likelihood of certain identities arising simply because they are the focus of said research. It also advocates expanding appeals higher loyalties as it is presented in accounts literature to include more than significant others. I found that women who are not mothers rarely think of themselves as childless unless prompted to do so. Rather than having situation-free childless identities, these identities are bound by context. Furthermore, some of my respondents had no childless identity to speak of because they were either never called upon to account for their childlessness or they ignored such attempts. Choice was utilized by nearly all of the women in my sample. Respondents who would clearly have been defined as involuntarily childless by other researchers presented themselves as women with reproductive choices who exercised control over their capacities to become mothers as well as control over how others perceive them.Includes bibliographical reference

Adapting an Analog Records Management System for the Ingest and Accession of Permanent Electronic Records

The Records and Archives Division of the Office of the Missouri Secretary of State (hereafter MSA) received two National Historical Publications and Records Commission (NHPRC) grants for the purpose of establishing an electronic records program at the Missouri State Archives. The first grant covered planning, staff training and a consultant who determined that minor modifications to the current State of Missouri Agency Records Tracking (SMART) System would allow for the ingest of permanent electronic records. The second grant funded the SMART upgrade and the purchase of data-grabbing equipment. The upgrade succeeded and MSA was able to ingest 150 GB of permanent electronic records via the SMART System

Resolving the cofactor-binding site in the proline biosynthetic enzyme human pyrroline-5-carboxylate reductase 1

Pyrroline-5-carboxylate reductase (PYCR) is the final enzyme in proline biosynthesis, catalyzing the NAD(P)H-dependent reduction of [?]1-pyrroline-5-carboxylate (P5C) to proline. Mutations in the PYCR1 gene alter mitochondrial function and cause the connective tissue disorder cutis laxa. Furthermore, PYCR1 is overexpressed in multiple cancers, and the PYCR1 knock-out suppresses tumorigenic growth, suggesting that PYCR1 is a potential cancer target. However, inhibitor development has been stymied by limited mechanistic details for the enzyme, particularly in light of a previous crystallographic study that placed the cofactor-binding site in the C-terminal domain rather than the anticipated Rossmann fold of the N-terminal domain. To fill this gap, we report crystallographic, sedimentation- velocity, and kinetics data for human PYCR1. Structures of binary complexes of PYCR1 with NADPH or proline determined at 1.9 Å resolution provide insight into cofactor and substrate recognition.WeseeNADPHbound to the Rossmann fold, over 25 Å from the previously proposed site. The 1.85 Å resolution structure of a ternary complex containing NADPH and a P5C/proline analog provides a model of the Michaelis complex formed during hydride transfer. Sedimentation velocity shows that PYCR1 forms a concentration-dependent decamer in solution, consistent with the pentamer-of-dimers assembly seen crystallographically. Kinetic and mutational analysis confirmed several features seen in the crystal structure, including the importance of a hydrogen bond between Thr-238 and the substrate as well as limited cofactor discrimination

Regulation of AUXIN RESPONSE FACTOR condensation and nucleo-cytoplasmic partitioning

Auxin critically regulates plant growth and development. Auxin-driven transcriptional responses are mediated through the AUXIN RESPONSE FACTOR (ARF) family of transcription factors. ARF protein condensation attenuates ARF activity, resulting in dramatic shifts in the auxin transcriptional landscape. Here, we perform a forward genetics screen for ARF hypercondensation, identifying an F-box protein, which we named AUXIN RESPONSE FACTOR F-BOX1 (AFF1). Functional characterization of SC

7-rhamnosylated flavonols modulate homeostasis of the plant hormone auxin and affect plant development

Flavonols are a group of secondary metabolites that affect diverse cellular processes. They are considered putative negative regulators of the transport of the phytohormone auxin, by which they influence auxin distribution and concomitantly take part in the control of plant organ development. Flavonols are accumulating in a large number of glycosidic forms. Whether these have distinct functions and diverse cellular targets is not well understood. The rol1-2 mutant of Arabidopsis thaliana is characterized by a modified flavonol glycosylation profile that is inducing changes in auxin transport and growth defects in shoot tissues. To determine whether specific flavonol glycosides are responsible for these phenotypes, a suppressor screen was performed on the rol1-2 mutant, resulting in the identification of an allelic series of UGT89C1, a gene encoding a flavonol 7-O-rhamnosyltransferase. A detailed analysis revealed that interfering with flavonol rhamnosylation increases the concentration of auxin precursors and auxin metabolites, whereas auxin transport is not affected. This finding provides an additional level of complexity to the possible ways by which flavonols influence auxin distribution and suggests that flavonol glycosides play an important role in regulating plant development

A modular analysis of the Auxin signalling network

Auxin is essential for plant development from embryogenesis onwards. Auxin acts in large part through regulation of transcription. The proteins acting in the signalling pathway regulating transcription downstream of auxin have been identified as well as the interactions between these proteins, thus identifying the topology of this network implicating 54 Auxin Response Factor (ARF) and Aux/IAA (IAA) transcriptional regulators. Here, we study the auxin signalling pathway by means of mathematical modeling at the single cell level. We proceed analytically, by considering the role played by five functional modules into which the auxin pathway can be decomposed: the sequestration of ARF by IAA, the transcriptional repression by IAA, the dimer formation amongst ARFs and IAAs, the feedback loop on IAA and the auxin induced degradation of IAA proteins. Focusing on these modules allows assessing their function within the dynamics of auxin signalling. One key outcome of this analysis is that there are both specific and overlapping functions between all the major modules of the signaling pathway. This suggests a combinatorial function of the modules in optimizing the speed and amplitude of auxin-induced transcription. Our work allows identifying potential functions for homo- and hetero-dimerization of transcriptional regulators, with ARF:IAA, IAA:IAA and ARF:ARF dimerization respectively controlling the amplitude, speed and sensitivity of the response and a synergistic effect of the interaction of IAA with transcriptional repressors on these characteristics of the signaling pathway. Finally, we also suggest experiments which might allow disentangling the structure of the auxin signaling pathway and analysing further its function in plants

Tryptophan-dependent auxin biosynthesis is required for HD-ZIP III-mediated xylem patterning

The development and growth of higher plants is highly dependent on the conduction of water and minerals throughout the plant by xylem vessels. In Arabidopsis roots the xylem is organized as an axis of cell files with two distinct cell fates: the central metaxylem and the peripheral protoxylem. During vascular development, high and low expression levels of the class III HD-ZIP transcription factors promote metaxylem and protoxylem identities, respectively. Protoxylem specification is determined by both mobile, ground tissue-emanating miRNA165/6 species, which downregulate, and auxin concentrated by polar transport, which promotes HD-ZIP III expression. However, the factors promoting high HD-ZIP III expression for metaxylem identity have remained elusive. We show here that auxin biosynthesis promotes HD-ZIP III expression and metaxylem specification. Several auxin biosynthesis genes are expressed in the outer layers surrounding the vascular tissue in Arabidopsis root and downregulation of HD-ZIP III expression accompanied by specific defects in metaxylem development is seen in auxin biosynthesis mutants, such as trp2-12, wei8 tar2 or a quintuple yucca mutant, and in plants treated with L-kynurenine, a pharmacological inhibitor of auxin biosynthesis. Some of the patterning defects can be suppressed by synthetically elevated HD-ZIP III expression. Taken together, our results indicate that polar auxin transport, which was earlier shown to be required for protoxylem formation, is not sufficient to establish a proper xylem axis but that root-based auxin biosynthesis is additionally required