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A number of studies have shown that the polyol pathway, consisting of aldose reductase (AR) and sorbitol dehydrogenase (SDH), contributes to ischemia-reperfusion (I/R)-induced myocardial infarction due to depletion of ATP. In this report we show that the polyol pathway in I/R heart also contributes to the impairment of sacro/endoplasmic reticulum Ca2+-ATPase (SERCA) and ryanodine receptor (RyR), two key players in Ca2+ signaling that regulates cardiac contraction. Rat hearts were isolated and retrogradely perfused with either Krebs’ buffer containing 1 μM AR inhibitor, zopolrestat, or 200 nM SDH inhibitor, CP-170,711, and challenged by 30 min of regional ischemia and 45 min of reperfusion. We found that post-ischemic contractile function of the isolated perfused hearts was improved by pharmacological inhibition of the polyol pathway. I/R-induced contractile dysfunction is most likely due to impairment in Ca2+ signaling and the activities of SERCA and RyR. All these abnormalities were significantly ameliorated by treatment with ARI or SDI. We showed that the polyol pathway activities increase the level of peroxynitrite, which enhances the tyrosine nitration of SERCA and irreversibly modify it to form SERCAC674-SO3H. This leads to reduced level of S-glutathiolated SERCA, contributing to its inactivation. The polyol pathway activities also deplete the level of GSH, leading to decreased active RyR, the S-glutathiolated RyR. Thus, in I/R heart, inhibition of polyol pathway improved the function of SERCA and RyR by protecting them from irreversible oxidation.
Polyol Pathway Impairs the Function of SERCA and RyR in Ischemic-Reperfused Rat Hearts by Increasing Oxidative Modifications of These Proteins
Wai Ho Tang,1 Gennadi M. Kravtsov, PhD,1 Martina Sauert,1 Xiao Yong Tong,3 Xiu yun Hou,3 Tak Ming Wong, PhD,1 Sookja K. Chung, PhD,2 and Stephen Sum Man Chung, PhD1,*
2011 Jul 1.
The epidermal growth factor receptor (EGFR) is a receptor tyrosine kinase that is an integral component of proliferative signaling. EGFRs on the cell surface become activated upon EGF binding and have an increased rate of endocytosis. Once in the cytoplasm, the EGF·EGFR complex is trafficked to the lysosome for degradation, and signaling is terminated. During trafficking, the EGFR kinase domain remains active, and the internalized EGFR can continue signaling to downstream effectors. Although effector activity varies based on the EGFR’s endocytic location, it is not clear how this occurs. In an effort to identify proteins that uniquely associate with the internalized, liganded EGFR in the early endosome, we developed an early endosome isolation strategy to analyze their protein composition. Post-nuclear supernatant from HeLa cells stimulated with and without EGF were separated on an isotonic 17% Percoll gradient. The gradient was fractionated, and early endosomal fractions were pooled and immunoisolated with an EEA1 mAb. The isolated endosomes were validated by immunoblot using antibodies against organelle-specific marker proteins and transmission EM. These early endosomes were also subjected to LC-MS/MS for proteomic analysis. Five proteins were detected in endosomes in a ligand-dependent manner: EGFR, RUFY1, STOML2, PTPN23, and CCDC51. Knockdown of RUFY1 or PTPN23 by RNAi indicated that both proteins play a role in EGFR trafficking. These experiments indicate that endocytic trafficking of activated EGFR changes the protein composition, membrane trafficking, and signaling potential of the early endosome.
endosome, epidermal growth factor receptor (EGFR), membrane trafficking, proteomics, receptor endocytosis
Proteomics reveals novel protein associations with early endosomes in an epidermal growth factor-dependent manner
Julie A. Gosney,‡ Daniel W. Wilkey,§ Michael L. Merchant,§ and Brian P. Ceresa‡,1
2018 Apr 20
High-throughput, non-invasive phenotyping is promising for evaluating crop nitrogen (N) use efficiency (NUE) and grain yield (GY) formation under field conditions, but its application for genotypes differing in morphology and phenology is still rarely addressed. This study therefore evaluates the spectral estimation of various dry matter (DM) and N traits, related to GY and grain N uptake (Nup) in high-yielding winter wheat breeding lines. From 2015 to 2017, hyperspectral canopy measurements were acquired on 26 measurement dates during vegetative and reproductive growth, and 48 vegetation indices from the visible (VIS), red edge (RE) and near-infrared (NIR) spectrum were tested in linear regression for assessing the influence of measurement stage and index selection. For most traits including GY and grain Nup, measurements at milk ripeness were the most reliable. Coefficients of determination (R²) were generally higher for traits related to maturity than for those related to anthesis canopy status. For GY (R² = 0.26-0.51 in the three years, p < 0.001), and most DM traits, indices related to the water absorption band at 970 nm provided better relationships than the NIR/VIS indices, including the normalized difference vegetation index (NDVI), and the VIS indices. In addition, most indices including RE bands, notably NIR/RE combinations, ranked above the NIR/VIS group. Due to index saturation, the index differentiation was most apparent in the highest-yielding year. For grain Nup and total Nup, the RE/VIS index MSR_705_445 and the simple ratio R780_R740 ranked highest, followed by other RE indices. Among the vegetative organs, R² values were mostly highest and lowest for leaf and spike traits, respectively. For each trait, index and partial least squares regression (PLSR) models were validated across years at milk ripeness, confirming the suitability of optimized index selection. PLSR improved the prediction errors of some traits but not consistently the R² values. The results suggest the use of sensor-based phenotyping as a useful support tool for screening of yield potential and NUE and for identifying contributing plant traits?which, due to their expensive and cumbersome destructive determination are otherwise not readily available. Water band and RE indices should be preferred over NIR/VIS indices for DM traits and N-related traits, respectively, and milk ripeness is suggested as the most reliable stage.
phenomics, smart farming, remote sensing, nitrogen use efficiency, yield prediction, red edge, water band indices, breeding
High-Throughput Field Phenotyping Traits of Grain Yield Formation and Nitrogen Use Efficiency: Optimizing the Selection of Vegetation Indices and Growth Stages
Lukas Prey, Yuncai Hu, and Urs Schmidhalter*