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Search indexed NASA NTRS and DOE OSTI research on propulsion, heat transfer, battery materials and energy systems. Follow report and document links to the original sources.

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At least 1,405 records · Page 78
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DOE OSTI2016

The up-regulation of long non-coding RNA CCAT2 indicates a poor prognosis for prostate cancer and promotes metastasis by affecting epithelial-mesenchymal transition

Recently, long noncoding RNAs (lncRNAs) have been shown to have critical regulatory roles in human cancer biology. LncRNA CCAT2 is a novel identified lncRNA that was previously reported to be up-regulated in different cancers, however, its role in prostate cancer remains unclear. The aim of this study was to investigate the expression and role of lncRNA CCAT2 in prostate cancer. The expression levels of lncRNA CCAT2 in PCa tissues and cell lines (DU145 and 22RV1) were evaluated by quantitative real-time PCR (qRT-PCR), and its association with prognosis of patients was analyzed by statistical analysis. Furthermore, the effect of CCAT2 on proliferation, migration, and invasion was studied in PCa cells. We found that the expression level of CCAT2 was higher in PCa tissues and cells compared to adjacent non-tumor tissues and normal prostate stromal immortalized cells WPMY-1. Kaplan-Meier survival analysis revealed that patients with high CCAT2 expression level had poorer overall survival and progression-free survival than those with low CCAT2 expression. Furthermore, multivariate analysis showed that the status of CCAT2 expression was an independent prognostic indicator for this disease. We also found that knockdown of CCAT2 could inhibit cell growth, migration, and invasion in vitro. In addition, knockdown of CCAT2 stimulated epithelial-mesenchymal transition (EMT) through abrogating N-cadherin, vimentin expression and intensifing the expression levels of E-cadherin. In conclusion, our data suggested that lncRNA CCAT2 was a novel molecule involved in PCa progression, which provided a potential prognostic biomarker and therapeutic target for new therapies in patients with prostate cancer. - Highlights: • LncRNA CCAT2 is identified with overexpression in prostate cancer tissues and cell lines. • High expression of lncRNA CCAT2 can be an independent prognostic factor for overall survival and progression-free survival. • Knockdown of lncRNA CCAT2 can inhibit cell growth, migration, and invasion in vitro study. • Knockdown of lncRNA CCAT2 stimulates metastasis of prostate cancer cell through regulating EMT-associated gene expression including N-cadherin, Vimentin and E-cadherin.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2016

The E3 ligase UBR5 regulates gastric cancer cell growth by destabilizing the tumor suppressor GKN1

Gastric cancer is the most common digestive malignant tumor worldwide and the underlying mechanisms are not fully understood. The E3 ligase UBR5 (also known as EDD1) is essentially involved in diverse types of cancer. Here we aimed to study the functions of UBR5 in human gastric cancer. We first analyzed the mRNA and protein levels of UBR5 in human gastric cancer tissues and the results showed that UBR5 was markedly increased in gastric cancer tissues compared with normal gastric mucosa or matched non-cancer gastric tissues. The relationship between UBR5 and survival of gastric cancer patients was analyzed and we found that high UBR5 expression was associated with poor overall and disease-free survival. We further tried to investigate the effects of UBR5 on gastric cancer cell growth in vitro and in vivo. Therefore, we knocked down UBR5 with lentivirus-mediated shRNA and found that UBR5 knockdown repressed in vitro proliferation and colony formation of gastric cancer cells AGS, MG803 and MNK1. In vivo xenograft experiment also demonstrated that UBR5 knockdown inhibited AGS growth. Finally, we explored the mechanism by which UBR5 contributed to the growth of gastric cancer cells. We found that UBR5 bound the tumor suppressor gastrokine 1 (GKN1) and increased its ubiquitination to reduce the protein stability of GKN1. GKN1 knockdown with lentivirus-mediated shRNA increased the in vitro colony formation and in vivo growth of AGS cells, and UBR5 knockdown was unable to affect the colony formation and in vivo growth of AGS cells when GKN1 was knocked down, indicating that GKN1 contributed to the effects of UBR5 in human gastric cancer cells. Taken together, UBR5 plays an essential role in gastric cancer and may be a potential diagnosis and treatment target for gastric cancer. - Highlights: • UBR5 expression is up-regulated in human gastric cancer. • UBR5 overexpression predicts poor survival. • UBR5 regulates gastric cancer growth in vitro and in vivo. • UBR5 reduces the protein stability of GKN1.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2016

PPARγ is regulated by miR-27b-3p negatively and plays an important role in porcine oocyte maturation

To elucidate the key miRNAs and the signalling pathways that are involved in porcine oocyte maturation, we performed a deep sequencing analysis of the miRNAs of pig germinal vesicle (GV) oocytes and metaphase II (MII) oocytes. Seven differentially expressed (DE) miRNAs were identified and the expression levels of miR-21 and miR-27b-3p were further confirmed by QPCR analysis. The target genes of 7 DE miRNAs were predicted and subjected to pathway analysis. Interestingly, fatty acid metabolism and fatty acid biosynthesis were the top two significantly enriched molecular functions during oocyte maturation. Heat map, which was built with 7 DE miRNAs and the enriched the molecular functions, revealed that miR-21, miR-27b-3p, miR-10a-5p and miR-10b-5p were involved in fatty acid metabolism. In particular, the regulatory role of miR-27b-3p on peroxisome proliferator-activated receptor-γ (PPARγ) was confirmed by their inversed expression patterns in GV and MII oocytes and luciferase report assays. In addition, we observed that PPARγ agonist (rosiglitazone) treatment significantly enhanced porcine oocyte maturation rate and early embryo developmental competent. Taken together, our results demonstrated that miR-27b and its target, PPARγ, play the vital roles in pig oocyte maturation through regulating the fatty acid metabolism. These data increased our understanding of the regulatory gene networks in porcine oocyte maturation and development. - Highlights: • Profiling of miRNAs in porcine GV and MII oocytes. • Fatty acid metabolism pathway is over-represented during porcine oogenesis. • PPARγ is regulated by miR-27b-3p negatively. • PPARγ promotes the rate of porcine oocyte maturation and blastocyst in vitro.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2017

Up-regulation of VEZT by small activating RNA inhibits the proliferation, invasion and migration of gastric cancer cells

Objective: To identify an effective saRNA sequence that can specifically up-regulate VEZT expression and to determine the influence of saRNA had on gastric cancer cell growth, proliferation, invasion and migration. Methods: Three various saRNAs, that target the VEZT gene promoter at different locations relative to the transcription start site were synthesized. A dsControl saRNA was synthesized as a negative control, and a specific shRNA was synthesized to knockdown VEZT and eliminate any off-target effects of the saRNA. Both SGC-7901 and M-28 cells were either transfected with the different saRNAs, or treated with Lipofectamine2000 alone. To determine the most effective saRNA, real-time PCR and Western blot were used to determine the VEZT mRNA and protein content, respectively, of each treatment group. After selection, both cell lines were treated with the chosen saRNA, dsControl or Lipofectamine2000. The saRNA treated cells were divided into two groups: the first group was used immediately in the experiments, and the second group was transfected with shRNA by using RNAi-Mate. The proliferation of cells transfected with saRNA, or saRNA and shRNA, as well as the other control cells, was detected by CCK-8. The invasive and migratory abilities were determined using the transwell chamber assay. Results: We identified the most effective saRNA via real-time PCR and Western blot. The selected saRNA inhibited the growth, invasion and migration of GC cells by specially reactivating VEZT. The real-time PCR and Western blot results showed that treatment with saRNA caused a significant up-regulation of VEZT, and an obvious decrease in the proliferative, invasive and migratory abilities; compared with the control groups (P < 0.01); furthermore, there were no significant differences among the control groups (P > 0.05). This phenomenon provides a theoretical basis for saRNA design and gene therapy for gastric cancer. - Highlights: • saRNA is a brand-new type of small non-coding RNA. It provides a brand-new therapeutic tool for gastric cancer. • In this manuscript, we designed the new saRNA that was effective for VEZT for the first time. • VEZT is a new TSG, and the reactivation through saRNA is never reported before. • It's the first time to report the RNAa in gastric cancer.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2017

miR-214 down-regulates ARL2 and suppresses growth and invasion of cervical cancer cells

Increasing evidence has shown that miRNAs are implicated in carcinogenesis and can function as oncogenes or tumor suppressor genes in human cancers. In this study, we confirmed that miR-214 is frequently down-regulated in cervical cancer compared with normal cervical tissues. Ectopic expression of miR-214 suppressed proliferation, migration and invasion of HeLa and C33A cervical cancer cells. Bioinformatics analysis revealed that ADP ribosylation factor like 2 (ARL2) was a potential target of miR-214 and was remarkably up-regulated in cervical cancer. Knockdown of ARL2 markedly inhibited cervical cancer cell proliferation, migration and invasion, similarly to over-expression of miR-214, indicating that ARL2 may function as an oncogene in cervical cancer. In conclusion, our study revealed that miR-214 acts as a tumor suppressor via inhibiting proliferation, migration and invasion of cervical cancer cells through targeting ARL2, and that both miR-214 and ARL2 may serve as prognostic or therapeutic targets for cervical cancer. - Highlights: • miR-214 targets ARL2. • ARL2 maybe an oncogene in cervical cancer. • ARL2 rescues miR-214.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2017

TEAD4-YAP interaction regulates tumoral growth by controlling cell-cycle arrest at the G1 phase

TEA domain transcription factor 4 (TEAD4), which has critical functions in the process of embryonic development, is expressed in various cancers. However, the important role of TEAD4 in human oral squamous cell carcinomas (OSCCs) remain unclear. Here we investigated the TEAD4 expression level and the functional mechanism in OSCC using quantitative reverse transcriptase-polymerase chain reaction, Western blot analysis, and immunohistochemistry. Furthermore, TEAD4 knockdown model was used to evaluate cellular proliferation, cell-cycle analysis, and the interaction between TEAD4 and Yes-associated protein (YAP) which was reported to be a transcription coactivator of cellular proliferation. In the current study, we found that TEAD4 expression increased significantly in vitro and in vivo and correlated with tumoral size in OSCC patients. TEAD4 knockdown OSCC cells showed decreased cellular proliferation resulting from cell-cycle arrest in the G1 phase by down-regulation of cyclins, cyclin-dependent kinases (CDKs), and up-regulation of CDK inhibitors. We also found that the TEAD4-YAP complex in the nuclei may be related closely to transcriptions of G1 arrest-related genes. Taken together, we concluded that TEAD4 might play an important role in tumoral growth and have potential to be a therapeutic target in OSCCs. - Highlights: • TEAD4 contributes to tumor progression in OSCCs. • TEAD4 knockdown results in cell-cycle arrest at the G1phase in OSCC cells. • In TEAD4 knockdown cells, the amount of YAP in the nucleus decreases. • Activation of the TEAD4-YAP complex is an important factor in OSCC tumor growth. • TEAD4 might be a critical biomarker and a therapeutic target for OSCCs.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2017

Characterization of the rax1 gene encoding a putative regulator of G protein signaling in Aspergillus fumigatus

The filamentous fungus Aspergillus fumigatus is the major cause of life threatening invasive aspergillosis, and its small hydrophobic asexual spores (conidia) are the major infection agent. To better understand biology of A. fumigatus, we have characterized the rax1 gene encoding a putative regulator of G protein signaling (RGS). The deletion (Δ) of rax1 results in restricted colony growth and highly reduced number of conidia in A. fumigatus. Transcript levels of the three central activators of asexual development abaA, brlA, and wetA are significantly reduced in the Δrax1 mutant. However, the Δrax1 conidia, but not vegetative cells, are specifically resistant against H{sub 2}O{sub 2} stress. The Δrax1 conidia accumulate higher mRNA levels of sakA encoding a key MAP kinase for stress response. Moreover, the Δrax1 conidia contain over five-fold amount of trehalose, an osmolyte and protein/membrane protectant. Transmission electron microscopy analyses indicate that the Δrax1 conidia have the thicker melanized-outermost cell wall layer compared to those of wild-type. In summary, Rax1 positively controls growth and development, and modulates intracellular trehalose amount, cell wall melanin levels in conidia, and spore resistance to H{sub 2}O{sub 2}. - Highlights: • Rax1 positively regulates vegetative growth and asexual development. • Deletion of rax1 resulted in spore resistance against H{sub 2}O{sub 2}. • Rax1 modulates trehalose amount and cell wall melanin levels in conidia.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2017

ER-α36 mediates estrogen-stimulated MAPK/ERK activation and regulates migration, invasion, proliferation in cervical cancer cells

Objective: Estrogen receptor alpha 36 (ER-α36), a truncated variant of ER-α, is different from other nuclear receptors of the ER-α family. Previous findings indicate that ER-α36 might be involved in cell growth, proliferation, and differentiation in carcinomas and primarily mediates non-genomic estrogen signaling. However, studies on ER-α36 and cervical cancer are rare. This study aimed to detect the expression of ER-α36 in cervical cancer; the role of ER-α36 in 17-β-estradiol (E2)-induced invasion, migration and proliferation of cervical cancer; and their probable molecular mechanisms. Methods: Immunohistochemistry and immunofluorescence were used to determine the location of ER-α36 in cervical cancer tissues and cervical cell lines. CaSki and HeLa cell lines were transfected with lentiviruses to establish stable cell lines with knockdown and overexpression of ER-α36. Wound healing assay, transwell invasion assay, and EdU incorporation proliferation assay were performed to evaluate the migration, invasion, and proliferation ability. The phosphorylation levels of mitogen-activated protein kinases/extracellular signal-regulated kinase (MAPK/ERK) signaling molecules were examined with western blot analysis. Results: ER-α36 expression was detected in both cervical cell lines and cervical cancer tissues. Downregulation of ER-α36 significantly inhibited cell invasion, migration, and proliferation. Moreover, upregulation of ER-α36 increased the invasion, migration, and proliferation ability of CaSki and HeLa cell lines. ER-α36 mediates estrogen-stimulated MAPK/ERK activation. Conclusion: ER-α36 is localized on the plasma membrane and cytoplasm in both cervical cancer tissues and cell lines. ER-α36 mediates estrogen-stimulated MAPK/ERK activation and regulates migration, invasion, proliferation in cervical cancer cells. - Highlights: • ER-α36 is expressed on both cervical cell lines and cervical cancer tissues. • ER-α36 mediates estrogen-stimulated cell invasion, migration and proliferation of cervical cancer cell lines. • ER-α36 mediates estrogen-stimulated MAPK/ERK activation in cervical cancer cell lines.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2017

Increased expression of microRNA-31-5p inhibits cell proliferation, migration, and invasion via regulating Sp1 transcription factor in HepG2 hepatocellular carcinoma cell line

Accumulating evidence has suggested that microRNA-31-5p (miR-31-5p) is dysfunctional in hepatocellular carcinoma (HCC). However, the molecular mechanism of HCC remains unclear. In this study, we investigated the role of miR-31-5p in tumor formation and development of HCC. The expression of miR-31-5p was detected in HCC tissues, corresponding adjacent tissues, normal liver tissues, and HCC cell lines. miR-31-5p mimics and an inhibitor were transfected into HepG2 cells to assess the effects of miR-31-5p on cell proliferation, apoptosis, cell cycle, migration, and invasion assays. Western blotting was used to detect the expression of Sp1 transcription factor (SP1), cyclin D1, and survivin in transfected HCC cells and control cells. The expression of miR-31-5p was significantly decreased in HCC cells and HCC tissues. Overexpression of miR-31-5p inhibited HCC cell growth, migration, and invasion. Overexpression of miR-31-5p reduced the expression of SP1 and cyclin D1, and knockdown of SP1 decreased cyclin D1 expression. The dual luciferase assay showed that miR-31-5p directly targeted SP1 in HepG2. Together, the results suggested that miR-31-5p acted as a tumor suppressor to regulate SP1, and that miR-31-5p could be used as a therapeutic target for the treatment of HCC. - Highlights: • The expression of miR-31-5p was significantly up-regulated in HCC tissues. • Increased expression of miR-31-5p inhibits cell proliferation, migration, and invasion. • Sp1 transcription factor is the target gene for miR-31-5p.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2017

Astrocytes regulate the expression of Sp1R3 on oligodendrocyte progenitor cells through Cx47 and promote their proliferation

Many degenerative diseases of the central nervous system are associated with demyelination. Oligodendrocyte progenitor cells (OPCs) are potential stem cells that can differentiate into various cell types, including oligodendrocytes (OLs). Promoting the proliferation and differentiation of OPCs into mature OLs that can myelinate axons is the key to stimulate remyelination. Here, we report that astrocytes (ASTs) increase the number of sphingosine-1-phosphate receptors 3 (S1pR3) on OPCs and promote OPCs proliferation through a direct contact via connexin 47 (Cx47). Our results demonstrate that ASTs can regulate the proliferation of OPCs through Cx47-mediated induction of S1pR3 expression on OPCs. Cx47/S1pR3 remarkably increases the number of OPCs and promotes cell transition from the G1 to the S phase. Furthermore, inhibiting either Cx47 or S1pR3 decreases OPC proliferation. In summary, ASTs regulate the expression of S1pR3 in OPCs via Cx47, which could be a valuable approach for promoting OPC proliferation. This strategy may therefore represent a potential treatment for neurological diseases caused by OLs death and demyelination. - Highlights: • Astrocytes through direct contact (connexin 47), can increase the number of OPCs surface proliferation receptors (S1pR3). • The increased number of OPCs on surface of proliferation receptors (S1pR3), promoted proliferation of OPCs. • Blocking S1pR3 could decrease the proliferation of OPCs.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2017

LIGHT is increased in patients with coronary disease and regulates inflammatory response and lipid metabolism in oxLDL-induced THP-1 macrophages

Inflammation is critical for the progression of hyperlipidemia. Although the exact mechanism through which inflammation affects hyperlipidemia is not very clear, evidence suggests that the tumor necrosis factor superfamily member 14 (TNFSF14/LIGHT)LIGHT might regulate lipid metabolism. In this study we investigated the expression of LIGHT in patients with different stages of coronary disease. The expression of lipid metabolism-related enzymes and inflammation-related proteins were further explored in oxidized low-density lipoproteins (oxLDL)-induced THP-1 macrophages. We found that LIGHT is highly expressed and companied with severe inflammations in patients with coronary disease. LIGHT significantly enhanced inflammation response in oxLDL-induced THP-1 macrophages. We further demonstrated that LIGHT markedly decreased the levels of lipolytic genes and increased the expressions of lipogenic genes in oxLDL-induced THP-1 macrophages. In addition, our results showed that LIGHT exerts its pro-inflammatory and pro-lipogenesis roles through activating nuclear factor-kappa B (NF-κB) signaling pathway. Taken together our study has demonstrated that LIGHT NF-κB-dependently exacerbates inflammation response and promotes lipid accumulation, and provided a new potential target for treatment of hyperlipidemia-related disease. - Highlights: • Patients with atherosclerosis display high levels of LIGHT expression. • LIGHT promotes inflammation and lipid accumulation in THP-1 macrophages. • LIGHT regulates lipid metabolism via NF-κB signaling.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2017

Site-specific regulation of oral mucosa-recruiting CD8{sup +} T cells in a mouse contact allergy model

Contact allergy is a T cell-mediated, delayed-type hypersensitivity generated by contact exposure of an allergen to the skin and mucosal surface. The clinical manifestations of allergic responses between the skin and oral mucosa vary and the differences in immunopathology have not been clarified. We generated hapten-induced contact hypersensitivity (CH) of the buccal mucosa (BM) in parallel studies with ear skin (ES) CH, and observed several characteristic findings of BM CH. The BM challenge induced more rapid and more severe inflammation than the ES challenge, with abundant granulocyte and CD8{sup +} T cell infiltration. However, these inflammatory responses diminished quickly. Recruiting CD8{sup +} T cells in the BM had higher ratios of CD62L{sup −}CD44{sup low-hi} memory-type cells, and showed impaired IFN-γ, greater PD-1, and comparable Ki-67 expression, suggesting that the recruiting-proliferating CD8{sup +} T cells were unable to differentiate into effector T cells and converted into exhausted T cells at the local site. This finding may explain the rapid recovery of the BM from severe inflammation. Preferentially greater expression of PD-1 ligand (B7-H1), was observed in the BM epithelium under the peak inflammation, and the absence of B7-H1 further accelerated CH responses, suggesting the occurrence of PD-1:B7-H1-mediated immune regulation at the local site. Our results may facilitate the understanding of the unique features of contact allergies in the oral mucosa, and guide the development of new strategies for control of contact allergy. - Highlights: • Contact allergy in the oral mucosa shows severe responses, but quick recovery. • Oral mucosa-infiltrating CD8{sup +} T cells have unique activatory and regulatory status. • PD-1:B7-H1-mediated pathway is involved in the site-specific regulation.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2017

Snail transcription factor NLS and importin β1 regulate the subcellular localization of Cathepsin L and Cux1

Several recent studies have highlighted an additional unexpected localization and site of action for Cathepsin L (Cat L) protease within the nucleus in breast, colon and prostate cancer, however, its role in the nucleus was unclear. It was proposed to mediate proteolytic processing of the transcription factor CCAAT-displacement protein/cut homeobox transcription factor (Cux1) from the full-length p200 isoform to generate the p110 and p90 isoforms, of which the p110 isoform was shown to act as a cell cycle regulator to accelerate entry into the S phase. The p110 isoform has also been shown to bind to the promoter regions of Snail and E-cadherin to activate Snail and inactivate E-cadherin transcription, thus promoting epithelial mesenchymal transition (EMT). Mechanistic studies on what drives Cat L nuclear localization have not been reported. Our hypothesis is that Snail shuttles into the nucleus with Cat L through binding to importin-β. Snail knockdown with siRNA in MDA-MB-468 breast cancer cells led to nuclear to cytoplasmic shuttling of Cat L and decreased levels of Cux1, while overexpression of Snail in MCF-7 breast cancer cells or HEK-293 human embryonic kidney cells led to increased nuclear expression of both Cat L and Cux1. Additionally, transient transfection of Snail NLS mutants not only abrogated Snail nuclear localization but also nuclear localization of Cat L and Cux1. Interestingly, importin β1 knockdown with siRNA decreased Snail and Cux1 levels, as well as nuclear localization of Cat L. Therefore, we show for the first time that the nuclear localization of Cat L and its substrate Cux1can be positively regulated by Snail NLS and importin β1, suggesting that Snail, Cat L and Cux1 all utilize importin β1 for nuclear import. - Highlights: • Snail expression is associated with increased nuclear expression of Cathepsin L (Cat L) and Cux1. • Snail NLS is required for nuclear localization of Cat L and Cux1. • Importin β1 is required for Snail, Cat L and Cux1 expression/nuclear localization.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2017

Androgen-responsive circular RNA circSMARCA5 is up-regulated and promotes cell proliferation in prostate cancer

Prostate cancer (PCa) is one of the most commonly diagnosed cancers in males worldwide. Circular RNA (circRNA) is a unique class of RNA transcribed by RNA polymerase II characterized by jointing 3′ and 5′ ends together via exon or intron circularization. However, the molecular functions of circRNAs in prostate cancer have rarely been explored. In present study, we found circ-SMARCA5 was up-regulated in prostate cancer samples compared to match normal tissues. We also observed circ-SMARCA5 expression was significantly induced after DHT treatment. Functional experiments showed circ-SMARCA5 acted as an oncogene in prostate cancer by promoting cell cycle and inhibiting cell apoptosis. We thought this study provided useful information for exploring circRNAs as potential therapeutic and prognostic targets for prostate cancer. - Highlights: • To our knowledge, this is the first study to explore the molecular functions of circRNA in prostate cancer. • Our study showed that circ-SMARCA5 promoted cell proliferation and inhibited cell apoptosis in prostate cancer. • circ-SMARCA5 was for the first time reported as an androgen-responsive circRNA. • circ-SMARCA5 was for the first time reported to be up-regulated in prostate cancer.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2017

MeWRKY20 and its interacting and activating autophagy-related protein 8 (MeATG8) regulate plant disease resistance in cassava

Highlights: • A systematic genome-wide of MeATGs is performed. • MeWRKY20 physically interacts with MeATG8a/8f/8h. • MeWRKY20 activates the transcription of MeATG8a. • MeATG8s and MeWRKY20 regulate defense response in cassava. As a highly conserved mechanism, autophagy is responsible for the transport of cytoplasmic constituents in the vacuoles or lysosomes. Moreover, autophagy is essential for plant development and various stress responses. In this study, 34 MeATGs were systematically identified in cassava, and their transcripts were commonly regulated by Xanthomonas axonopodis pv manihotis (Xam). Through transient expression in Nicotiana benthamiana, the subcellular locations of 4 MeATG8s were revealed. Notably, MeWRKY20 was identified as physical interacting protein of MeATG8a/8f/8h and upstream transcriptional activator of MeATG8a. Through virus-induced gene silencing (VIGS) in cassava, we found that MeATG8-silenced and MeWRKY20-silenced plants resulted in disease sensitive, with less callose depositions and lower autophagic activity. This study may facilitate our understanding of the upstream MeWRKY20 and underlying target as well as interacting proteins of MeATG8s in immune response. Taken together, MeWRKY20 and MeATG8a/8f/8h are essential for disease resistance against bacterial blight by forming various transcriptional modules and interacting complex in cassava.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2017

Dihydroceramide is a key metabolite that regulates autophagy and promotes fibrosis in hepatic steatosis model

Highlights: • FFA uptake increases sphingolipids in liver cells. • Dihydroceramide product correlates with autophagosome accumulation. • Dihydroceramide impairs autophagic flux and increases lipid droplets. • Rapamycin recuperates fibrosis by recovering autophagic flux by increased dihydroceramide. Non-alcoholic fatty liver disease (NAFLD) is an increasingly common chronic liver disease worldwide. Sphingolipids are a family of lipids that play essential roles as critical regulators in metabolic disorders. Some sphingolipids are known key factors in metabolic dysfunction. However, the precise effect of dihydroceramide on NAFLD remains unknown. Here, we report how dihydroceramide in autophagosome accumulation activates fibrogenesis in human liver Chang cells treated with free fatty acids (FFA). According to LC/MS lipid profiling, FFA increased the levels of sphingolipids and triacylglycerol (TG). To demonstrate the potential role of dihydroceramide metabolism in autophagy, several sphingolipid synthesis inhibitors were used. Increased dihydroceramide led to impairment of autophagic flux, resulting in increased TG storage in lipid droplets (LD) and upregulated expression of fibrosis markers. Hepatic stellate cells (HSCs, LX-2 cells) were co-cultured with Chang cells to assess the potential fibrogenic response to dihydroceramide, Treatment with rapamycin recovered autophagic flux in Chang cells and fibrogenesis in the co-culture system. Our results identified a critical function of dihydroceramide metabolism in autophagy. It could play an important role in the progression of NAFLD associated with lipid over-accumulation. Therefore, preventing autophagic flux by regulating dihydroceramide could be a potential strategic approach for providing therapy for NAFLD.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

KLF2 attenuates bleomycin-induced pulmonary fibrosis and inflammation with regulation of AP-1

Highlights: • KLF2 is downregulated in BLM-treated rats. • KLF2 alleviates BLM-induced lung fibrosis. • KLF2 suppresses the production of pro-inflammatory cytokines. • KLF2 prevents TGF-β1 induced lung fibrosis in vitro. • Effects of KLF2 on AP-1. Pulmonary fibrosis (PF) is a chronic, fibrosing interstitial pneumonia and devastating disease. Here we investigated the potential roles of Kruppel-like factor 2 (KLF2) on pulmonary fibrosis and inflammation response. A mouse model of pulmonary fibrosis was established by intratracheal injection of bleomycin (BLM). The mRNA and protein levels of KLF2 were assayed by RT-PCR and Western blotting respectively. The extent of lung fibrosis was determined using hematoxylin and eosin (HE) staining and Masson's trichrome staining, and the hydroxyproline content was quantified. RT-PCR was used to evaluate the mRNA expression of collagen type 1a1 (col1a1), col3a1, α-SMA, TNF-α, IL-1β and IL-6. The concentrations of TNF-α, IL-1β, and IL-6 in bronchoalveolar lavage fluid (BALF) and lung tissue were examined by ELISA. Also, the effects of KLF2 on activator protein-1 (AP-1) were evaluated by measuring the c-Jun and c-Fos protein levels. We found that KLF2 was remarkably downregulated in BLM-treated rats, both in mRNA and protein levels. Additionally, overexpression of KLF2 attenuated the destruction of the alveolar space and pulmonary interstitial collagen hyperplasia, and deposition reduced the expression of col1a1, col3a1, and α-SMA, and blocked the production of TNF-α, IL-1β, and IL-6 in BALF and lung tissue in vivo. Moreover, adenoviral transduction of KLF2 inhibited TGF-β1-induced expression of col1a1, col3a1, and α-SMA in vitro. Mechanically, BLM up-regulated c-Jun and c-Fos expression, which was impeded by KLF2 overexpression. Taken together, our data indicate that KLF2 attenuates pulmonary fibrosis and inflammation, possibly through the regulation of AP-1.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2017

MiR-150 deficiency ameliorated hepatosteatosis and insulin resistance in nonalcoholic fatty liver disease via targeting CASP8 and FADD-like apoptosis regulator

The prevalence of Non-alcoholic fatty liver diseases (NAFLD) increased rapidly in the world. However, the pathogenesis of is still unclear. Hepatic steatosis and insulin resistance are considered to be central to the pathophysiology of NAFLD. MicroRNAs are short non-coding RNAs and has been reported to be involved in pathogenesis of NAFLD and related metabolic diseases. Here, we investigated the mechanisms by which miR-150 regulate hepatic steatosis and insulin resistance in high fat diet (HFD) induced NAFLD model. The expression of miR-150 was up-regulated dramatically in both human NAFLD patients and HFD mice model, as well as in hepatocytes treated with oleic acid. miR-150 deficiency ameliorated the hepatic steatosis and insulin resistance significantly in NAFLD mice. miR-150 deficiency decreased the expression of genes related to fatty acid uptake, synthesis and gluconeogenesis, while increased the expression of genes related to fatty acid β-oxidation. Further, we identified that CFLAR is a direct downstream target of miR-150. Overexpression of miR-150 reduced both the mRNA and protein levels of CFLAR in vitro. And overexpression of miR-150 significantly inhibited the luciferase activity of CFLAR 3′-UTR, while the effect of miR-150 was blocked when the binding site of miR-150 within the CFLAR 3′-UTR was mutated. We also found that miR-150 deficiency decreased the expression of p-Jnk1 and p-Ask1, while the effect of miR-150 on steatosis and insulin signaling was blocked by CFLAR overexpression. In conclusion, our data indicated that miR-150 potentially contributes to the hepatic steatosis and insulin resistance in NAFLD. miR-150/CFLAR pathway may be a new therapeutic strategy against NAFLD.

60 APPLIED LIFE SCIENCES↗