1985 — 1989 |
Komuniecki, Richard W |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Volatile Acid Formation by Parasitic Helminths
Parasitic helminths pose significant health problems throughout the world but, in many cases, adequate means of chemotherapy still are not available. It is clear that many helminths exhibit a predominantly anaerobic energy metabolism which differs substantially from that of the host, and it appears that many potent anthelmintics specifically interfere with helminth energy metabolism. However, before the differences between host and parasite can be fully exploited, the energy-generating reactions operating in these parasites need to be examined in more detail. The present study is designed to characterize the regulation and energetics of volatile fatty acid synthesis in mitochondria isolated from the porcine nematode, Ascaris suum. This organism has served as an excellent model for many studies of helminth metabolism. Its mitochondria function anaerobically, using unsaturated organic acids, instead of O2, as terminal electron-acceptors and it excretes acetate, propionate and the branched-chain fatty acids (BFA), 2-methylbutyrate and 2-methylvalerate, as end products of carbohydrate degradation. However, little is known about the factors controlling their formation. In the present study, the rates of pyruvate decarboxylation and BFA formation in intact mitochondria will be correlated with the activity state of the pyruvate dehydrogenase complex and the levels of a number of physiologically important intramitochondrial metabolites. In addition, pyruvate dehydrogenase phosphatase and the membrane-bound electron-transport components involved in BFA synthesis will be isolated and characterized. Since all helminths exhibiting anaerobic mitochondrial metabolisms excrete acetate and contain similar electron-transport systems, these studies should have broad application. Hopefully, once the enzymatic differences between host and parasite metabolism have been identified and evaluated, a more sensible approach to chemotherapy can be adopted. In addition, the more that is known about the biochemical mode of action of anthelmintics, the more rational efficient drug usage will become.
|
1 |
1988 — 1991 |
Komuniecki, Patricia (co-PI) [⬀] Wheelock, Margaret [⬀] Komuniecki, Richard |
N/AActivity Code Description: No activity code was retrieved: click on the grant title for more information |
Acquistion of a Zeiss Axiophot Photomicroscope
This is a request for funds to obtain a fluorescence microscope that will be used primarily by two research groups. One group is investigating the mechanism by which human epithelial cells recognize and adhere to each other. The process seems to be mediated by an integral membrane protein which has been purified and against which antibodies have been produced. The second group is investigating proteins involved in the shift between aerobic and anaerobic metabolism in parasitic helminths. Mitochondrial proteins have been purified and antibodies generated here also. The antibodies in both studies will be fluorescently labelled and then used as probes to localize the proteins of interest in the cellular structures, using the requested instrumentation.
|
0.915 |
1990 — 2004 |
Komuniecki, Richard W |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Volatile Acid Formation in Parasitic Helminths
Many effective anthelminthics inhibit muscle contraction, either by directly interfering with processes at the neuromuscular junction or inhibiting energy metabolism. Since the mitochondrial metabolism of many parasitic helminths is predominantly anaerobic and differs substantially from that of the host, it is a potential site for chemotherapeutic attack. During the previous grant period, a number of important insights into the regulation of the enzymes involved in the mitochondrial metabolism of the parasitic nematode, Ascaris suum, have been described. The present study is designed to extend these studies and concentrate on differences that have been identified between host and parasite. Specifically, a novel component of the ascarid pyruvate dehydrogenase complex (PDC) will be further characterized and its role and that of E2 acetylation in the regulation of PDC activity by its associated kinase and phosphatase will be identified. In addition, the regulation of NADH-dependent 2-methyl branched-chain enoyl CoA reduction will be characterized using submitochondrial particles and purified ascarid enzymes, and the generation of a proton-gradient and ADP phosphorylation measured directly. Since Ca++ appears to play a unique role in the regulation of ascarid mitochondrial metabolism, Ca++ fluxes in isolated ascarid mitochondria will be measured and correlated with end-product formation and energy- generation. Preliminary experiments have suggested that mitochondrial volume is an important regulator of organic acid formation in these isolated mitochondria so that its role also will be described. These studies will permit the development of well characterized preparations of ascarid mitochondria, capable of volatile organic acid synthesis at physiological levels, and should provide a much clearer picture of energy-generation in these often studied, but still poorly understood organelles. Since these mitochondria are often used to screen compounds for potential anthelminthic activity, a clear understanding of their function is essential to intercept results meaningfully, especially since they differ so profoundly from the corresponding mammalian organelles.
|
1 |
2000 — 2009 |
Komuniecki, Richard Walter |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Regulation of Pharyngeal Pumping in Parasitic Nematodes
DESCRIPTION (provided by the applicant): Biogenic amines, such as serotonin, tyramine, octopamine and dopamine regulate a number of key processes in parasitic nematodes, including, feeding, pharyngeal pumping, locomotion, egg-laying and many more complex behaviors. During the previous grant period, we have begun to characterize the pharmacology, coupling and localization of a number of biogenic amine receptors in both free-living and parasitic nematodes with the goal of identifying the specific receptors regulating these key processes. During the course of these studies, we have also identified a novel tyraminergeric signaling pathway and presynaptic 5-HT receptor-linked signaling complexes that are localized by a multi-PDZ domain containing scaffolding protein that appear to be essential for efficient signal transduction. The present study is focused on the definitive identification of the biogenic amine receptors regulating pharyngeal pumping in the parasitic nematode, Ascaris suum, on the assumption that interference with these receptors, especially by stimulating inhibitory inputs, as has been described for ivermectin and glutamate-gated chloride channels, will have a deleterious effect of the worms. In addition, we hypothesize that receptor localization can be as important as receptor function for effective signal transduction and plan to test this hypothesis using mutants and a "dominant negative" approach, once the other components of these multi-protein signaling complexes have been confirmed by direct protein interaction and in vivo co-localization.
|
1 |
2008 — 2012 |
Komuniecki, Richard Walter |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Locomotion in Parasitic Nematodes
[unreadable] DESCRIPTION (provided by applicant): Parasitic nematodes are a major cause of human morbidity and contribute significantly to a loss of Disability Adjusted Life Years. More importantly, effective chemotherapy is still not available to control some infections, such as lymphatic filariasis or river blindness. Perhaps less well appreciated, but equally important for proper human nutrition and health, is the devastating economic impact of nematode parasites on livestock and plants and new anti-helminthic and drug targets are both desperately needed in all settings. Serotonin (5-HT) dramatically inhibits locomotion and the contraction of body wall muscle in a variety of free-living and parasitic nematodes, suggesting that serotonergic signaling may be an excellent target for drug discovery. We propose to use a dual systems approach, designed to exploit the experimental advantages of both the C. elegans and A. suum model systems, to characterize the receptors and downstream signaling components in the pathway of 5-HT inhibition of locomotion. First, the role of 5-HT signaling in the inhibition of locomotion will be characterized in C. elegans by bioinformatics, the heterologous expression of cloned receptors and the use of putative 5-HT receptor null mutants, with special focus on the identification of the receptors and their downstream signaling pathways. Second, potential orthologues of the relevant C. elegans 5-HT receptors will be cloned and characterized from A. suum and the proposed pathway of 5-HT inhibition of locomotion generated from C. elegans tested by examination of relevant physiological end points in A. suum (i.e. locomotion and muscle contraction). Because of the enormous diversity among nematodes, processes in C. elegans may not be duplicated exactly in the parasites, so the results of this study should not only identify key targets in serotonergic signaling pathways, but also highlight potential differences between these two important model systems. Parasitic nematodes cause significant medical, veterinary and agricultural problems worldwide. The present studies are designed to characterize the receptors and downstream signaling components involved in the 5-HT inhibition of locomotion. Locomotion is a key target for anti-nematodal drug discovery and these studies should provide basic insights into pathways regulating neurotransmitter release and the activity state of body wall muscle, as well as identifying a wealth of potential novel molecular targets for drug discovery. [unreadable] [unreadable] PROJECT NARRATIVE Parasitic nematodes cause significant medical, veterinary and agricultural problems worldwide. These studies are designed to characterize the receptors and downstream signaling components involved in serotonin inhibition of locomotion. Locomotion is a key target for anti-nematodal drug discovery and these studies should provide insight into pathways regulating neurotransmitter release and the activity state of body wall muscle, as well as identifying a wealth of potential novel drug targets. [unreadable] [unreadable] [unreadable]
|
1 |
2015 |
Komuniecki, Richard Walter |
R15Activity Code Description: Supports small-scale research projects at educational institutions that provide baccalaureate or advanced degrees for a significant number of the Nation’s research scientists but that have not been major recipients of NIH support. The goals of the program are to (1) support meritorious research, (2) expose students to research, and (3) strengthen the research environment of the institution. Awards provide limited Direct Costs, plus applicable F&A costs, for periods not to exceed 36 months. This activity code uses multi-year funding authority; however, OER approval is NOT needed prior to an IC using this activity code. |
Paralysis of Nematode Parasites
? DESCRIPTION (provided by applicant): Nematode infections are a major cause of human morbidity and contribute significantly to a loss of Disability Adjusted Life Years. More importantly, in many cases, such as filarial infection, effective chemotherapy is still not availabe. Perhaps less well appreciated, but equally important for human health, is the devastating economic impact of parasitic nematodes on livestock and plants, and new anthelmintic and drug targets are both desperately needed in all settings. Most anthelmintic in use against nematode infections act as agonists at key receptors and cause paralysis by interfering with muscle contraction and/or locomotion. The overall objective of this application is to promote the development of new anthelmintic that target locomotion to cause paralysis. We have characterized key monoamine receptors regulating locomotion using an innovative 'dual systems' approach that exploited the experimental advantages of the C. elegans and Ascaris suum models. Highlighting the utility of this approach, C. elegans molecular genetics was instrumental in isolating key receptor genes; bioinformatics approaches then identified corresponding parasitic nematode cDNAs, and ultimately, four key monoamine receptors were identified as promising possible anthelmintic targets. In the present study we will develop innovative anthelmintic drug screening strategies based on these receptors by characterizing their agonist sensitivities in heterologous cells, and expressing them in C. elegans, to create 'chimeric' nematodes to confirm orthodoxy and allow agonist screening under physiological conditions. We will establish the sites of action and physiological roles of these receptors in the locomotors circuitry of parasitic nematodes by direct functional localization and electrophysiological approaches. These studies will not only identify definitively key monoamine receptors regulating locomotion in parasitic nematodes, but because of the enormous diversity among nematodes, they will also highlight the potential differences between these two important model systems. Locomotion is critical to the survival of parasitic nematodes, and drugs that inhibit locomotion can successfully clear parasitic nematode infection. These studies will identify a wealth of potential novel molecular targets for drug discovery.
|
1 |