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1. chinaXiv:201605.01479 [pdf]

Identification of lipid droplet structure-like/resident proteins in Caenorhabditis elegans

Na, Huimin; Zhang, Peng; Chen, Yong; Zhu, Xiaotong; Liu, Yi; Liu, Yangli; Xie, Kang; Yang, Fuquan; Zhang, Hong; Liu, Pingsheng; Na, Huimin; Zhang, Peng; Zhu, Xiaotong; Liu, Yangli; Xie, Kang; Xu, Ningyi; Mak, Ho Yi; Xu, Ningyi; Mak, Ho Yi; Yu, Yong
Subjects: Biology >> Biophysics >> Biochemistry & Molecular Biology

The lipid droplet (LD) is a cellular organelle that stores neutral lipids in cells and has been linked with metabolic disorders. Caenorhabditis elegans has many characteristics which make it an excellent animal model for studying LDs. However, unlike in mammalian cells, no LD structure-like/resident proteins have been identified in C. elegans, which has limited the utility of this model for the study of lipid storage and metabolism. Herein based on three lines of evidence, we identified that MDT-28 and DHS-3 previously identified in C. elegans LD proteome were two LD structure-like/resident proteins. First, MDT-28 and DHS-3 were found to be the two most abundant LD proteins in the worm. Second, the proteins were specifically localized to LDs and we identified the domains responsible for this targeting in both proteins. Third and most importantly, the depletion of MDT-28 induced LD clustering while DHS-3 deletion reduced triacylglycerol content (TAG). We further characterized the proteins finding that MDT-28 was ubiquitously expressed in the intestine, muscle, hypodermis, and embryos, whereas DHS-3 was expressed mainly in intestinal cells. Together, these two LD structure-like/resident proteins provide a basis for future mechanistic studies into the dynamics and functions of LDs in C. elegans. (C) 2015 Elsevier B.V. All rights reserved.

submitted time 2016-05-12 Hits699Downloads296 Comment 0

2. chinaXiv:201605.01476 [pdf]

Oxidovanadium(IV) sulfate-induced glucose uptake in HepG2 cells through IR/Akt pathway and hydroxyl radicals

Zhao, Qian; Chen, Deliang; Zhang, Fang; Ding, Wenjun; Liu, Pingsheng; Wei, Taotao
Subjects: Biology >> Biophysics >> Biochemistry & Molecular Biology

The insulin-mimetic and anti-diabetic properties of vanadium and related compounds have been well documented both in vitro and in vivo. However, the molecular basis of the link between vanadium and the insulin signaling pathway in diabetes mellitus is not fully described. We investigated the effects of reactive oxygen species (ROS) induced by oxidovanadium(IV) sulfate (VOSO4) on glucose uptake and the insulin signaling pathway in human hepatoma cell line HepG2. Exposure of cells to VOSO4 (5-50 mu M) resulted in an increase in glucose uptake, insulin receptor (IR) and protein kinase B (Akt) phosphorylation and intracellular ROS generation. Using Western blot, we found that catalase and sodium formate, but not superoxide dismutase, prevented the increase of hydroxyl radical (center dot OH) generation and significantly decreased VOSO4-induced IR and Akt phosphorylation. These results suggest that VOSO4-induced center dot OH radical, which is a signaling species, promotes glucose uptake via the IR/Akt signaling pathway. (C) 2015 Elsevier Inc. All rights reserved.

submitted time 2016-05-12 Hits399Downloads238 Comment 0

3. chinaXiv:201605.01452 [pdf]

Proteomic analysis of murine testes lipid droplets

Wang, Weiyi; Wei, Suning; Su, Xueying; Du, Congkuo; Li, Fengjuan; Geng, Bin; Xu, Guoheng; Li, Linghai; Liu, Pingsheng
Subjects: Biology >> Biophysics

Testicular Leydig cells contain abundant cytoplasmic lipid droplets (LDs) as a cholesteryl-ester store for releasing cholesterols as the precursor substrate for testosterone biosynthesis. Here, we identified the protein composition of testicular LDs purified from adult mice by using mass spectrometry and immunodetection. Among 337 proteins identified, 144 were previously detected in LD proteomes; 44 were confirmed by microscopy. Testicular LDs contained multiple Rab GTPases, chaperones, and proteins involved in glucuronidation, ubiquination and transport, many known to modulate LD formation and LD-related cellular functions. In particular, testicular LDs contained many members of both the perilipin family and classical lipase/esterase superfamily assembled predominately in adipocyte LDs. Thus, testicular LDs might be regulated similar to adipocyte LDs. Remarkably, testicular LDs contained a large number of classical enzymes for biosynthesis and metabolism of cholesterol and hormonal steroids, so steroidogenic reactions might occur on testicular LDs or the steroidogenic enzymes and products could be transferred through testicular LDs. These characteristics differ from the LDs in most other types of cells, so testicular LDs could be an active organelle functionally involved in steroidogenesis.

submitted time 2016-05-12 Hits238Downloads159 Comment 0

4. chinaXiv:201605.01413 [pdf]

Lysine Malonylation Is Elevated in Type 2 Diabetic Mouse Models and Enriched in Metabolic Associated Proteins

Du, Yipeng; Zhou, Bo; He, Xiaolong; Wei, Peng; Liu, Pingsheng; Wei, Taotao; Cai, Tanxi; Xue, Peng; Yang, Fuquan; Cai, Tanxi; Xue, Peng; Yang, Fuquan; Li, Tingting; Cai, Tanxi; Zhou, Bo; He, Xiaolong; Wei, Peng
Subjects: Biology >> Biophysics

Protein lysine malonylation, a newly identified protein post-translational modification (PTM), has been proved to be evolutionarily conserved and is present in both eukaryotic and prokaryotic cells. However, its potential roles associated with human diseases remain largely unknown. In the present study, we observed an elevated lysine malonylation in a screening of seven lysine acylations in liver tissues of db/db mice, which is a typical model of type 2 diabetes. We also detected an elevated lysine malonylation in ob/ob mice, which is another model of type 2 diabetes. We then performed affinity enrichment coupled with proteomic analysis on liver tissues of both wild-type (wt) and db/db mice and identified a total of 573 malonylated lysine sites from 268 proteins. There were more malonylated lysine sites and proteins in db/db than in wt mice. Five proteins with elevated malonylation were verified by immunoprecipitation coupled with Western blot analysis. Bioinformatic analysis of the proteomic results revealed the enrichment of malonylated proteins in metabolic pathways, especially those involved in glucose and fatty acid metabolism. In addition, the biological role of lysine malonylation was validated in an enzyme of the glycolysis pathway. Together, our findings support a potential role of protein lysine malonylation in type 2 diabetes with possible implications for its therapy in the future.

submitted time 2016-05-12 Hits580Downloads357 Comment 0

5. chinaXiv:201605.01335 [pdf]

Dynamics of the Lipid Droplet Proteome of the Oleaginous Yeast Rhodosporidium toruloides

Zhu, Zhiwei; Gong, Zhiwei; Zhang, Sufang; Lin, Xinping; Shen, Hongwei; Zou, Hanfa; Zhao, Zongbao K.; Ding, Yunfeng; Yang, Li; Zhang, Congyan; Xie, Zhensheng; Yang, Fuquan; Zhao, Xudong; Liu, Pingsheng; Zhang, Sufang; Shen, Hongwei; Zhao, Zongbao K.
Subjects: Biology >> Biophysics

Lipid droplets (LDs) are ubiquitous organelles that serve as a neutral lipid reservoir and a hub for lipid metabolism. Manipulating LD formation, evolution, and mobilization in oleaginous species may lead to the production of fatty acid-derived biofuels and chemicals. However, key factors regulating LD dynamics remain poorly characterized. Here we purified the LDs and identified LD-associated proteins from cells of the lipid-producing yeast Rhodosporidium toruloides cultured under nutrient-rich, nitrogen-limited, and phosphorus-limited conditions. The LD proteome consisted of 226 proteins, many of which are involved in lipid metabolism and LD formation and evolution. Further analysis of our previous comparative transcriptome and proteome data sets indicated that the transcription level of 85 genes and protein abundance of 77 proteins changed under nutrient-limited conditions. Such changes were highly relevant to lipid accumulation and partially confirmed by reverse transcription-quantitative PCR. We demonstrated that the major LD structure protein Ldp1 is an LD marker protein being upregulated in lipid-rich cells. When overexpressed in Saccharomyces cerevisiae, Ldp1 localized on the LD surface and facilitated giant LD formation, suggesting that Ldp1 plays an important role in controlling LD dynamics. Our results significantly advance the understanding of the molecular basis of lipid overproduction and storage in oleaginous yeasts and will be valuable for the development of superior lipid producers.

submitted time 2016-05-11 Hits426Downloads235 Comment 0

6. chinaXiv:201605.01235 [pdf]

Lipid droplet remodeling and interaction with mitochondria in mouse brown adipose tissue during cold treatment

Yu, Jinhai; Zhang, Shuyan; Cui, Liujuan; Na, Huimin; Zhu, Xiaotong; Yang, Fuquan; Liu, Pingsheng; Yu, Jinhai; Na, Huimin; Zhu, Xiaotong; Wang, Weiyi; Xu, Guoheng; Li, Linghai; Christian, Mark; Cui, Liujuan
Subjects: Biology >> Biophysics >> Biochemistry & Molecular Biology

Brown adipose tissue (BAT) maintains animal body temperature by non-shivering thermogenesis, which is through uncoupling protein 1 (UCP1) that uncouples oxidative phosphorylation and utilizes beta-oxidation of fatty acids released from triacylglycerol (TAG) in lipid droplets (LDs). Increasing BAT activity and "browning" other tissues such as white adipose tissue (WAT) can enhance the expenditure of excess stored energy, and in turn reduce prevalence of metabolic diseases. Although many studies have characterized the biology of BAT and brown adipocytes, BAT LDs especially their activation induced by cold exposure remain to be explored. We have isolated LDs from mouse interscapular BAT and characterized the full proteome using mass spectrometry. Both morphological and biochemical experiments showed that the LDs could tightly associate with mitochondria. Under cold treatment mouse BAT started expressing LD structure protein PLIN-2/ADRP and increased expression of PLIN1 Both hormone sensitive lipase (HSL) and adipose TAG lipase (ATGL) were increased in LDs. In addition, isolated BAT LDs showed increased levels of the mitochondrial protein UCP1, and prolonged cold exposure could stimulate BAT mitochondrial cristae biogenesis. These changes were in agreement with the data from transcriptional analysis. Our results provide the BAT LD proteome for the first time and show that BAT LDs facilitate heat production by coupling increasing TAG hydrolysis through recruitment of ATGL and HSL to the organelle and expression of another LD resident protein PLIN2/ADRP, as well as by tightly associating with activated mitochondria. These findings will benefit the study of BAT activation and the interaction between LDs and mitochondria. (C) 2015 Elsevier B.V. All rights reserved.

submitted time 2016-05-11 Hits448Downloads259 Comment 0

7. chinaXiv:201605.00723 [pdf]

Comparative proteomics reveals abnormal binding of ATGL and dysferlin on lipid droplets from pressure overload-induced dysfunctional rat hearts

Li, Linghai; Zhang, Huina; Li, Linghai; Zhang, Huina; Wang, Jifeng; Zhang, Shuyan; Xu, Shimeng; Shu, Qingbo; Yang, Fuquan; Liu, Pingsheng; Wang, Weiyi; Hong, Yun; Zheng, Min; Xu, Shimeng; Shu, Qingbo; Li, Juanfen; Qian, Zongjie;
Subjects: Biology >> Biophysics

Excessive retention of neutral lipids in cardiac lipid droplets (LDs) is a common observation in cardiomyopathy. Thus, the systematic investigation of the cardiac LD proteome will help to dissect the underlying mechanisms linking cardiac steatosis and myocardial dysfunction. Here, after isolation of LDs from normal and dysfunctional Sprague-Dawley rat hearts, we identified 752 heart-associated LD proteins using iTRAQ quantitative proteomic method, including 451 proteins previously unreported on LDs. The most noteworthy finding was the identification of the membrane resealing protein, dysferlin. An analysis of dysferlin truncation mutants indicated that its C2 domain was responsible for its LD localization. Quantitative proteomic results further determined that 27 proteins were increased and 16 proteins were decreased in LDs from post pressure overload-induced dysfunctional hearts, compared with normal hearts. Notably, adipose triacylglycerol lipase (ATGL) was dramatically decreased and dysferlin was substantially increased on dysfunctional cardiac LDs. This study for the first time reveals the dataset of the heart LD proteome in healthy tissue and the variation of it under cardiac dysfunction. These findings highlight an association between the altered LD protein localization of dysferlin and ATGL and myocardial dysfunction.

submitted time 2016-05-05 Hits412Downloads233 Comment 0

8. chinaXiv:201605.00707 [pdf]

Comparative Proteomic Study of Fatty Acid-treated Myoblasts Reveals Role of Cox-2 in Palmitate-induced Insulin Resistance

Chen, Xiulan; Wei, Shasha; Yang, Fuquan; Chen, Xiulan; Wei, Shasha; Yang, Fuquan; Xu, Shimeng; Deng, Yaqin; Li, Yiran; Liu, Pingsheng; Xu, Shimeng; Wei, Shasha; Deng, Yaqin; Li, Yiran;
Subjects: Biology >> Biophysics

Accumulated studies demonstrate that saturated fatty acids (FAs) such as palmitic acid (PA) inhibit insulin signaling in skeletal muscle cells and monounsaturated fatty acids such as oleic acid (OA) reverse the effect of PA on insulin signaling. The detailed molecular mechanism of these opposite effects remains elusive. Here we provide a comparative proteomic study of skeletal myoblast cell line C2C12 that were untreated or treated with PA, and PA plus OA. A total of 3437 proteins were quantified using SILAC in this study and 29 proteins fall into the pattern that OA reverses PA effect. Expression of some these proteins were verified using qRT-PCR and Western blot. The most significant change was cyclooxygenase-2 (Cox-2). In addition to whole cell comparative proteomic study, we also compared lipid droplet (LD)-associated proteins and identified that Cox-2 was one of three major altered proteins under the FA treatment. This finding was then confirmed using immunofluorescence. Finally, Cox-2 selective inhibitor, celecoxib protected cells from PA-reduced insulin signaling Akt phosphorylation. Together, these results not only provide a dataset of protein expression change in FA treatment but also suggest that Cox-2 and lipid droplets (LDs) are potential players in PA- and OA-mediated cellular processes.

submitted time 2016-05-05 Hits354Downloads218 Comment 0

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