Gamma-glutamylcysteine synthetase (gamma-GCS), also known as glutamate-cysteine ligase (EC 6.3.2.2), is the rate-limiting enzyme in the synthesis of glutathione (GSH). The gene GLCLC encodes the catalytic subunit while GLCLR encodes the regulatory subunit. Although it has been shown that GLCLC can respond to a variety of stresses by increased transcription, it is not known whether a similar response occurs for GLCLR. Nor is it known whether post-transcriptional regulation of either gene product is altered during stress. The present investigation was undertaken to explore transcriptional and post-transcriptional regulation of GLCLC and GLCLR gene products when HepG2 cells were challenged with the radiation sensitizer diethyl maleate (DEM). Expression of steady-state GLCLC and GLCLR mRNA was enhanced 5-20-fold after DEM challenge. Nuclear run-off assays were performed on unstressed and stressed cells to determine whether the increased expression of GLCLC and GLCLR mRNA was due to altered transcriptional activity of these genes. The DEM treatment increased the transcription rates of both genes 2-5-fold. In unstressed HepG2 cells, the half-life of GLCLC mRNA transcripts was approximately 4 h. In contrast, the half-life of GLCLR transcripts was approximately 8 h. In cells treated with DEM, the half-lives of all transcripts were increased, indicating that message stabilization contributed to the increased expression of gene products. Finally, a PEST algorithm has identified a PEST (proline, glutamate, serine, threonine) motif within the catalytic subunit of gamma-GCS, suggesting that this subunit might exhibit conditional proteolytic regulation. These results imply that regulation of the products of the GLCLC and GLCLR genes may be altered at multiple levels during exposure to stress.
A series of studies were conducted to examine the residual activity and toxicity of the ecdysone agonists tebufenozide and methoxyfenozide to codling moth, Cydia pomonella (L.), and oriental fruit moth, Grapholita molesta (Busck), in North Carolina apple systems. Methoxyfenozide exhibited greater activity than tebufenozide against codling moth eggs in dose-response bioassays, with a 4.5- and 5.3-fold lower LC50 value to eggs laid on fruit treated before or after oviposition, respectively. Oriental fruit moth eggs were 57- and 12-fold less sensitive to methoxyfenozide than were codling moth eggs on fruit treated before and after oviposition, respectively. Methoxyfenozide was effective in reducing larval entries of both codling moth and oriental fruit moth in field residual activity bioassays, exhibiting activity for at least 28 d after application. Residue breakdown on fruit was approximately 80% at 28 d after treatment for both methoxyfenozide and tebufenozide, with the most rapid residue decline (60%) occurring during the first 14 d after application. Two applications of methoxyfenozide applied at 14-d intervals provided better canopy coverage and higher residue levels than one application. Spray volume (683 versus 2,057 liters/ha) did not affect the efficacy of methoxyfenozide. Leaf and fruit expansion during the season was measured to determine potential plant-growth dilution effects on residual activity. There was very little increase in leaf area after mid May, but increase in fruit surface area over the season was described by a second order polynomial regression. Implications for codling moth and oriental fruit moth management programs are discussed.
A series of studies were conducted to examine the residual activity and toxicity of the ecdysone agonists tebufenozide and methoxyfenozide to codling moth, Cydia pomonella (L.), and oriental fruit moth, Grapholita molesta (Busck), in North Carolina apple systems. Methoxyfenozide exhibited greater activity than tebufenozide against codling moth eggs in dose-response bioassays, with a 4.5- and 5.3-fold lower LC50 value to eggs laid on fruit treated before or after oviposition, respectively. Oriental fruit moth eggs were 57- and 12-fold less sensitive to methoxyfenozide than were codling moth eggs on fruit treated before and after oviposition, respectively. Methoxyfenozide was effective in reducing larval entries of both codling moth and oriental fruit moth in field residual activity bioassays, exhibiting activity for at least 28 d after application. Residue breakdown on fruit was approximately 80% at 28 d after treatment for both methoxyfenozide and tebufenozide, with the most rapid residue decline (60%) occurring during the first 14 d after application. Two applications of methoxyfenozide applied at 14-d intervals provided better canopy coverage and higher residue levels than one application. Spray volume (683 versus 2,057 liters/ha) did not affect the efficacy of methoxyfenozide. Leaf and fruit expansion during the season was measured to determine potential plant-growth dilution effects on residual activity. There was very little increase in leaf area after mid May, but increase in fruit surface area over the season was described by a second order polynomial regression. Implications for codling moth and oriental fruit moth management programs are discussed.
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