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by
Tao Xu |
Total Records (
3 ) for
Tao Xu |
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Ameeta K. Agarwal
,
Tao Xu
,
Melissa R. Jacob
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Qin Feng
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Michael C. Lorenz
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Larry A. Walker
and
Alice M. Clark
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Sampangine, a plant-derived alkaloid found in the Annonaceae family, exhibits strong inhibitory activity against the opportunistic fungal pathogens Candida albicans, Cryptococcus neoformans, and Aspergillus fumigatus. In the present study, transcriptional profiling experiments coupled with analyses of mutants were performed in an effort to elucidate its mechanism of action. Using Saccharomyces cerevisiae as a model organism, we show that sampangine produces a transcriptional response indicative of hypoxia, altering the expression of genes known to respond to low-oxygen conditions. Several additional lines of evidence obtained suggest that these responses could involve effects on heme. First, the hem1Δ mutant lacking the first enzyme in the heme biosynthetic pathway showed increased sensitivity to sampangine, and exogenously supplied hemin partially rescued the inhibitory activity of sampangine in wild-type cells. In addition, heterozygous mutants with deletions in genes involved in five out of eight steps in the heme biosynthetic pathway showed increased susceptibility to sampangine. Furthermore, spectral analyses of pyridine extracts indicated significant accumulation of free porphyrins in sampangine-treated cells. Transcriptional profiling experiments were also performed with C. albicans to investigate the response of a pathogenic fungal species to sampangine. Taking into account the known differences in the physiological responses of C. albicans and S. cerevisiae to low oxygen, significant correlations were observed between the two transcription profiles, suggestive of heme-related defects. Our results indicate that the antifungal activity of the plant alkaloid sampangine is due, at least in part, to perturbations in the biosynthesis or metabolism of heme. |
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Caixia Lv
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Maorong Chen
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Geliang Gan
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Lifen Wang
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Tao Xu
and
Jiuping Ding
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Large conductance, voltage- and Ca2+-activated K+ (BK) channels encoded by the mslo α and β2 subunits exist abundantly in rat chromaffin cells, pancreatic β cells, and DRG neurons. The extracellular loop of hβ2 acting as the channel regulator influences the rectification and toxin sensitivity of BK channels, and the inactivation domain at its N terminus induces rapid inactivation. However, the regulatory mechanism, especially the trafficking mechanism of hβ2, is still unknown. With the help of immunofluorescence and patch clamp techniques, we determine that the hβ2 subunit alone resides in the endoplasmic reticulum, suggesting that trafficking mechanism of hβ2 differs from that of hβ1 opposite to what we predicted previously. We further demonstrate that a four-turn α helical segment at the N terminus of hβ2 prevents the surface expression of hβ2, that is, the helical segment itself is a retention signal. Using the c-Myc epitope-tagged extracellular loop of hβ2, we reveal that the most accessible site by antibody is located at the middle of the extracellular loop, which might provide clues to understand how the auxiliary β subunits regulates the toxin sensitivity and the rectification of BK-type channels. |
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Li Jiang
,
Junmei Fan
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Li Bai
,
Yan Wang
,
Yu Chen
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Lu Yang
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Liangyi Chen
and
Tao Xu
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Insulin-stimulated GLUT4 translocation to the plasma membrane constitutes a key process for blood glucose control. However, convenient and robust assays to monitor this dynamic process in real time are lacking, which hinders current progress toward elucidation of the underlying molecular events as well as screens for drugs targeting this particular pathway. Here, we have developed a novel dual colored probe to monitor the translocation process of GLUT4 based on dual color fluorescence measurement. We demonstrate that this probe is more than an order of magnitude more sensitive than the current technology for detecting fusion events from single GLUT4 storage vesicles (GSVs). A small fraction of fusion events were found to be of the "kiss-and-run" type. For the first time, we show that insulin stimulation evokes a ~40-fold increase in the fusion of GSVs in 3T3-L1 adipocytes, compared with basal conditions. The probe can also be used to monitor the prefusion behavior of GSVs. By quantifying both the docking and fusion rates simultaneously, we demonstrate a proportional inhibition in both docking and fusion of GSVs by a dominant negative mutant of AS160, indicating a role for AS160 in the docking of GSVs but not in the regulation of GSV fusion after docking. |
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