Member Of Heat Shock Protein Family Research Articles

GroEL protein of the Leptospira spp. interacts with host proteins and induces cytokines secretion on macrophages

BackgroundLeptospirosis is a zoonotic disease caused by infection with spirochetes from Leptospira genus. It has been classified into at least 17 pathogenic species, with more than 250 serologic variants. This wide distribution may be a result of leptospiral ability to colonize the renal tubules of mammalian hosts, including humans, wildlife, and many domesticated animals. Previous studies showed that the expression of proteins belonging to the microbial heat shock protein (HSP) family is upregulated during infection and also during various stress stimuli. Several proteins of this family are known to have important roles in the infectious processes in other bacteria, but the role of HSPs in Leptospira spp. is poorly understood. In this study, we have evaluated the capacity of the protein GroEL, a member of HSP family, of interacting with host proteins and of stimulating the production of cytokines by macrophages.ResultsThe binding experiments demonstrated that the recombinant GroEL protein showed interaction with several host components in a dose-dependent manner. It was also observed that GroEL is a surface protein, and it is secreted extracellularly. Moreover, two cytokines (tumor necrosis factor-α and interleukin-6) were produced when macrophages cells were stimulated with this protein.ConclusionsOur findings showed that GroEL protein may contribute to the adhesion of leptospires to host tissues and stimulate the production of proinflammatory cytokines during infection. These features might indicate an important role of GroEL in the pathogen-host interaction in the leptospirosis.

Prognostic and Functional Significant of Heat Shock Proteins (HSPs) in Breast Cancer Unveiled by Multi-Omics Approaches.

Simple SummaryIn this study, we investigated the expression pattern and prognostic significance of the heat shock proteins (HSPs) family members in breast cancer (BC) by using several bioinformatics tools and proteomics investigations. Our results demonstrated that, collectively, HSPs were deregulated in BC, acting as both oncogene and onco-suppressor genes. In particular, two different HSP-clusters were significantly associated with a poor or good prognosis. Interestingly, the HSPs deregulation impacted gene expression and miRNAs regulation that, in turn, affected important biological pathways involved in cell cycle, DNA replication, and receptors-mediated signaling. Finally, the proteomic identification of several HSPs members and isoforms revealed much more complexity of HSPs roles in BC and showed that their expression is quite variable among patients. In conclusion, we elaborated two panels of HSPs that could be further explored as potential biomarkers for BC progression and prognosis.Heat shock proteins (HSPs) are a well-characterized molecular chaperones protein family, classified into six major families, according to their molecular size. A wide range of tumors have been shown to express atypical levels of one or more HSPs, suggesting that they could be used as biomarkers. However, the collective role and the possible coordination of HSP members, as well as the prognostic significance and the functional implications of their deregulated expression in breast cancer (BC) are poorly investigated. Here, we used a systematic multi-omics approach to assess the HSPs expression, the prognostic value, and the underlying mechanisms of tumorigenesis in BC. By using data mining, we showed that several HSPs were deregulated in BC and significantly correlated with a poor or good prognosis. Functional network analysis of HSPs co-expressed genes and miRNAs highlighted their regulatory effects on several biological pathways involved in cancer progression. In particular, these pathways concerned cell cycle and DNA replication for the HSPs co-expressed genes, and miRNAs up-regulated in poor prognosis and Epithelial to Mesenchymal Transition (ETM), as well as receptors-mediated signaling for the HSPs co-expressed genes up-regulated in good prognosis. Furthermore, the proteomic expression of HSPs in a large sample-set of breast cancer tissues revealed much more complexity in their roles in BC and showed that their expression is quite variable among patients and confined into different cellular compartments. In conclusion, integrative analysis of multi-omics data revealed the distinct impact of several HSPs members in BC progression and indicate that collectively they could be useful as biomarkers and therapeutic targets for BC management.

Comparative transcriptomic analysis brings new insights into the response to acute temperature acclimation in burbot (Lota lota lota)

Temperature is a crucial environmental factor affecting the survival and growth of aquatic organisms. However, the global warming caused by the growth of greenhouse gas emission has severely affected the sustainable development of global nature ecosystem. The change in water temperature has caused a disorder in the aquatic ecosystems. Lota lota lota is a unique member of cold-freshwater codfish; it has an important economic value like its marine relatives. It is also a good model for studying the molecular mechanisms of temperature adaptability of fish. In this study, RNA sequencing based on the Illumina HiSeq™ 2500 platform was used to reveal gene expression at the transcriptome level. Transcriptome profiles were generated from liver and brain samples from fish exposed to (0 °C, 17 °C and 28 °C). A total of 30,958 annotated differentially expressed genes (DEGs) were identified in all comparisons, with more DEGs in high-temperature groups (1668) than in low-temperature groups (1110). GO analysis was divided into three extensive categories and 55 subcategories. A series of candidate genes and pathways associated with temperature responsiveness was detected, mainly including heat-shock protein (HSP) family, fatty acid desaturase (FAD) and antigen processing and presentation. The expression levels of HSP70 were highest amongst those of the HSP family members in the high-temperature groups, implying that HSP70 was an efficient biomarker of high temperature challenge in L. lota lota. An obvious upward trend in the expression of FAD was conducive to regulating fatty acid composition and maintaining membrane fluidity under low-temperature stress. In addition, evidence indicated that elevated temperature had a large influence on immune function and heat stress could upregulate the immune-related pathways in L. lota lota.

DNAJC5 promotes hepatocellular carcinoma cells proliferation though regulating SKP2 mediated p27 degradation

DNAJC5 (DnaJ heat shock protein family (Hsp40) member C5), also known as cysteine tandem protein (CSPα), is important for maintaining the normal function of nerve tissues, but its oncogenic function remains unknown. Here, we report a unique mechanism underlying the oncogenic function of DNAJC5. DNAJC5 protein expression is highly detectable in human hepatocellular carcinoma (HCC) tissues and is strongly related to a poor prognosis among HCC patients. DNAJC5 overexpression promotes HCC cell proliferation and reduced the ratio of cells in G1 phase of the cell cycle. Furthermore, DNAJC5 interacts with SKP2 and enhances the degradation of p27 (a cyclin-dependent kinase inhibitor1B) by promoting formation of the SKP2-p27 complex. In contrast, DNAJC5 knockdown rescues the SKP2-mediated decrease in p27 protein levels. These results reveal that the DNAJC5-SKP2-p27 pathway is a novel mechanism for the oncogenic function of DNAJC5 in HCC.

Transcriptional Enhancer Factor Domain Family member 4 Exerts an Oncogenic Role in Hepatocellular Carcinoma by Hippo-Independent Regulation of Heat Shock Protein 70 Family Members.

Transcriptional enhancer factor domain family member 4 (TEAD4) is a downstream effector of the conserved Hippo signaling pathway, regulating the expression of genes involved in cell proliferation and differentiation. It is up‐regulated in several cancer types and is associated with metastasis and poor prognosis. However, its role in hepatocellular carcinoma (HCC) remains largely unexplored. Using data from The Cancer Genome Atlas, we found that TEAD4 was overexpressed in HCC and was associated with aggressive HCC features and worse outcome. Overexpression of TEAD4 significantly increased proliferation and migration rates in HCC cells in vitro as well as tumor growth in vivo. Additionally, RNA sequencing analysis of TEAD4‐overexpressing HCC cells demonstrated that TEAD4 overexpression was associated with the up‐regulation of genes involved in epithelial‐to‐mesenchymal transition, proliferation, and protein‐folding pathways. Among the most up‐regulated genes following TEAD4 overexpression were the 70‐kDa heat shock protein (HSP70) family members HSPA6 and HSPA1A. Chromatin immunoprecipitation–quantitative real‐time polymerase chain reaction experiments demonstrated that TEAD4 regulates HSPA6 and HSPA1A expression by directly binding to their promoter and enhancer regions. The pharmacologic inhibition of HSP70 expression in TEAD4‐overexpressing cells reduced the effect of TEAD4 on cell proliferation. Finally, by overexpressing TEAD4 in yes‐associated protein (YAP)/transcriptional coactivator with PDZ binding motif (TAZ)‐knockdown HCC cells, we showed that the effect of TEAD4 on cell proliferation and its regulation of HSP70 expression does not require YAP and TAZ, the main effectors of the Hippo signaling pathway. Conclusion: A novel Hippo‐independent mechanism for TEAD4 promotes cell proliferation and tumor growth in HCC by directly regulating HSP70 family members.

Cryo-EM structure of human mitochondrial HSPD1

SummaryChaperonins play an important role in folding newly synthesized or translocated proteins in all organisms. The bacterial chaperonin GroEL has served as a model system for the understanding of these proteins. In comparison, its human homolog, known as mitochondrial heat shock protein family member D1 (HSPD1) is poorly understood. Here, we present the structure of HSPD1 in the apo state determined by cryo-electron microscopy (cryo-EM). Unlike GroEL, HSPD1 forms mostly single ring assemblies in the absence of co-chaperonin (HSPE1). Comparison with GroEL shows a rotation and increased flexibility of the apical domain. Together with published structures of the HSPD1/HSPE1 co-chaperonin complex, this work gives insight into the structural changes that occur during the catalytic cycle. This new understanding of HSPD1 structure and its rearrangements upon complex formation may provide new insights for the development of HSPD1-targeting treatments against a diverse range of diseases including glioblastoma.

Bi-allelic pathogenic variations in DNAJB11 cause Ivemark II syndrome, a renal-hepatic-pancreatic dysplasia

DNAJB11 (DnaJ Heat Shock Protein Family (Hsp40) Member B11) heterozygous loss of function variations have been reported in autosomal dominant cystic kidney disease with extensive fibrosis, associated with maturation and trafficking defect involving both the autosomal dominant polycystic kidney disease protein polycystin-1 and the autosomal dominant tubulointerstitial kidney disease protein uromodulin. Here we show that biallelic pathogenic variations in DNAJB11 lead to a severe fetal disease including enlarged cystic kidneys, dilation and proliferation of pancreatic duct cells, and liver ductal plate malformation, an association known as Ivemark II syndrome. Cysts of the kidney were developed exclusively from uromodulin negative tubular segments. In addition, tubular cells from the affected kidneys had elongated primary cilia, a finding previously reported in ciliopathies. Thus, our data show that the recessive disease associated with DNAJB11 variations is a ciliopathy rather than a disease of the autosomal dominant tubulointerstitial kidney disease spectrum, and prompt screening of DNAJB11 in fetal hyperechogenic/cystic kidneys.

Metformin rescues muscle function in BAG3 myofibrillar myopathy models

ABSTRACT Dominant de novo mutations in the co-chaperone BAG3 cause a severe form of myofibrillar myopathy, exhibiting progressive muscle weakness, muscle structural failure, and protein aggregation. To elucidate the mechanism of disease in, and identify therapies for, BAG3 myofibrillar myopathy, we generated two zebrafish models, one conditionally expressing BAG3P209L and one with a nonsense mutation in bag3. While transgenic BAG3P209L-expressing fish display protein aggregation, modeling the early phase of the disease, bag3 -/- fish exhibit exercise dependent fiber disintegration, and reduced swimming activity, consistent with later stages of the disease. Detailed characterization of the bag3 -/- fish, revealed an impairment in macroautophagic/autophagic activity, a defect we confirmed in BAG3 patient samples. Taken together, our data highlights that while BAG3P209L expression is sufficient to promote protein aggregation, it is the loss of BAG3 due to its sequestration within aggregates, which results in impaired autophagic activity, and subsequent muscle weakness. We therefore screened autophagy-promoting compounds for their effectiveness at removing protein aggregates, identifying nine including metformin. Further evaluation demonstrated metformin is not only able to bring about the removal of protein aggregates in zebrafish and human myoblasts but is also able to rescue the fiber disintegration and swimming deficit observed in the bag3 −/- fish. Therefore, repurposing metformin provides a promising therapy for BAG3 myopathy. Abbreviations:ACTN: actinin, alpha; BAG3: BAG cochaperone 3; CRYAB: crystallin alpha B; DES: desmin; DMSO: dimethyl sulfoxide; DNAJB6: DnaJ heat shock protein family (Hsp40) member B6; dpf: days post fertilization; eGFP: enhanced green fluorescent protein; FDA: Food and Drug Administration; FHL1: four and a half LIM domains 1; FLNC: filamin C; hpf: hours post-fertilization; HSPB8: heat shock protein family B [small] member 8; LDB3/ZASP: LIM domain binding 3; MYOT: myotilin; TTN: titin; WT: wild-type.

AN UNUSUAL CASE OF ACUTE HYPOXIC RESPIRATORY FAILURE IN A PATIENT WITH FIBRILLARY GLOMERULONEPHRITIS

SESSION TITLE: Fellows Critical Care Posters SESSION TYPE: Fellow Case Report Posters PRESENTED ON: October 18-21, 2020 INTRODUCTION: Fibrillary glomerulonephritis (FGN) is a rare primary glomerular disease (incidence <1%) characterized by immune-complex mediated GN with amyloid-like fibrils which are IgG positive and Congo red negative(1). We present a case of refractory respiratory failure with co-existing FGN. CASE PRESENTATION: A 58-year-old woman with hypertension and chronic hematuria/ proteinuria (presumed IgA nephropathy but no biopsy) presented to the hospital with fever and dyspnea for 2 days. Due to moderate hypoxemia she required invasive mechanical ventilation (IMV). CXR showed bilateral diffuse airspace opacities. Lab work up showed proteinuria, hematuria, elevated CRP and normal creatinine. ANCA panel, complements, anti-GBM antibody, anti-SSA/SSB, RF and anti Scl-70 were negative. BAL showed no evidence of DAH, neutrophilic alveolitis and sterile cultures. Echocardiogram was normal. Despite lung protective IMV, PEEP adjustment as per ARDS net and adjuvant therapies (inhaled nitric oxide, steroids and paralytics), severe hypoxemia persisted. She developed AKI requiring renal replacement therapy. She received appropriate antibiotics for ESBL E. coli UTI and C. diff. colitis. Kidney biopsy showed FGN. She underwent plasma exchange, intravenous immunoglobulin therapy followed by rituximab per management protocol used in other pulmonary renal syndromes. Due to worsening of severe ARDS refractory to medical therapy, VV-ECMO was initiated in week 4. She was unable to be weaned off ECMO support (>30 days) which lead to her death. DISCUSSION: FGN was first described in 1977(2). Fibrils characterizing FGN are confined to glomeruli and stain intensely by IF for IgG, C3, ?, and ?, strongly suggesting that the fibrils are composed of a complex of antibodies and antigens as seen in our case(3). It was described in association with underlying malignancy, hepatitis C, diabetes or autoimmune disease. Nephrotic range proteinuria, hematuria and hypertension are usual presentations. Recent discovery of major fibril component, DNA-J heat-shock protein family member B9 (DNAJB9), has made the diagnosis of FGN by immunohistochemistry possible(4,5). Nearly half of patients with FGN progress to ESRD within 4 years. Benefit from immunosuppressive therapy is unestablished. Transplant is not curative as recurrence is not uncommon. Very few cases of extra-renal manifestations of FGN have been reported. Only cases associated with pulmonary hemorrhage were successfully treated with steroids(6). CONCLUSIONS: Our case reports a rare extra-renal manifestation of FGN causing refractory respiratory failure. Further research to identify exact pathogenesis and successful treatment is imperative. Reference #1: 1. Fibrillary Glomerulonephritis: Clinicopathological features and atypical cases from a multi-institutional cohort, CJASN Dec 2019, 14 (12) 1741-50 2. Nephrotic syndrome associated with amyloid-like glomerular deposits. Nephron. 1977;18:301-308 Reference #2: 3. Fibrillary Glomerulonephritis: An update, Kidney Int Rep. 2019 Jul; 4(7): 917-922 4. DNA J heat shock protein family B member 9 is a novel biomarker for fibrillary GN. J Am Soc Nephrol. 2018;29:51–56 Reference #3: 5. DNA J homolog subfamily B member 9 is a putative autoantigen in fibrillary GN. J Am Soc Nephrol. 2018; 29:231–239 6. Fibrillary Glomerulonephritis: Another cause of bleeding lung. Case reports, ATS 2017 conference DISCLOSURES: No relevant relationships by Moises Cossio, source=Web Response No relevant relationships by Domingo Franco-Palacios, source=Web Response No relevant relationships by Kushagra Gupta, source=Web Response No relevant relationships by Shahzad Hussain, source=Web Response No relevant relationships by Venkateswara Kollipara, source=Web Response No relevant relationships by Toribiong Uchel, source=Web Response

Molecular characterization and expression patterns of heat shock proteins in Spodoptera littoralis, heat shock or immune response?

The Egyptian cotton leaf worm, Spodoptera littoralis (Boisd.), is a major agricultural lepidopterous pest causing extensive damage in a variety of crops including vegetable, cotton, fodder, and fiber crops. Heat shock protein (HSP) family members play important roles in protecting insects against environmental stressors. In this study, we characterized three putative heat shock proteins (SpliHsp70, SpliHsp90, and SpliHSF) from S. littoralis and analyzed their expression levels in response to heat, cold, ultraviolet irradiation, Bacillus thuringiensis, and Spodoptera littoralis nucleopolyhedrovirus treatments. Significant upregulation of SpliHsp70 was observed in female pupae, while the highest expression levels of SpliHsp90 and SpliHSF were found in female adults. Heat shock triggered increases in SpliHsp levels compared to cold treatment. SpliHsp90 exhibited the highest expression levels during the first 30 min of UV treatment. Both bacterial and viral pathogenic agents effected the regulation of Hsps in S. littoralis. These findings suggest that SpliHsp genes might play significant roles in the response to biotic and abiotic stress, as well as in the regulation of developmental stages.

Abstract 5972: HLJ1 deficiency enhanced diethylnitrosamine-induced hepatocarcinogenesis in a gene targeting mouse model

Abstract Liver cancer is the sixth most common cancer and the fourth leading cause of cancer-related death worldwide. The conventional treatments such as surgical resection and liver transplantation are applicable to only a small proportion of patients and prolongation of survival is restricted. Therefore, it is urgent to develop cancer biomarkers for defining cancer risk, diagnosis, prognosis and even find new therapeutic targets. Human Liver DnaJ-Like Protein (HLJ1), which belongs to DnaJ heat shock protein family (HSP40) member B4 (DNAJB4), has been reported to act as a tumor suppressor in NSCLC, colorectal cancer, melanoma metastasis. However, the precise role of HLJ1 in liver cancer tumorigenesis and the progress of HCC carcinogenesis has not been elucidated yet. Thus, we investigate whether HLJ1 has tumor-suppressive or oncogenic role on DEN-induced liver cancer development in a mouse model. Wild-type (WT) and HLJ1 knockout (HLJ1−/−) mice are injected with 20 mg/kgbw DEN at postnatal day 15. Thereafter, 50 mg/kgbw DEN is administrated weekly to mice between age 4 to 12 weeks. Mice were sacrificed at age 30 and 36 weeks to assess the tumor count and size. In another group, mice are injected with DEN and age day 15 and then 500 ppm PB is administrated in drinking water until mice were sacrifice at age 52 weeks. The liver tissues are frozen and fixed for western blotting and IHC analysis. For cDNA microarray analysis, gene expression microarray samples were from 6-8 weeks wild-type and HLJ1−/− mice livers. When sacrificed at age 30 and 36 weeks, HLJ1−/− mice exhibited higher tumor multiplicity, serum ALT, AST levels, and larger macroscopic tumor nodules than WT mice. In DEN+PB treatment group, higher incidence rate can be observed in HLJ1−/− mice than in WT mice. This indicated HLJ1 protects liver from DEN-induced HCC carcinogenesis and knockout of HLJ1 confers to more severe HCC induction. Furthermore, p-STAT3 expression was higher in normal part of HLJ1−/− mice liver compared to that of WT mice liver, which suggest the role of HLJ1 in regulating p-STAT3 signaling in the early stage of tumor formation. Using IHC staining and western blotting to analyze HLJ1 expression level, we found half of the tumors express higher HLJ1 than adjacent normal part did, which suggested a tumor-promoting role of HLJ1 after tumor initiation and formation. In cDNA microarray analysis, network analysis of genes enriched in HCC map folder revealed involved objects JunB, c-Myc, p21 and Cyclin D1, which was further confirmed by quantitative PCR. p21 and Cyclin D1 were highly expressed in the liver of HLJ1−/− comparing to that of WT mice, which suggested HLJ1-related genes p21 and Cyclin D1 may have roles in HCC formation. Clinical information of 362 liver cancer patients from TCGA database exhibited that higher expression of HLJ1 in tumor part is correlated to lower disease-specific survival rate. It indicated that HLJ1 may be a tumor marker for HCC prognosis and clinical outcome prediction. It also suggested that HLJ1 may play an oncogenic role after tumor formation. In summary, HLJ1 has protective role against DEN-induced hepatocarcinogenesis through down-regulated p-STAT3 and cyclin D1 whereas it could also be an enhancer to promote tumor progression after tumor formation. Hence, HLJ1 could be a target for liver cancer therapy and a prognostic biomarker in the future. Citation Format: Wei-Jia Luo, Min-Hui Chien, Jeremy J. W. Chen, Sung-Liang Yu, Pan-Chyr Yang, Kang-Yi Su. HLJ1 deficiency enhanced diethylnitrosamine-induced hepatocarcinogenesis in a gene targeting mouse model [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 5972.

Elevated levels of autoantibodies against DNAJC2 in sera of patients with atherosclerotic diseases

BackgroundSerum antibody markers have been increasingly identified not only for cancer and autoimmune diseases but also for atherosclerosis-related diseases such as acute ischemic stroke (AIS), acute myocardial infarction (AMI), diabetes mellitus (DM), and chronic kidney disease (CKD). Biomarkers for transient ischemic attack (TIA) and non-ST segment elevation acute coronary syndrome (NSTEACS) are potentially useful for detection of early phase of atherosclerotic changes against AIS and AMI, respectively. MethodsWe utilized serological identification of antigens by recombinant cDNA expression cloning (SEREX) using a human aortic endothelial cell cDNA phage library and sera from patients with TIA or NSTEACS. Serum antibody levels were measured by amplified luminescent proximity homogeneous assay-linked immunosorbent assay (AlphaLISA) using purified recombinant antigens. ResultsScreening of sera from patients with TIA identified DnaJ heat shock protein family (Hsp40) member C2 (DNAJC2) as a candidate antigen, which was also isolated by SEREX screening using sera of patients with NSTEACS. The validation cohort revealed significantly higher DNAJC2 antibody (DNAJC2-Ab) levels in the sera of patients with TIA or AIS than those in healthy donors (HDs). Multivariate logistic regression analysis indicated that the predictive odds ratios (OR) of DNAJC2-Ab levels for TIA and AIS were 2.54 (95% confidence interval [CI]: 1.36–4.74, p = 0.0034) and 2.14 (95% CI: 1.39–3.30, p = 0.0005), respectively. Serum DNAJC2-Ab levels were also higher in patients with AMI, DM, and CKD than those in HDs. ConclusionSerum DNAJC2-Ab level may be useful for early detection of atherosclerotic lesions, which lead to AIS and AMI.

Cancer Antigen Discovery Is Enabled by RNA Sequencing of Highly Purified Malignant and Nonmalignant Cells.

Broadly expressed, highly differentiated tumor-associated antigens (TAA) can elicit antitumor immunity. However, vaccines targeting TAAs have demonstrated disappointing clinical results, reflecting poor antigen selection and/or immunosuppressive mechanisms. Here, a panel of widely expressed, novel colorectal TAAs were identified by performing RNA sequencing of highly purified colorectal tumor cells in comparison with patient-matched colonic epithelial cells; tumor cell purification was essential to reveal these genes. Candidate TAA protein expression was confirmed by IHC, and preexisting T-cell immunogenicity toward these antigens tested. The most promising candidate for further development is DNAJB7 [DnaJ heat shock protein family (Hsp40) member B7], identified here as a novel cancer-testis antigen. It is expressed in many tumors and is strongly immunogenic in patients with cancers originating from a variety of sites. DNAJB7-specific T cells were capable of killing colorectal tumor lines in vitro, and the IFNγ+ response was markedly magnified by control of immunosuppression with cyclophosphamide in patients with cancer. This study highlights how prior methods that sequence whole tumor fractions (i.e., inclusive of alive/dead stromal cells) for antigen identification may have limitations. Through tumor cell purification and sequencing, novel candidate TAAs have been identified for future immunotherapeutic targeting.

The expression profiles and prognostic values of HSPs family members in Head and neck cancer

BackgroundHead and neck squamous cell carcinoma (HNSC) ranks as the sixth most common malignancy. The identification of highly specific and sensitive prognostic markers and potential drug targets can contribute to enhanced patient prognosis and individualized treatments. Heat shock proteins (HSPs) act as molecular chaperones and play a crucial role in maintaining cell homeostasis. Recently, research has indicated that HSPs also act as “evil chaperones” in cancer development.MethodsIn this study, we assessed the expression of HSPs in HNSC patients using the ONCOMINE, GEPIA, and UALCAN databases. Mutations of HSP genes were also analysed using the cBioPortal database. Additionally, the expression levels of HSPs were verified using the Human Protein Altas (THPA) database.ResultsWe found that the expression levels of HSPH1, HSPD1, SERPINH1, HSPA4, and HSP90AA1 were significantly higher in tissues from HNSC patients compared with normal tissues. Moreover, HSPH1, HSPD1, SERPINH1, HSPA4 and HSP90AA1 expressions were linked to disease progression. Survival analysis with the GEPIA and OncoLnc databases indicated that upregulation of HSPH1, HSPD1, SERPINH1, HSPA4 and HSP90AA1 was related to poor overall survival (OS).ConclusionThis study suggests that the HSPH1, HSPD1, SERPINH1, HSPA4 and HSP90AA1 genes are potential clinical targets and prognostic biomarkers for patients with HNSC.

Amyloid-β oligomers are captured by the DNAJB6 chaperone: Direct detection of interactions that can prevent primary nucleation

A human molecular chaperone protein, DnaJ heat shock protein family (Hsp40) member B6 (DNAJB6), efficiently inhibits amyloid aggregation. This inhibition depends on a unique motif with conserved serine and threonine (S/T) residues that have a high capacity for hydrogen bonding. Global analysis of kinetics data has previously shown that DNAJB6 especially inhibits the primary nucleation pathways. These observations indicated that DNAJB6 achieves this remarkably effective and sub-stoichiometric inhibition by interacting not with the monomeric unfolded conformations of the amyloid-β symbol (Aβ) peptide but with aggregated species. However, these pre-nucleation oligomeric aggregates are transient and difficult to study experimentally. Here, we employed a native MS-based approach to directly detect oligomeric forms of Aβ formed in solution. We found that WT DNAJB6 considerably reduces the signals from the various forms of Aβ (1–40) oligomers, whereas a mutational DNAJB6 variant in which the S/T residues have been substituted with alanines does not. We also detected signals that appeared to represent DNAJB6 dimers and trimers to which varying amounts of Aβ are bound. These data provide direct experimental evidence that it is the oligomeric forms of Aβ that are captured by DNAJB6 in a manner which depends on the S/T residues. We conclude that, in agreement with the previously observed decrease in primary nucleation rate, strong binding of Aβ oligomers to DNAJB6 inhibits the formation of amyloid nuclei.

Pharmacologic dissection of the overlapping impact of heat shock protein family members on platelet function

BackgroundPlatelets play a pivotal role in hemostasis, wound healing, and inflammation, and are thus implicated in a variety of diseases, including cancer. Platelet function is associated with release of granule content, cellular shape change, and upregulation of receptors that promote establishment of a thrombus and maintenance of hemostasis. ObjectivesThe role of heat shock proteins (Hsps) in modulating platelet function has been studied for a number of years, but comparative roles of individual Hsps have not been thoroughly examined. MethodsWe utilized a panel of specific inhibitors of Hsp40, Hsp70, Hsp90, and Grp94 (the endoplasmic reticulum homolog of Hsp90) to assess their impact on several aspects of platelet function. ResultsInhibition of each of the aforementioned Hsps reduced alpha granule release. In contrast, there was some selectivity in impacts on dense granule release. Thromboxane synthesis was impaired after exposure to inhibitors of Hsp40, Hsp90, and Grp94, but not after inhibition of Hsp70. Both expression of active glycoprotein IIb/IIIa (GPIIb/IIIa) and fibrinogen‐induced platelet shape change were diminished by our inhibitors. In contrast, aggregation was selectively abrogated after inhibition of Hsp40 or Hsp90. Lastly, activated platelet–cancer cell interactions were reduced by inhibition of both Hsp70 and Grp94. ConclusionsThese data suggest the importance of Hsp networks in regulating platelet activity.

Nutrigenomic Effects of Long-Term Grape Pomace Supplementation in Dairy Cows

Simple SummaryThe aim of this study was to evaluate the effect of grape pomace (GP), the polyphenol-rich agricultural by-product, on dairy cows’ whole-blood transcriptome, milk production and composition. Twelve lactating Holstein-Friesian cows were randomly assigned to two groups; the first received a GP-supplemented diet for 60 days (group GP), whereas the second was given only a basal diet (CTR). The results reveal 40 protein-coding genes differentially expressed in the GP group when compared with the CTR group, but no effects were noticed on milk production, concentrations of crude protein, fat, casein, lactose and urea, or somatic cell count. Compared to CTR, GP had a transcriptomic signature mainly reflecting a reinforced immunogenic response.The increasing demand for more animal products put pressure on improving livestock production efficiency and sustainability. In this context, advanced animal nutrition studies appear indispensable. Here, the effect of grape pomace (GP), the polyphenol-rich agricultural by-product, was evaluated on Holstein-Friesian cows’ whole-blood transcriptome, milk production and composition. Two experimental groups were set up. The first one received a basal diet and served as a control, while the second one received a 7.5% GP-supplemented diet for a total of 60 days. Milk production and composition were not different between the group; however, the transcriptome analysis revealed a total of 40 genes significantly affected by GP supplementation. Among the most interesting down-regulated genes, we found the DnaJ heat-shock protein family member A1 (DNAJA1), the mitochondrial fission factor (MFF), and the impact RWD domain protein (IMPACT) genes. The gene set enrichment analysis evidenced the positive enrichment of ‘interferon alpha (IFN-α) and IFN-γ response’, ‘IL6-JAK-STAT3 signaling’ and ‘complement’ genes. Moreover, the functional analysis denoted positive enrichment of the ‘response to protozoan’ and ‘negative regulation of viral genome replication’ biological processes. Our data provide an overall view of the blood transcriptomic signature after a 60-day GP supplementation in dairy cows which mainly reflects a GP-induced immunomodulatory effect.

The Predicted Key Molecules, Functions, and Pathways That Bridge Mild Cognitive Impairment (MCI) and Alzheimer's Disease (AD).

To elucidate the key molecules, functions, and pathways that bridge mild cognitive impairment (MCI) and Alzheimer's disease (AD), we investigated open gene expression data sets. Differential gene expression profiles were analyzed and combined with potential MCI- and AD-related gene expression profiles in public databases. Then, weighted gene co-expression network analysis was performed to identify the gene co-expression modules. One module was significantly negatively associated with MCI samples, in which gene ontology function and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis showed that these genes were related to cytosolic ribosome, ribosomal structure, oxidative phosphorylation, AD, and metabolic pathway. The other two modules correlated significantly with AD samples, in which functional and pathway enrichment analysis revealed strong relationships of these genes with cytoplasmic ribosome, protein binding, AD, cancer, and apoptosis. In addition, we regarded the core genes in the module network closely related to MCI and AD as bridge genes and submitted them to protein interaction network analysis to screen for major pathogenic genes according to the connectivity information. Among them, small nuclear ribonucleoprotein D2 polypeptide (SNRPD2), ribosomal protein S3a (RPS3A), S100 calcium binding protein A8 (S100A8), small nuclear ribonucleoprotein polypeptide G (SNRPG), U6 snRNA-associated Sm-like protein LSm3 (LSM3), ribosomal protein S27a (RPS27A), and ATP synthase F1 subunit gamma (ATP5C1) were not only major pathogenic genes of MCI, but also bridge genes. In addition, SNRPD2, RPS3A, S100A8, SNRPG, LSM3, thioredoxin (TXN), proteasome 20S subunit alpha 4 (PSMA4), annexin A1 (ANXA1), DnaJ heat shock protein family member A1 (DNAJA1), and prefoldin subunit 5 (PFDN5) were not only major pathogenic genes of AD, but also bridge genes. Next, we screened for differentially expressed microRNAs (miRNAs) to predict the miRNAs and transcription factors related the MCI and AD modules, respectively. The significance score of miRNAs in each module was calculated using a hypergeometric test to obtain the miRNApivot-Module interaction pair. Thirty-four bridge regulators were analyzed, among which hsa-miR-519d-3p was recognized as the bridge regulator between MCI and AD. Our study contributed to a better understanding of the pathogenic mechanisms of MCI and AD, and might lead to the development of a new strategy for clinical diagnosis and treatment.

Cell Stress Induced Stressome Release Including Damaged Membrane Vesicles and Extracellular HSP90 by Prostate Cancer Cells.

Tumor cells exhibit therapeutic stress resistance-associated secretory phenotype involving extracellular vesicles (EVs) such as oncosomes and heat shock proteins (HSPs). Such a secretory phenotype occurs in response to cell stress and cancer therapeutics. HSPs are stress-responsive molecular chaperones promoting proper protein folding, while also being released from cells with EVs as well as a soluble form known as alarmins. We have here investigated the secretory phenotype of castration-resistant prostate cancer (CRPC) cells using proteome analysis. We have also examined the roles of the key co-chaperone CDC37 in the release of EV proteins including CD9 and epithelial-to-mesenchymal transition (EMT), a key event in tumor progression. EVs derived from CRPC cells promoted EMT in normal prostate epithelial cells. Some HSP family members and their potential receptor CD91/LRP1 were enriched at high levels in CRPC cell-derived EVs among over 700 other protein types found by mass spectrometry. The small EVs (30–200 nm in size) were released even in a non-heated condition from the prostate cancer cells, whereas the EMT-coupled release of EVs (200–500 nm) and damaged membrane vesicles with associated HSP90α was increased after heat shock stress (HSS). GAPDH and lactate dehydrogenase, a marker of membrane leakage/damage, were also found in conditioned media upon HSS. During this stress response, the intracellular chaperone CDC37 was transcriptionally induced by heat shock factor 1 (HSF1), which activated the CDC37 core promoter, containing an interspecies conserved heat shock element. In contrast, knockdown of CDC37 decreased EMT-coupled release of CD9-containing vesicles. Triple siRNA targeting CDC37, HSP90α, and HSP90β was required for efficient reduction of this chaperone trio and to reduce tumorigenicity of the CRPC cells in vivo. Taken together, we define “stressome” as cellular stress-induced all secretion products, including EVs (200–500 nm), membrane-damaged vesicles and remnants, and extracellular HSP90 and GAPDH. Our data also indicated that CDC37 is crucial for the release of vesicular proteins and tumor progression in prostate cancer.

Genomewide analysis of circular RNA in pituitaries of normal and heat-stressed sows

BackgroundAs a newly characterized type of noncoding RNA, circular RNA (circRNA) has been shown to have functions in diverse biological processes of animals. It has been reported that several noncoding RNAs may regulate animals’ response to heat stress which can be easily induced by hyperthermia in summer. However, the expression and functions of circRNAs in the pituitary of sows and whether they participate in heat stress adaption are still unclear.ResultsIn this study, we found that high temperature over the thermoneutral zone of sows during the summer increased the serum heat shock protein 70 (HSP70) level, decreased the superoxide dismutase (SOD) vitality and prolactin (PRL) concentration, and induced heat stress in sows. Then, we explored circRNA in the pituitary of heat-stressed and normal sows using RNA sequencing and bioinformatics analysis. In total, 12,035 circRNAs were detected, with 59 circRNAs differentially expressed, including 42 up-regulated and 17 down-regulated circRNAs in pituitaries of the heat-stressed sows. Six randomly selected circRNAs were identified through reverse transcription PCR followed by DNA sequencing and other 7 randomly selected differentially expressed circRNAs were verified by quantitative real-time PCR analysis. The predicted target genes regulated by circRNAs through sponging microRNAs (miRNAs) were enriched in metabolic pathway. Furthermore, the predicted circRNA–miRNA–mRNA interactions showed that some circRNAs might sponge miRNAs to regulate pituitary-specific genes and heat shock protein family members, indicating circRNA’s roles in pituitary hormone secretion and heat stress response.ConclusionsOur results provided a meaningful reference to understand the functions of circRNA in the porcine pituitary and the mechanisms by which circRNA may participate in animals’ response to heat stress.