GABA accumulation and the hypersensitive response in isolated mesophyll cells treated with the G protein activator mastoparan

GABA (y-amino butyric acid) is a non-protein amino acid synthesized through the a-decarboxylation of L-glutamate. This reaction is catalyzed by L-glutamate decarboxylase (EC 4.1.1.15), a cytosolic Ca2+/calmodulin-stimulated enzyme. The purpose of this study is to determine whether or not GABA accumulation is associated with the hypersensitive response of isolated Asparagus sprengeri mesophyll cells. The addition of 25 J.lM mastoparan, a G protein activator, to suspensions of isolated asparagus mesophyll cells significantly increased GABA synthesis and cell death. Cell death was assessed using Evan's blue dye and fluorescein diacetate tests for cell viability. In addition, mastoparan stimulated pH-dependent alkalinization of the external medium, and a rapid and large 02 consumption followed by a loss of photosynthetic activity. The rate of 02 consumption and the net decrease in 02 in the dark was enhanced by light. The inactive mastoparan analogue Mas17 was ineffective in stimulating GABA accumulation, medium alkalinization, 02 uptake and cell death. Accumulation of H202 in response tomastoparan was not detected, however, mastoparan caused the cell-dependent degradation of added H202. The pH dependence of mastoparan-stimulated alkalinization suggests cellular electrolyte leakage, while the consumption of 02 corresponds to the oxidative burst in which 02 at the cell surface is reduced to form various active oxygen species. The results are indicative of the "hypersensitive response" of plants to pathogen attack, namely, the death of cells in the locality of pathogen invasion. The data are compatible with a model in which mastoparan triggers G protein activity, subsequent intracellular signal transduction pathway/s, and the hypersensitive response. It is postulated that the physiological elicitation of the hypersensitive response involves G protein signal transduction. The synthesis of GABA during the hypersensitive response has not been documented previously; however the role/s of GABA synthesis in the hypersensitive response, if any, remain unclear.

Isolation and characterization of a chitinase-secreting mutant of mortierella pusilla with altered interaction with the mycoparasite piptocephalis virginiana

Mortierella pusilla is a susceptible host and supports good growth of the mycoparasite, Piptocephalis virginiana. Uninucleate spores of M. pusilla were sUbjected to N-methyl-N'-nitro-nitrosoguanidine (MNNG). To attain a high mutation frequency , a 1o-minute exposure to 10 mg/ml MNNG was used and lead to the survival of about 10 % of the spores. The exposed spores then were plated on chitin or milk plates. Approximately 30,000 colonies were examined after mutagenesis on the screening media. A strain, MUT23 , with abnormal slow growth morphology was found to delay parasitism by £. virginiana. The particular morphology was not due to auxotrophy, because this strain displayed normal hyphae when glucose was used as the sole carbon source. One interesting phenomenon was that MUT23 showed an extensive clearing zone around the colony on colloidal chitin agar after 20-25 d. On the same conditions, wild type strain did not show this phenotype. In addition, the MUT23 strain produced the same normal hypha as the wild type strain when it was grown on colloidal chitin agar. The MUT23 was also able to produce more spores on colloidal chitin agar than on malt-yeast extract and minimal media. The parasite germ tubes formed appressoria at the point of contact on the cell surface of wild type and MUT23 grown for 6 days cell surface but not on the cel surface of MUT23 grown for 2 days. Thus, interaction between MUT23 strain and the mycoparasite was dependent on MUT23 age. The effect of MUT23 filtrate on germination of the parasite was tested. Lysis of germinated spores of the parasite were observed in concentrated MUT23 filtered solution. MUT23 was compared to the wild type strain for their chitinase production in sUbmerged culture. The chitinase isozymes of both wild type and MUT23 were shown by immunoblotting. Eight distinct chitinase molecules were detected. MUT23 showed markedly higher chitinase activity than the wild type cultured in chitin-containing medium. Maximum chitinase activities of MUT23 were 13.5 fold higher at 20 day of the culture then that of wild type.

The establishment of two novel bovine cell lines by transfection with the putative transforming region of bovine adenovirus type 3

Human adenoviruses (Ads), members of the family adenoviridae, are medium-sized DNA viruses which have been used as valuable research tools for the study of RNA processing, oncogenic transformation, and for the development of viral vectors for use in gene delivery and immunization technology. The left 12% of the linear Ad genollle codes for products which are necessary for the efficient replication of the virus, as well as being responsible for the forlllation of tumors in animallllodels. The establishlllent of the 293 cell line, by immortalization of human embryonic kidney cells with th~ E1 region of Ad type S (AdS), has facilitated extensive manipulation of the Ads and the development of recombinant Ad vectors. The study of bovine adenoviruses (BAVs), which cause mild respiratory and gastrointestinal infections in cattle has, on the other hand, been limited primarily to that of infectivity, immunology and clinicallllanifestations. As a result, any potential as gene delivery vehicles has not yet been realized. Continued research into the molecular biolo~gy of BAVs and the development of recolllbinant vectors would benefit from the development of a cell line analogous to that of the 293 cells. In an attelllpt to establish such a cell line, the recombinant plaslllid pKC-neo was constructed, containing the left 0-19.7% of the BAV type 3 (BAV3) genome, and the selectable marker for resistance to the aminoglycoside G418, a neomycin derivative. The plasmid construct was then used to transfect both the Madin-Darby bovine kidney (MDBK) -iicell line and primary bovine lung cells, after which G418-resistant foci were selected for analysis. Two cell lines, E61 (MDBK) and E24 (primary lung), were subsequently selected and analysed for DNA content, revealing the presence of the pKC-neo sequences in their respective genomes. In addition, BAV3 RNA transcripts were detected in the E61 cells. Although the presence of E1 products has yet to be confirmed in both cell lines, the E24 cells exhibit a phenotype characteristic of partial transformation by E1. The apparent immortalization of the primary lung cells will permit exploitation of their ability to take up exogenous DNA at high efficiency.

The practical examination and how it correlates with the triple jump, tutorial, and written examination

This study explored the relationship between the practical examination and other course evaluation methods~ specifically, the triple jump, tutorial, and written examination. Studies correlating academic and clinical grades tended to indicate that they may not be highly correlated because each evaluation process contributes different kinds of information regarding student knowledge, skills, and attitudes. Six hypotheses were generated stating a positive relationship between the four evaluation methods. A correlation matrix was produced of the Pearson Product Moment correlation co-efficients on the four evaluation methods in the second and third year Occupational Therapy Technique and Clinical Problem Solving courses of the 1988 and 1989 graduates (n~45). The results showed that the highest correlations existed between the triple jump and the tutorial grades and the lowest correlations existed between the practical examination and written examination grades. Not all of the correlations~ however~ reached levels of significance. The correlations overall. though, were only low to moderate at best which indicates that the evaluation methods may be measuring different aspects of student learning. This conclusion supports the studies researched. The implications and significance of this study is that it will assist the faculty in defining what the various evaluation methods measure which will in turn promote more critical input into curriculum development for the remaining years of the program.

The encounter with the real and post-Soviet trauma : fantasy construction in Russian popular cinema /

This thesis examines the impact of the Soviet Union's collapse on the Russian Symbolic as represented through popular cinema of the post-Soviet period. The disintegration of the USSR in 1991 became one of the most traumatic experiences for many Russian people. The trauma of the collapse of the Soviet Union penetrated the everyday reality of the Russian Symbolic, leaving the traces-symptoms in different cultural fonns like literature, arts, television and cinema. Because popular culture usually reacts very quickly to any social, political and economical shifts in society, it is an excellent barometer for deeper changes in society. Focusing on postSoviet popular cinema, this thesis analyzes the symptoms of cultural and individual trauma occasioned by the momentous changes of the 1990's. This study is grounded in post-analytic theory of Jacques Lacan and its interpretation by Slavoj Zizek, which emphases the traumatic encounter with the Real as a "hard core" of our reality. According to this paradigm, a new chain of signifiers is structured around the traumatic breach in the Symbolic, initiating a process of fantasy construction to deal with consequences of trauma and, thus, to support our Symbolic order. This thesis examines three major fantasy constructions - drinking, traveling to a "happy land" and family reunion and money - in popular films by Alexander Rogozhkin, Yurij Mamin, Georgij Shengelia, Dmitrij Astrakhan, Valerij Todorovskij, Alexej Balabanov, Sergej Bodrov Jr. and Petr Buslov. According to Zizek, enjoyment underlies any fantasy constructions, and that is why after the intrusion of the Real every individual and culture should go through the process of fantasizing about some substitutes which can help to minimize the traumatic effect and which can lead to a partial enjoyment. By analyzing the fantasies about drinking, "happy land", reconstruction of the family bonds and money in Russian popular cinema since 1991, this thesis demonstrates how the traumatic engagement with the Real affected the everyday lives of Russian people, and how individuals tried to fill the gap, the lack, in the post-Soviet Symbolic and "return" the lost feeling of unity and plenitude.

Outline of the evolution of the Falls of Niagara, contrast with the Falls of Zambesi /

At head of title: For the International Geological Congress.

A compressed view of the points to be discussed, in treating with the United States of America : with an appendix /

Appendix: On the conduct of the government of United States towards the Indian tribes: p.[129]-139.

Letter from the Secretary of State to Mr. Monroe, on the subject of the attack on the Chesapeake : the correspondence of Mr. Monroe with the British Government ; and also Mr. Madison's correspondence with Mr. Rose on the same subject. --

March 23, 1808. Printed by order of the House of Representatives.

Message from the President of the United States, transmitting copies of a letter from the British Secretary of State for Foreign Affairs, to the Secretary of State, with the answer of the latter.--

January 6, 1814. Ordered to lie on the table.

A true and impartial account of the actions fought at Chippawa & Lundy's Lane during the last war with the United States /

On leaf preceding the t.p.: Battles of Chippawa and Lundy's Lane, by Charles Anderson, guide to the battle ground.