scholarly journals Data-Independent Acquisition-Based Proteome and Phosphoproteome Profiling Reveals Early Protein Phosphorylation and Dephosphorylation Events in Arabidopsis Seedlings upon Cold Exposure

2021 ◽  
Vol 22 (23) ◽  
pp. 12856
Author(s):  
Jinjuan Tan ◽  
Zhongjing Zhou ◽  
Hanqian Feng ◽  
Jiayun Xing ◽  
Yujie Niu ◽  
...  
Keyword(s):  
Protein Phosphorylation ◽  
Molecular Mechanisms ◽  
Cold Treatment ◽  
Temperature Drop ◽  
Mass Spectrometry Analysis ◽  
Cold Response ◽  
E3 Ligases ◽  
Data Independent Acquisition ◽  
Early Protein ◽  
Cold Adaption

Protein phosphorylation plays an important role in mediating signal transduction in cold response in plants. To better understand how plants sense and respond to the early temperature drop, we performed data-independent acquisition (DIA) method-based mass spectrometry analysis to profile the proteome and phosphoproteome of Arabidopsis seedlings upon cold stress in a time-course manner (10, 30 and 120 min of cold treatments). Our results showed the rapid and extensive changes at the phosphopeptide levels, but not at the protein abundance levels, indicating cold-mediated protein phosphorylation and dephosphorylation events. Alteration of over 1200 proteins at phosphopeptide levels were observed within 2 h of cold treatment, including over 140 kinases, over 40 transcriptional factors and over 40 E3 ligases, revealing the complexity of regulation of cold adaption. We summarized cold responsive phosphoproteins involved in phospholipid signaling, cytoskeleton reorganization, calcium signaling, and MAPK cascades. Cold-altered levels of 73 phosphopeptides (mostly novel cold-responsive) representing 62 proteins were validated by parallel reaction monitoring (PRM). In summary, this study furthers our understanding of the molecular mechanisms of cold adaption in plants and strongly supports that DIA coupled with PRM are valuable tools in uncovering early signaling events in plants.

scholarly journals iTRAQ-based quantitative proteomics analysis of cantaloupe (Cucumis melo var. saccharinus) after cold storage

2020 ◽  
Author(s):  
Wen Song ◽  
Fengxian Tang ◽  
Wenchao Cai ◽  
Qin Zhang ◽  
Fake Zhou ◽  
...  
Keyword(s):  
Cold Stress ◽  
Cold Tolerance ◽  
Proteomic Analysis ◽  
Molecular Mechanisms ◽  
Treatment Time ◽  
Chilling Injury ◽  
Cold Treatment ◽  
Cold Response ◽  
Low Temperature Storage ◽  
Metabolic Analysis

Abstract Background: During the low temperature storage, cantaloupe is susceptible to the cold stress, resulting in the loss of edible and commercial quality. To ascertain the molecular mechanisms of cold tolerance in cantaloupe, cold-sensitive cultivar Goldqueen-308 (GE) and cold-tolerant cultivar Jiashi-310 (JS) were used for quantitative proteomic analysis with iTRAQ in parallel. Results: In this work, two commercial cultivars were treated at 0.5℃ for 0, 12 and 24 days. Phenotypes assays showed that GE suffered a more severe damage as the cold treatment time extended. Proteomic analysis revealed that the number of differentially expressed proteins (DEPs) changed significantly over time in cold-exposed cantaloupe. Comparing with GE, JS responded in a prompter manner in terms of expressing cold-responding proteins during the similarly cold treatment. Furthermore, much more different groups of proteins were mobilized in response to the cold treatment in JS comparing with GE. Metabolic analysis indicated that more amino acids were up-regulated in JS during the early phases of cold stress. This study also identified some DEPs since they were up-regulated in JS or down-regulated in GE in terms of molecular mechanisms, which were mainly related to carbohydrate and energy metabolism, structure proteins, ROS scavening, amino acid metabolic and signaling transduction. Moreover, iTRAQ analysis was confirmed to be reliable via the results of phenotypes assays, metabolic analysis and q-PCR validation. Conclusion: By proteomics information,we found that the prompt response and the significant mobilization of proteins in JS maintained a higher level of cold tolerance, and the delay of cold response in GE could be a critical reason for the severe chilling injury. The candidate proteins we found will be the basis of future studies for further investigations and our findings may help to better understand the novel mechanisms of cold tolerance in cantaloupe.

scholarly journals Comparative proteomics analysis reveals the molecular mechanism of enhanced cold tolerance through ROS scavenging in winter rapeseed (Brassica napus L.)

PLoS ONE ◽  
2021 ◽  
Vol 16 (1) ◽  
pp. e0243292
Author(s):  
Wenbo Mi ◽  
Zigang Liu ◽  
Jiaojiao Jin ◽  
Xiaoyun Dong ◽  
Chunmei Xu ◽  
...  
Keyword(s):  
Cold Stress ◽  
Cold Tolerance ◽  
Molecular Mechanisms ◽  
Early Stage ◽  
Cold Treatment ◽  
Ros Scavenging ◽  
Brassica Napus L ◽  
Membrane Injury ◽  
Data Independent Acquisition ◽  
Winter Rapeseed

Two winter rapeseed cultivars, “NS” (cold tolerant) and “NF” (cold sensitive), were used to reveal the morphological, physiological, and proteomic characteristics in leaves of plants after treatment at -4°C for 12 h(T1) and 24 h(T2), and at room temperature(T0), to understand the molecular mechanisms of cold tolerance. Antioxidant activity and osmotic adjustment ability were higher, and plasma membrane injury was less obvious, in NS than in NF under cold stress. We detected different abundant proteins (DAPs) related to cold tolerance in winter rapeseed through data-independent acquisition (DIA). Compared with NF, A total of 1,235 and 1,543 DAPs were identified in the NSs under T1 and T2, respectively. Compared with NF, 911 proteins were more abundant in NS only after cold treatment. Some of these proteins were related to ROS scavenging through four metabolic pathways: lysine degradation; phenylalanine, tyrosine, and tryptophan; flavonoid biosynthesis; and ubiquinone and other terpenoid-quinone biosynthesis. Analysis of these proteins in the four candidate pathways revealed that they were rapidly accumulated to quickly enhance ROS scavenging and improve the cold tolerance of NS. These proteins were noticeably more abundant during the early stage of cold stress, which was critical for avoiding ROS damage.

scholarly journals iTRAQ-based quantitative proteomics analysis of cantaloupe (Cucumis melo var. saccharinus) after cold storage

2020 ◽  
Author(s):  
Wen Song ◽  
Fengxian Tang ◽  
Wenchao Cai ◽  
Qin Zhang ◽  
Fake Zhou ◽  
...  
Keyword(s):  
Cold Stress ◽  
Cold Tolerance ◽  
Proteomic Analysis ◽  
Molecular Mechanisms ◽  
Treatment Time ◽  
Chilling Injury ◽  
Cold Treatment ◽  
Cold Response ◽  
Low Temperature Storage ◽  
Metabolic Analysis

Abstract Background: During the low temperature storage, cantaloupe is susceptible to the cold stress, resulting in the loss of edible and commercial quality. To ascertain the molecular mechanisms of cold tolerance in cantaloupe, cold-sensitive cultivar Goldqueen-308 (GE) and cold-tolerant cultivar Jiashi-310 (JS) were used for quantitative proteomic analysis with iTRAQ in parallel. Results: In this work, two commercial cultivars were treated at 0.5℃ for 0, 12 and 24 days. Phenotypes assays showed that GE suffered a more severe damage as the cold treatment time extended. Proteomic analysis revealed that the number of differentially expressed proteins (DEPs) changed significantly over time in cold-exposed cantaloupe. Comparing with GE, JS responded in a prompter manner in terms of expressing cold-responding proteins during the similarly cold treatment. Furthermore, much more different groups of proteins were mobilized in response to the cold treatment in JS comparing with GE. Metabolic analysis indicated that more amino acids were up-regulated in JS during the early phases of cold stress. This study also identified some DEPs since they were up-regulated in JS or down-regulated in GE in terms of molecular mechanisms, which were mainly related to carbohydrate and energy metabolism, structure proteins, ROS scavening, amino acid metabolic and signaling transduction. Moreover, iTRAQ analysis was confirmed to be reliable via the results of phenotypes assays, metabolic analysis and q-PCR validation. Conclusion: By proteomics information,we found that the prompt response and the significant mobilization of proteins in JS maintained a higher level of cold tolerance, and the delay of cold response in GE could be a critical reason for the severe chilling injury. The candidate proteins we found will be the basis of future studies for further investigations and our findings may help to better understand the novel mechanisms of cold tolerance in cantaloupe.

The Role of Ubiquitin E3 Ligase in Atherosclerosis

2020 ◽  
Vol 28 (1) ◽  
pp. 152-168
Author(s):  
Zhi-Xiang Zhou ◽  
Zhong Ren ◽  
Bin-Jie Yan ◽  
Shun-Lin Qu ◽  
Zhi-Han Tang ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Vascular Smooth Muscle ◽  
Smooth Muscle Cell ◽  
Muscle Cell ◽  
Vascular Smooth Muscle Cell ◽  
Protein Modification ◽  
Molecular Mechanisms ◽  
Cholesterol Metabolism ◽  
Atherosclerotic Cardiovascular Disease ◽  
E3 Ligases

: Atherosclerosis is a chronic inflammatory vascular disease. Atherosclerotic cardiovascular disease is the main cause of death in both developed and developing countries. Many pathophysiological factors, including abnormal cholesterol metabolism, vascular inflammatory response, endothelial dysfunction and vascular smooth muscle cell proliferation and apoptosis, contribute to the development of atherosclerosis and the molecular mechanisms underlying the development of atherosclerosis are not fully understood. Ubiquitination is a multistep post-translational protein modification that participates in many important cellular processes. Emerging evidence suggests that ubiquitination plays important roles in the pathogenesis of atherosclerosis in many ways, including regulation of vascular inflammation, endothelial cell and vascular smooth muscle cell function, lipid metabolism and atherosclerotic plaque stability. This review summarizes important contributions of various E3 ligases to the development of atherosclerosis. Targeting ubiquitin E3 ligases may provide a novel strategy for the prevention of the progression of atherosclerosis.

scholarly journals Cold-Active β-Galactosidases: Insight into Cold Adaption Mechanisms and Biotechnological Exploitation

Marine Drugs ◽  
2021 ◽  
Vol 19 (1) ◽  
pp. 43
Author(s):  
Marco Mangiagalli ◽  
Marina Lotti
Keyword(s):  
Low Temperature ◽  
Molecular Mechanisms ◽  
Building Blocks ◽  
Cold Active ◽  
Cold Adaption ◽  
Glycosyl Donor ◽  
Pharmaceutical Industries ◽  
Biotechnological Processes ◽  
Hydrolysis Of ◽  
Insight Into

β-galactosidases (EC 3.2.1.23) catalyze the hydrolysis of β-galactosidic bonds in oligosaccharides and, under certain conditions, transfer a sugar moiety from a glycosyl donor to an acceptor. Cold-active β-galactosidases are identified in microorganisms endemic to permanently low-temperature environments. While mesophilic β-galactosidases are broadly studied and employed for biotechnological purposes, the cold-active enzymes are still scarcely explored, although they may prove very useful in biotechnological processes at low temperature. This review covers several issues related to cold-active β-galactosidases, including their classification, structure and molecular mechanisms of cold adaptation. Moreover, their applications are discussed, focusing on the production of lactose-free dairy products as well as on the valorization of cheese whey and the synthesis of glycosyl building blocks for the food, cosmetic and pharmaceutical industries.

scholarly journals Polymodal Functionality of C. elegans OLL Neurons in Mechanosensation and Thermosensation

2021 ◽  
Author(s):  
Yuedan Fan ◽  
Wenjuan Zou ◽  
Jia Liu ◽  
Umar Al-Sheikh ◽  
Hankui Cheng ◽  
...  
Keyword(s):  
Glutamate Receptor ◽  
Sensory Neurons ◽  
Molecular Mechanisms ◽  
Temperature Sensitive ◽  
Cold Sensation ◽  
Cold Response ◽  
C Elegans ◽  
Sensory Modalities ◽  
A Cell ◽  
Bona Fide

AbstractSensory modalities are important for survival but the molecular mechanisms remain challenging due to the polymodal functionality of sensory neurons. Here, we report the C. elegans outer labial lateral (OLL) sensilla sensory neurons respond to touch and cold. Mechanosensation of OLL neurons resulted in cell-autonomous mechanically-evoked Ca2+ transients and rapidly-adapting mechanoreceptor currents with a very short latency. Mechanotransduction of OLL neurons might be carried by a novel Na+ conductance channel, which is insensitive to amiloride. The bona fide mechano-gated Na+-selective degenerin/epithelial Na+ channels, TRP-4, TMC, and Piezo proteins are not involved in this mechanosensation. Interestingly, OLL neurons also mediated cold but not warm responses in a cell-autonomous manner. We further showed that the cold response of OLL neurons is not mediated by the cold receptor TRPA-1 or the temperature-sensitive glutamate receptor GLR-3. Thus, we propose the polymodal functionality of OLL neurons in mechanosensation and cold sensation.

scholarly journals Identification of a PGXPP degron motif in dishevelled and structural basis for its binding to the E3 ligase KLHL12

Open Biology ◽  
2020 ◽  
Vol 10 (6) ◽  
pp. 200041 ◽  
Author(s):  
Zhuoyao Chen ◽  
Gregory A. Wasney ◽  
Sarah Picaud ◽  
Panagis Filippakopoulos ◽  
Masoud Vedadi ◽  
...  
Keyword(s):  
Ubiquitin Ligase ◽  
Molecular Mechanisms ◽  
Hydrophobic Interactions ◽  
E3 Ubiquitin Ligase ◽  
Structural Basis ◽  
E3 Ligases ◽  
Kelch Domain ◽  
Ring E3 Ligases ◽  
Ring E3 ◽  
New Protein

Wnt signalling is dependent on dishevelled proteins (DVL1-3), which assemble an intracellular Wnt signalosome at the plasma membrane. The levels of DVL1-3 are regulated by multiple Cullin-RING E3 ligases that mediate their ubiquitination and degradation. The BTB-Kelch protein KLHL12 was the first E3 ubiquitin ligase to be identified for DVL1-3, but the molecular mechanisms determining its substrate interactions have remained unknown. Here, we mapped the interaction of DVL1-3 to a ‘PGXPP' motif that is conserved in other known partners and substrates of KLHL12, including PLEKHA4, PEF1, SEC31 and DRD4. To determine the binding mechanism, we solved a 2.4 Å crystal structure of the Kelch domain of KLHL12 in complex with a DVL1 peptide that bound with low micromolar affinity. The DVL1 substrate adopted a U-shaped turn conformation that enabled hydrophobic interactions with all six blades of the Kelch domain β-propeller. In cells, the mutation or deletion of this motif reduced the binding and ubiquitination of DVL1 and increased its stability confirming this sequence as a degron motif for KLHL12 recruitment. These results define the molecular mechanisms determining DVL regulation by KLHL12 and establish the KLHL12 Kelch domain as a new protein interaction module for a novel proline-rich motif.

scholarly journals Mapping Interactome Networks of FOSL1 and FOSL2 in Human Th17 Cells

2021 ◽  
Author(s):  
Ankitha Shetty ◽  
Santosh D. Bhosale ◽  
Subhash Kumar Tripathi ◽  
Tanja Buchacher ◽  
Rahul Biradar ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Cell Differentiation ◽  
Th17 Cell ◽  
Th17 Cells ◽  
Molecular Mechanisms ◽  
Rna Binding ◽  
Affinity Purification ◽  
Mass Spectrometry Analysis ◽  
Binding Partners ◽  
Th17 Cell Differentiation

Dysregulated function of Th17 cells has implications in immunodeficiencies and autoimmune disorders. Th17 cell-differentiation is orchestrated by a complex network of transcription factors, including several members of the activator protein (AP-1) family. Among these, FOSL1 and FOSL2 influence the effector responses of Th17 cells. However, the molecular mechanisms underlying their functions are unclear, owing to the poorly characterized protein interaction networks of these factors. Here, we establish the first interactomes of FOSL1 and FOSL2 in human Th17 cells, using affinity purification–mass spectrometry analysis. In addition to the known JUN proteins, we identified several novel binding partners of FOSL1 and FOSL2. Gene ontology analysis found a major fraction of these interactors to be associated with RNA binding activity, which suggests new mechanistic links. Intriguingly, 29 proteins were found to share interactions with FOSL1 and FOSL2, and these included key regulators of Th17-fate. We further validated the binding partners identified in this study by using parallel reaction monitoring targeted mass spectrometry and other methods. Our study provides key insights into the interaction-based signaling mechanisms of FOSL1 and FOSL2 that potentially govern Th17 cell-differentiation and associated pathologies.

scholarly journals Changes in Protein Phosphorylation during Salivary Gland Degeneration in Haemaphysalis longicornis

2020 ◽  
Vol 58 (2) ◽  
pp. 161-171
Author(s):  
Qi Xiao ◽  
Yuhong Hu ◽  
Xiaohong Yang ◽  
Jianna Tang ◽  
Xiaoshuang Wang ◽  
...  
Keyword(s):  
Salivary Gland ◽  
Rna Interference ◽  
Protein Phosphorylation ◽  
Quantitative Study ◽  
Quantitative Proteomics ◽  
Blood Feeding ◽  
Study Data ◽  
Haemaphysalis Longicornis ◽  
Phosphorylated Proteins ◽  
Data Independent Acquisition

The ticks feed large amount of blood from their hosts and transmit pathogens to the victims. The salivary gland plays an important role in the blood feeding. When the female ticks are near engorgement, the salivary gland gradually loses its functions and begins to rapidly degenerate. In this study, data-independent acquisition quantitative proteomics was used to study changes in the phosphorylation modification of proteins during salivary gland degeneration in <i>Haemaphysalis longicornis</i>. In this quantitative study, 400 phosphorylated proteins and 850 phosphorylation modification sites were identified. Trough RNA interference experiments, we found that among the proteins with changes in phosphorylation, apoptosis-promoting Hippo protein played a role in salivary gland degeneration.

scholarly journals Neuronal Proteomic Analysis of the Ubiquitinated Substrates of the Disease-Linked E3 Ligases Parkin and Ube3a

2018 ◽  
Vol 2018 ◽  
pp. 1-14 ◽  
Author(s):  
Aitor Martinez ◽  
Juanma Ramirez ◽  
Nerea Osinalde ◽  
Jesus M. Arizmendi ◽  
Ugo Mayor
Keyword(s):  
Proteasome Inhibitors ◽  
Brain Dysfunction ◽  
Mass Spectrometry Analysis ◽  
E3 Ligases ◽  
Spectrometry Analysis ◽  
Ubiquitinated Proteins ◽  
Neuronal Proteins ◽  
Culture Models ◽  
First Time ◽  
Cell Culture Models

Both Parkin and UBE3A are E3 ubiquitin ligases whose mutations result in severe brain dysfunction. Several of their substrates have been identified using cell culture models in combination with proteasome inhibitors, but not in more physiological settings. We recently developed theUbbiostrategy to isolate ubiquitinated proteins in flies and have now identified by mass spectrometry analysis the neuronal proteins differentially ubiquitinated by those ligases. This is an example of how flies can be used to provide biological material in order to reveal steady state substrates of disease causing genes. Collectively our results provide new leads to the possible physiological functions of the activity of those two disease causing E3 ligases. Particularly, in the case of Parkin the novelty of our data originates from the experimental setup, which is not overtly biased by acute mitochondrial depolarisation. In the case of UBE3A, it is the first time that a nonbiased screen for its neuronal substrates has been reported.

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