Hsp70- and Hsp90-Mediated Regulation of the Conformation of p53 DNA Binding Domain and p53 Cancer Variants.

The activity of the tumor suppressor p53 has to be timed and balanced closely to prevent untimely induction of cell death. The stability of p53 depends on the ubiquitin ligase Mdm2 but also on Hsp70 and Hsp90 chaperones that interact with its DNA binding domain (DBD). Using hydrogen exchange mass spectrometry and biochemical methods, we analyzed conformational states of wild-type p53-DBD at physiological temperatures and conformational perturbations in three frequent p53 cancer mutants. We demonstrate that the Hsp70/Hdj1 system shifts the conformational equilibrium of p53 toward a flexible, more mutant-like, DNA binding inactive state by binding to the DNA binding loop. The analyzed cancer mutants are likewise destabilized by interaction with the Hsp70/Hdj1 system. In contrast, Hsp90 protects the DBD of p53 wild-type and mutant proteins from unfolding. We propose that the Hsp70 and Hsp90 chaperone systems assume complementary functions to optimally balance conformational plasticity with conformational stability.

Exploring the Roles of Key Mediators IKBKE and HSPA1A in Alzheimer's Disease and Hepatocellular Carcinoma through Bioinformatics Analysis.

Recent studies have hinted at a potential link between Alzheimer's Disease (AD) and cancer. Thus, our study focused on finding genes common to AD and Liver Hepatocellular Carcinoma (LIHC), assessing their promise as diagnostic indicators and guiding future treatment approaches for both conditions. Our research utilized a broad methodology, including differential gene expression analysis, Weighted Gene Co-expression Network Analysis (WGCNA), gene enrichment analysis, Receiver Operating Characteristic (ROC) curves, and Kaplan-Meier plots, supplemented with immunohistochemistry data from the Human Protein Atlas (HPA) and machine learning techniques, to identify critical genes and significant pathways shared between AD and LIHC. Through differential gene expression analysis, WGCNA, and machine learning methods, we identified nine key genes associated with AD, which served as entry points for LIHC analysis. Subsequent analyses revealed IKBKE and HSPA1A as shared pivotal genes in patients with AD and LIHC, suggesting these genes as potential targets for intervention in both conditions. Our study indicates that IKBKE and HSPA1A could influence the onset and progression of AD and LIHC by modulating the infiltration levels of immune cells. This lays a foundation for future research into targeted therapies based on their shared mechanisms.

Integrating single-nucleus sequence profiling to reveal the transcriptional dynamics of Alzheimer's disease, Parkinson's disease, and multiple sclerosis.

BACKGROUND: Alzheimer's disease (AD), Parkinson's disease (PD), and multiple sclerosis (MS) are three nervous system diseases that partially overlap clinically and genetically. However, bulk RNA-sequencing did not accurately detect the core pathogenic molecules in them. The availability of high-quality single cell RNA-sequencing data of post-mortem brain collections permits the generation of a large-scale gene expression in different cells in human brain, focusing on the molecular features and relationships between diseases and genes. We integrated single-nucleus RNA-sequencing (snRNA-seq) datasets of human brains with AD, PD, and MS to identify transcriptomic commonalities and distinctions among them. METHODS: The snRNA-seq datasets were downloaded from Gene Expression Omnibus (GEO) database. The Seurat package was used for snRNA-seq data processing. The uniform manifold approximation and projection (UMAP) were utilized for cluster identification. The FindMarker function in Seurat was used to identify the differently expressed genes. Functional enrichment analysis was carried out using the Gene Set Enrichment Analysis (GSEA) and Gene ontology (GO). The protein-protein interaction (PPI) analysis of differentially expressed genes (DEGs) was analyzed using STRING database ( http://string-db.org ). SCENIC analysis was performed using utilizing pySCENIC (v0.10.0) based on the hg19-tss-centered-10 kb-10species databases. The analysis of potential therapeutic drugs was analyzed on Connectivity Map ( https://clue.io ). RESULTS: The gene regulatory network analysis identified several hub genes regulated in AD, PD, and MS, in which HSPB1 and HSPA1A were key molecules. These upregulated HSP family genes interact with ribosome genes in AD and MS, and with immunomodulatory genes in PD. We further identified several transcriptional regulators (SPI1, CEBPA, TFE3, GRHPR, and TP53) of the hub genes, which has important implications for uncovering the molecular crosstalk among AD, PD, and MS. Arctigenin was identified as a potential therapeutic drug for AD, PD, and MS. CONCLUSIONS: Together, the integrated snRNA-seq data and findings have significant implications for unraveling the shared and unique molecular crosstalk among AD, PD, and MS. HSPB1 and HSPA1A as promising targets involved in the pathological mechanisms of neurodegenerative diseases. Additionally, the identification of arctigenin as a potential therapeutic drug for AD, PD, and MS further highlights its potential in treating these neurological disorders. These discoveries lay the groundwork for future research and interventions to enhance our understanding and treatment of AD, PD, and MS.

The profile of HSPA1A gene expression and its association with heat tolerance in crossbred cattle and the tropically adapted dwarf Vechur and Kasaragod.

Certain livestock breeds are adapted to hot and humid environments, and these breeds have genetics that could be useful in a changing climate. The expression of several genes has been identified as a useful biomarker for heat stress. In this study, the responses to heat exposure of heat-tolerant Vechur and Kasaragod cattle found in Kerala state in India (also known as dwarf Bos taurus indicus) were compared to crossbred cattle (crosses of Bos t. taurus with Bos t. indicus). At various time points during heat exposure, rectal temperature and the expression of HSPA1A were determined, and the relationship between them was characterized. We characterized HSPA1A mRNA in Vechur cattle and performed molecular clock analysis. The expression of HSPA1A between the lineages and at different temperature humidity index (THI) was significant. There were significant differences between the expression profiles of HSPA1A in Kasaragod and crossbred (p < 0.01) and Vechur and crossbred (p < 0.01) cattle, but no significant difference in expression was observed between Vechur and Kasaragod cattle. The genetic distance between Vechur, B. grunniens, B. t. taurus, and B. t. indicus was 0.0233, 0.0059, and 0.007, respectively. The genetic distance between Vechur and the Indian dwarf breed Malnad Gidda was 0.0081. A molecular clock analysis revealed divergent adaptive evolution of Vechur cattle to B. t. taurus, with adaptations to the high temperatures and humidity that are prevalent in their breeding tract in Kerala, India. These results could also prove useful in selecting heat-tolerant animals using HSPA1A as a marker.

Heat-Induced Proteotoxic Stress Response in Placenta-Derived Stem Cells (PDSCs) Is Mediated through HSPA1A and HSPA1B with a Potential Higher Role for HSPA1B.

Placenta-derived stem cells (PDSCs), due to unique traits such as mesenchymal and embryonic characteristics and the absence of ethical constraints, are in a clinically and therapeutically advantageous position. To aid in stemness maintenance, counter pathophysiological stresses, and withstand post-differentiation challenges, stem cells require elevated protein synthesis and consequently augmented proteostasis. Stem cells exhibit source-specific proteostasis traits, making it imperative to study them individually from different sources. These studies have implications for understanding stem cell biology and exploitation in the augmentation of therapeutic applications. Here, we aim to identify the primary determinants of proteotoxic stress response in PDSCs. We generated heat-induced dose-responsive proteotoxic stress models of three stem cell types: placental origin cells, the placenta-derived mesenchymal stem cells (pMSCs), maternal origin cells, the decidua parietalis mesenchymal stem cells (DPMSCs), and the maternal-fetal interface cells, decidua basalis mesenchymal stem cells (DBMSCs), and measured stress induction through biochemical and cell proliferation assays. RT-PCR array analysis of 84 genes involved in protein folding and protein quality control led to the identification of Hsp70 members HSPA1A and HSPA1B as the prominent ones among 17 significantly expressed genes and with further analysis at the protein level through Western blotting. A kinetic analysis of HSPA1A and HSPA1B gene and protein expression allowed a time series evaluation of stress response. As identified by protein expression, an active stress response is in play even at 24 h. More prominent differences in expression between the two homologs are detected at the translational level, alluding to a potential higher requirement for HSPA1B during proteotoxic stress response in PDSCs.

Transcriptomic and Proteomic Analysis of CRISPR/Cas9-Mediated ARC-Knockout HEK293 Cells.

Arc/Arg3.1 (activity-regulated cytoskeletal-associated protein (ARC)) is a critical regulator of long-term synaptic plasticity and is involved in the pathophysiology of schizophrenia. The functions and mechanisms of human ARC action are poorly understood and worthy of further investigation. To investigate the function of the ARC gene in vitro, we generated an ARC-knockout (KO) HEK293 cell line via CRISPR/Cas9-mediated gene editing and conducted RNA sequencing and label-free LC-MS/MS analysis to identify the differentially expressed genes and proteins in isogenic ARC-KO HEK293 cells. Furthermore, we used bioluminescence resonance energy transfer (BRET) assays to detect interactions between the ARC protein and differentially expressed proteins. Genetic deletion of ARC disturbed multiple genes involved in the extracellular matrix and synaptic membrane. Seven proteins (HSPA1A, ENO1, VCP, HMGCS1, ALDH1B1, FSCN1, and HINT2) were found to be differentially expressed between ARC-KO cells and ARC wild-type cells. BRET assay results showed that ARC interacted with PSD95 and HSPA1A. Overall, we found that ARC regulates the differential expression of genes involved in the extracellular matrix, synaptic membrane, and heat shock protein family. The transcriptomic and proteomic profiles of ARC-KO HEK293 cells presented here provide new evidence for the mechanisms underlying the effects of ARC and molecular pathways involved in schizophrenia pathophysiology.

S-Glutathionylation of human inducible Hsp70 reveals a regulatory mechanism involving the C-terminal α-helical lid.

Heat shock protein 70 (Hsp70) proteins are a family of ancient and conserved chaperones. Cysteine modifications have been widely detected among different Hsp70 family members in vivo, but their effects on Hsp70 structure and function are unclear. Here, we treated HeLa cells with diamide, which typically induces disulfide bond formation except in the presence of excess GSH, when glutathionylated cysteines predominate. We show that in these cells, HspA1A (hHsp70) undergoes reversible cysteine modifications, including glutathionylation, potentially at all five cysteine residues. In vitro experiments revealed that modification of cysteines in the nucleotide-binding domain of hHsp70 is prevented by nucleotide binding but that Cys-574 and Cys-603, located in the C-terminal α-helical lid of the substrate-binding domain, can undergo glutathionylation in both the presence and absence of nucleotide. We found that glutathionylation of these cysteine residues results in unfolding of the α-helical lid structure. The unfolded region mimics substrate by binding to and blocking the substrate-binding site, thereby promoting intrinsic ATPase activity and competing with binding of external substrates, including heat shock transcription factor 1 (Hsf1). Thus, post-translational modification can alter the structure and regulate the function of hHsp70.

LASP1 promotes proliferation, metastasis, invasion in head and neck squamous cell carcinoma and through direct interaction with HSPA1A.

LIM and SH3 protein 1 (LASP1) is a specific focal adhesion protein that promotes metastasis in a variety of tumours. However, its role in head and neck squamous cell carcinoma (HNSCC) has not been fully validated. The purpose of this study was to analyse the interaction of LASP1 and its binding partner in HNSCC. The expression of LASP1 and HSPA1A in HNSCC was analysed by real-time PCR and Western blot. The effects of LASP1 on the biology behaviour of HNSCC cell lines were observed in vivo and in vitro. Co-immunoprecipitation analysis was performed to confirm the interaction between LASP1 and HSPA1A. LASP1 was highly expressed in HNSCC and associated with poor prognosis for patients. LASP1 also promoted cell proliferation, colony formation, invasion and cell cycle G2/M phase transition. Heat shock protein family A member 1A (HSPA1A) is identified as a chaperone protein of LASP1 and co-localized in the cytoplasm. HSPA1A positively regulates the interaction of LASP1 with P-AKT and enhances the malignant behaviour of HNSCC cells. LASP1 and HSPA1A are both up-regulated in HNSCC, and directly binds to each other. Double inhibition of LASP1 and HSPA1A expression may be an effective method for the treatment of HNSCC.

Inhibition of HSP90 and Activation of HSF1 Diminish Macrophage NLRP3 Inflammasome Activity in Alcohol-Associated Liver Injury.

BACKGROUND: Activation of NLRP3 in liver macrophages contributes to alcohol-associated liver disease (ALD). Molecular chaperone heat shock protein (HSP) 90 facilitates NLRP3 inflammasome activity during infections and inflammatory diseases. We previously reported that HSP90 is induced in ALD and regulates proinflammatory cytokines, tumor necrosis factor alpha, and IL-6. Whether HSP90 affects IL-1ß and IL-18 regulated by NLRP3 inflammasome in ALD is unknown. Here, we hypothesize that HSP90 modulated NLRP3 inflammasome activity and affects IL-1ß and IL-18 secretion in ALD. METHODS: The expression of HSP90AA1 and NLRP3 inflammasome genes was evaluated in human alcoholic livers and in mouse model of ALD. The importance of HSP90 on NLRP3 inflammasome activation in ALD was evaluated by administering HSP90 inhibitor, 17-dimethylaminoethylamino-17-demethoxygeldanamycin (17-DMAG) to mice subjected to ALD, and in vitro to bone marrow-derived macrophages (BMDM) stimulated with LPS and ATP. The effect of activation of HSF1/HSPA1A axis during HSP90 inhibition or direct activation during heat shock of BMDMs on NLRP3 activity and secretion of downstream cytokines was evaluated. RESULTS: We found positive correlation between induction of HSP90 and NLRP3 inflammasome genes in human alcoholic cirrhotic livers. Administration of 17-DMAG in mouse model of ALD significantly down-regulated NLRP3 inflammasome-mediated caspase-1 (CASP-1) activity and cytokine secretion, with reduction in ALD. 17-DMAG-mediated decrease in NLRP3 was restricted to liver macrophages. Using BMDMs, we show that inhibition of HSP90 prevented CASP-1 activity, and Gasdermin D (GSDMD) cleavage, important in release of active IL-1ß and IL-18. Interestingly, activation of the heat shock factor 1 (HSF1)/HSPA1A axis, either during HSP90 inhibition or by heat shock, decreased NLRP3 inflammasome activity and reduced secretion of cytokines. CONCLUSION: Our studies indicate that inhibition of HSP90 and activation of HSF1/HSPA1A reduce IL-1ß and IL-18 via decrease in NLRP3/CASP-1 and GSDMD activity in ALD.

Andrographolide and Its 14-Aryloxy Analogues Inhibit Zika and Dengue Virus Infection.

Andrographolide is a labdene diterpenoid with potential applications against a number of viruses, including the mosquito-transmitted dengue virus (DENV). In this study, we evaluated the anti-viral activity of three 14-aryloxy analogues (ZAD-1 to ZAD-3) of andrographolide against Zika virus (ZIKV) and DENV. Interestingly, one analogue, ZAD-1, showed better activity against both ZIKV and DENV than the parental andrographolide. A two-dimension (2D) proteomic analysis of human A549 cells treated with ZAD-1 compared to cells treated with andrographolide identified four differentially expressed proteins (heat shock 70 kDa protein 1 (HSPA1A), phosphoglycerate kinase 1 (PGK1), transketolase (TKT) and GTP-binding nuclear protein Ran (Ran)). Western blot analysis confirmed that ZAD-1 treatment downregulated expression of HSPA1A and upregulated expression of PGK1 as compared to andrographolide treatment. These results suggest that 14-aryloxy analogues of andrographolide have the potential for further development as anti-DENV and anti-ZIKV agents.

Increase of Hspa1a and Hspa1b genes in the resting B cells of Sirt1 knockout mice.

Sirt1, also known as the longevity gene, is an NAD+-dependent class III histone deacetylase that has been extensively studied in multiple areas of research including cellular metabolism, longevity, cancer, autoimmunity, and immunity. However, little is known about the function of Sirt1 in B cells. This study aimed to investigate the role of Sirt1 in the expression pattern of mRNAs in the resting B cells of mice. CD19+ B cell-specific inducible Sirt1 knockout (KO) mice were divided into tamoxifen-treated Sirt1 KO group (S19T) or control group (S19). mRNAs extracted from resting B cells of both groups were analyzed for differentially expressed genes (DEG) using microarray. DEG analysis showed significant differential expression of 20 genes, of which Hspa1a and Hspa1b showed the highest fold change (FC) in S19T compared with S19 (p value < 0.01 and FC > 3). Further, Kyoto Encyclopedia of Genes and Genomes analysis identified pathways associated with diseases, organismal systems, and antigen processing and presentation. Additionally, the pathways known to involve Hspa1a and Hspa1b were also activated in the S19T group. On the other hand, after in vitro stimulation with lipopolysaccharide, cell viability and IgM production were significantly decreased in Sirt1 KO B cells, while expressions of TNF-α, IL-6, and IL-10 were increased. In summary, our study reveals that Sirt1 may maintain the quiescent state in resting B cells by suppressing the increase of Hspa1a and Hspa1b. This work provides a foundation for further studies on the functional roles of Sirt1 in B cells.

Knockdown of Heat Shock Proteins HSPA6 (Hsp70B') and HSPA1A (Hsp70-1) Sensitizes Differentiated Human Neuronal Cells to Cellular Stress.

Heat shock proteins are involved in cellular repair and protective mechanisms that counter characteristic features of neurodegenerative diseases such as protein misfolding and aggregation. The HSPA (Hsp70) multigene family includes the widely studied HSPA1A (Hsp70-1) and the little studied HSPA6 (Hsp70B') which is present in the human genome and not in mouse and rat. The effect of knockdown of HSPA6 and HSPA1A expression was examined in relation to the ability of differentiated human SH-SY5Y neuronal cells to tolerate thermal stress. Low dose co-application of celastrol and arimoclomol, which induces Hsps, enhanced the ability of differentiated neurons to survive heat shock. Small interfering RNA (siRNA) knockdown of HSPA6 and HSPA1A resulted in loss of the protective effect of co-application of celastrol/arimoclomol. More pronounced effects on neuronal viability were apparent at 44 °C heat shock compared to 43 °C. siRNA knockdown suggests that HSPA6 and HSPA1A contribute to protection of differentiated human neuronal cells from cellular stress.

Development of HSPA1A promoter-driven luciferase reporter gene assays in human cells for assessing the oxidative damage induced by silver nanoparticles.

The exponential increase in the total number of engineered nanoparticles in consumer products requires novel tools for rapid and cost-effective toxicology screening. In order to assess the oxidative damage induced by nanoparticles, toxicity test systems based on a human HSPA1A promoter-driven luciferase reporter in HepG2, LO2, A549, and HBE cells were established. After treated with heat shock and a group of silver nanoparticles (AgNPs) with different primary particle sizes, the cell viability, oxidative damage, and luciferase activity were determined. The time-dependent Ag(+) ions release from AgNPs in cell medium was also evaluated. Our results showed that heat shock produced a strong time-dependent induction of relative luciferase activity in the four luciferase reporter cells. Surprisingly, at 4h of recovery, the relative luciferase activity was >98× the control level in HepG2-luciferase cells. Exposure to different sizes of AgNPs resulted in activation of the HSPA1A promoter in a dose-dependent manner, even at low cytotoxic or non-cytotoxic doses. The smaller (5nm) AgNPs were more potent in luciferase induction than the larger (50 and 75nm) AgNPs. These results were generally in accordance with the oxidative damage indicated by malondialdehyde concentration, reactive oxygen species induction and glutathione depletion, and Ag(+) ions release in cell medium. Compared with the other three luciferase reporter cells, the luciferase signal in HepG2-luciferase cells is obviously more sensitive and stable. We conclude that the luciferase reporter cells, especially the HepG2-luciferase cells, could provide a valuable tool for rapid screening of the oxidative damage induced by AgNPs.

Localization of heat shock protein HSPA6 (HSP70B') to sites of transcription in cultured differentiated human neuronal cells following thermal stress.

Heat shock proteins (Hsps) are a set of highly conserved proteins that are involved in cellular repair and protective mechanisms. In order to identify potential stress-sensitive sites in differentiated SH-SY5Y human neuronal cells, localization of two inducible members of the HSPA (HSP70) family was investigated, namely HSPA6 (HSP70B') and HSPA1A (HSP70-1). Following heat shock, yellow fluorescent protein (YFP)-tagged HSPA6 and HSPA1A proteins localized to nuclear speckles that are enriched in RNA splicing factors (identified by SC35 and SON marker proteins) and then to the granular component of the nucleolus (identified by nucleophosmin). Subsequently, YFP-HSPA6 protein, but not YFP-HSPA1A, localized to the periphery of nuclear speckles that are sites of RNA transcription. The HSPA6 gene is present in the human genome but not in genomes of rat and mouse. Hence, current animal models of neurodegenerative diseases are lacking a potentially protective member of the HSPA family. Potential stress-sensitive sites were identified in differentiated human SH-SY5Y cells by the localization of HSPA6 (HSP70B') and HSPA1A (HSP70-1) to nuclear components following heat shock. HSPA6 and HSPA1A rapidly moved to nuclear speckles, enriched in RNA splicing factors, then to the granular layer of the nucleolus. Subsequently, HSPA6 exhibited a novel localization not observed for the more widely studied HSPA1A, namely association with the periphery of nuclear speckles that are sites of transcription. HS = heat shock; HSPA6 = HSP70B' protein; HSPA1A = HSP70-1 protein.

Development block of golden hamster ICSI embryos is associated with decreased expression of HDAC1, HSPA1A and MYC.

We have investigated the mechanism for embryo development block in vitro and to improve the development rate of golden hamster embryos in vitro. Intracytoplasmic sperm injection (ICSI) technique was used to produce golden hamster ICSI embryos. The changes in the histone acetylation and the expression of histone deacetylase and related genes were analyzed by immunocytochemical staining and real-time PCR both in golden hamster in vivo embryos and in ICSI embryos. Aged oocytes significantly increased the oocyte spontaneous activation rate. In vitro cultured ICSI embryos suffered from severe development block in M199TE medium. Expression of histone deacetylase 1 (HDAC1) was significantly decreased in the nuclei of the arrested ICSI 2-cell embryos, and its nuclear and cytoplasmic expression pattern was also markedly altered. The acetylation level of H4K5, however, was not significantly changed between golden hamster in vivo embryos and ICSI embryos. HSPA1A and MYC, the marker genes for zygotic genome activation (ZGA), were transcriptionally decreased in arrested ICSI 2-cell embryos. Transcription of HDAC1 was also downregulated in these embryos, whereas the mRNA expression of the proapoptotic gene, BAX, was not changed. These results indicate that the golden hamster ICSI embryo development block during ZGA is associated with decreased nuclear expression and altered expression of HDAC1. HSPA1A, MYC, and HDAC1 mRNA levels, which decrease, resulting in ZGA failure.

The use of CO2 laser in vulvar lichen sclerosus treatment - molecular evidence.

Vulvar lichen sclerosus is chronic and difficult to treat disorder, which offer is recurrent and leads to multiple complications. The limited efficacy of pharmacologic treatment directed the search for new therapies including use of CO2 laser. In our study we focused on collagen and elastin gene expression as well as heat shock proteins and p53 expression in two patients with vulvar lichen sclerosus who underwent CO2 laser therapy. In both patients we observed decreased clinical symptoms observed by an experienced gynecologist as well as significant changes in gene expression before and after laser treatment.

Targeting HSPA1A in ARID2-deficient lung adenocarcinoma.

Somatic mutations of the chromatin remodeling gene ARID2 are observed in ∼7% of human lung adenocarcinomas (LUADs). However, the role of ARID2 in the pathogenesis of LUADs remains largely unknown. Here we find that ARID2 expression is decreased during the malignant progression of both human and mice LUADs. Using two KrasG12D -based genetically engineered murine models, we demonstrate that ARID2 knockout significantly promotes lung cancer malignant progression and shortens overall survival. Consistently, ARID2 knockdown significantly promotes cell proliferation in human and mice lung cancer cells. Through integrative analyses of ChIP-Seq and RNA-Seq data, we find that Hspa1a is up-regulated by Arid2 loss. Knockdown of Hspa1a specifically inhibits malignant progression of Arid2-deficient but not Arid2-wt lung cancers in both cell lines as well as animal models. Treatment with an HSPA1A inhibitor could significantly inhibit the malignant progression of lung cancer with ARID2 deficiency. Together, our findings establish ARID2 as an important tumor suppressor in LUADs with novel mechanistic insights, and further identify HSPA1A as a potential therapeutic target in ARID2-deficient LUADs.

Corrigendum: Long Noncoding RNA NONHSAT079852.2 Contributes to GBM Recurrence by Functioning as a ceRNA for has-mir-10401-3p to Facilitate HSPA1A Upregulation.

[This corrects the article DOI: 10.3389/fonc.2021.636632.].

Long Noncoding RNA NONHSAT079852.2 Contributes to GBM Recurrence by Functioning as a ceRNA for has-mir-10401-3p to Facilitate HSPA1A Upregulation.

Glioblastoma multiforme (GBM) is the most common brain malignancy and major cause of high mortality in patients with GBM, and its high recurrence rate is its most prominent feature. However, the pathobiological mechanisms involved in recurrent GBM remain largely unknown. Here, whole-transcriptome sequencing (RNA-sequencing, RNA-Seq) was used in characterizing the expression profile of recurrent GBM, and the aim was to identify crucial biomarkers that contribute to GBM relapse. Differentially expressed RNAs in three recurrent GBM tissues compared with three primary GBM tissues were identified through RNA-Seq. The function and mechanism of a candidate long noncoding RNA (lncRNA) in the progression and recurrence of GBM were elucidated by performing comprehensive bioinformatics analyses, such as functional enrichment analysis, protein-protein interaction prediction, and lncRNA-miRNA-mRNA regulatory network construction, and a series of in vitro assays. As the most significantly upregulated gene identified in recurrent GBM, HSPA1A is mainly related to antigen presentation and the MAPK signaling pathway, as indicated by functional enrichment analysis. HSPA1A was predicted as the target gene of the lncRNA NONHSAT079852.2. qRT-PCR revealed that NONHSAT079852.2 was significantly elevated in recurrent GBM relative to that in primary GBM, and high NONHSAT079852.2 expression was associated with the poor overall survival rates of patients with GBM. The knockdown of NONHSAT079852.2 successfully induced tumor cell apoptosis, inhibited the proliferation, migration, invasion and the expression level of HSPA1A in glioma cells. NONHSAT079852.2 was identified to be a sponge for hsa-miR-10401-3p through luciferase reporter assay. Moreover, HSPA1A was targeted and regulated by hsa-miR-10401-3p. Collectively, the results suggested that NONHSAT079852.2 acts as a sponge of hsa-mir-10401-3p and thereby enhances HSPA1A expression, promotes tumor cell proliferation and invasion, and leads to the progression and recurrence of GBM. This study will provide new insight into the regulatory mechanisms of NONHSAT079852.2-mediated competing endogenous RNA in the pathogenesis of recurrent GBM and evidence of the potential of lncRNAs as diagnostic biomarkers or potential therapeutic targets.

ACOT4 accumulation via AKT-mediated phosphorylation promotes pancreatic tumourigenesis.

The acyl-CoA thioesterase (ACOT) family catalyses the hydrolysis of acyl-CoA thioesters to their corresponding non-esterified fatty acid and coenzyme A (CoA). Increasing evidence suggests that cancer cells generally have altered lipid metabolism in different aspects. However, the roles of the ACOT family in cancer, especially in pancreatic ductal carcinoma (PDAC), are largely unknown. In the present study, we mined data to determine the clinical significance of all eleven ACOT genes among nine major solid tumour types from TCGA database and found that the expression of ACOT4 in PDAC was negatively correlated with patient survival, establishing ACOT4 as a potential biomarker of PDAC. Depletion of ACOT4 attenuated the proliferation and tumour formation of PDAC cells. Using mass spectrometry, HSPA1A was found to associate with ACOT4. Furthermore, we found that phosphorylation of ACOT4 at S392 by AKT decreased the binding of ACOT4 to HSPA1A, resulting in ACOT4 accumulation. The ACOT4 elevation promotes pancreatic tumourigenesis by producing excessive CoA to support tumour cell metabolism. Thus, our study expands the relationship between AKT signalling and lipid metabolism and establishes a functional role of ACOT4 in PDAC.