OBJECTIVETo investigate the expression and significance of HSP27, HSP60, HSP70 and HSP90 alpha in esophageal squamous cell carcinoma (ESCC) and tissues along the incision margin (TIM).

METHODSThe presence and the level of expression of HSP27, HSP60, HSP70 and HSP90 alpha were determined in 168 specimens from ESCC and 42 from tissues along TIM by EnVision immunohistochemistry and Western blotting, to compare their positive staining rates and explore the correlation between their expressions and clinicopathologic features in ESCC.

RESULTSThe positive staining rates of HSP27, HSP60, HSP70 and HSP90 alpha in ESCC and TIM were 62.0% and 42.1%, 92.7% and 63.2%, 57.9% and 22.2%, and 33.7% and 18.5%, respectively. There was very significant difference between the expression of HSP60 and HSP70 in ESCC and TIM (P < 0.01), but not significant about HSP27 and HSP90 alpha (P > 0.05). The positive staining rate of HSP27 declined with the lower grade of differentiation of ESCC (P < 0.05).

CONCLUSIONThe present findings suggest that the expression of HSPs of different molecular weight in ESCC and TIM is a common event. The level of expressions of HSP60 and HSP70 are higher than those in TIM. HSP60 and HSP70 expression correlated with the biological behavior of ESCC. The expression of HSP27 was positively correlated to the grade of differentiation of ESCC. Overexpression of HSP27 may be associated to the differentiation of squamous cell carcinoma.

Blotting, Western ; Carcinoma, Squamous Cell ; metabolism ; pathology ; Cell Differentiation ; Chaperonin 60 ; metabolism ; Chi-Square Distribution ; Esophageal Neoplasms ; metabolism ; pathology ; Esophagus ; chemistry ; pathology ; HSP27 Heat-Shock Proteins ; HSP70 Heat-Shock Proteins ; metabolism ; HSP90 Heat-Shock Proteins ; metabolism ; Heat-Shock Proteins ; metabolism ; Humans ; Immunohistochemistry ; Lymphatic Metastasis ; Neoplasm Invasiveness ; Neoplasm Proteins ; metabolism

OBJECTIVETo explore the mechanism of podophyllotoxin nanostructured lipid carriers (POD-NLC)-induced apoptosis of VK2/E6E7 cells mediated by endoplasmic reticulum stress (ERS).

METHODSVK2/E6E7 cells cultured in vitro were exposed to 0.125, 0.25, and 0.5 µg/ml POD-NLC or blank NLC for 24 h. The intracellular calcium concentration was measured by laser scanning confocal microscopy (LSCM), and the expression levels of GRP78, GRP94, and calpain2 mRNA and proteins in the cells were detected using RT-PCR and Western blotting.

RESULTSCompared with the control cells, the cells exposed to POD-NLC showed a concentration-dependent increase of intracellular calcium concentration (P<0.01), and the differences were statistically significant between different dose groups (P<0.05). RT-PCR and Western blotting showed that POD-NLC up-regulated GRP78, GRP94 and calpain2 mRNA and proteins expressions, which showed significant differences between blank-NLC and the control groups (P>0.05).

CONCLUSIONPOD-NLC induces apoptosis of VK2/E6E7 cells possibly by triggering the endoplasmic reticulum stress response.

Apoptosis ; Calcium ; metabolism ; Calpain ; metabolism ; Cell Line ; Endoplasmic Reticulum Stress ; HSP70 Heat-Shock Proteins ; metabolism ; Heat-Shock Proteins ; metabolism ; Humans ; Membrane Proteins ; metabolism ; Nanostructures ; chemistry ; Podophyllotoxin ; chemistry ; RNA, Messenger

OBJECTIVETo investigate lead distribution and the change of 78 000 glucose regulated protein (GRP78) in various organs of weaned rats challenged with low-level maternal origin lead.

METHODSMale littermates, bred from the female Fisher 344 rats gavaged with lead acetate or sodium acetate (1 ml of 10 mg/ml per day per animal) with male Fisher 344 rats without lead treatment, were divided into 4 groups including control (group A), gestation plus lactation (group B), gestation only (group C), and lactation only (group D). Each group had 6 litters. These littermates were weaned and terminated at postnatal day 21. Lead contents and GRP78 levels in various organs of these littermates were determined by atomic absorbance spectrometry (AAS) and Western blotting analysis, respectively.

RESULTSMaternal lead was observed to transfer to littermates through gestation and lactation. Concentrations of littermate blood lead in groups A to D were (0.0010+/-0.0010), (0.1420+/-0.0190), (0.0250+/-0.0040), and (0.1490+/-0.0160) microg/ml, respectively. Concentrations of littermate brain lead in groups A to D were (0.0005+/-0.0005), (0.1120+/-0.0130), (0.0125+/-0.0042), and (0.0700+/-0.0058) microg/g, respectively. Concentrations of littermate kidney lead in groups A to D were (0.0050+/-0.0050), (1.0400+/-0.1000), (0.1040+/-0.0330), and (0.9920+/-0.0850) microg/g, respectively. Concentrations of littermate liver lead in groups A to D were (0.0030+/-0.0050), (0.3600+/-0.0550), (0.0567+/-0.0126), and (0.3030+/-0.0310) microg/g, respectively. Blood, brain, kidney and liver lead concentrations in groups B and D were significantly higher than those in group C and differences were 5-10 folds. Arbitrary units of littermate leukocytic GRP78 concentration normalized with actin protein in groups A to D were 1.000+/-0.038, 1.180+/-0.060, 0.998+/-0.109, and 1.290+/-0.110, respectively. Arbitrary units of littermate brain GRP78 concentration normalized with actin protein level in groups A to D were 0.996+/-0.128, 0.922+/-0.246, 1.150+/-0.170, and 0.750+/-0.126, respectively.

CONCLUSIONLead in maternal bodies could be transferred to litter bodies through gestation and lactation and distributed in various organs. Lead might also changed GRP78 expression in leukocytes.

Animals ; Animals, Newborn ; Brain ; metabolism ; Female ; HSP70 Heat-Shock Proteins ; metabolism ; Kidney ; chemistry ; Lead ; metabolism ; Leukocytes ; metabolism ; Liver ; chemistry ; Male ; Membrane Proteins ; metabolism ; Pregnancy ; Rats ; Rats, Inbred F344

Protein palmitoylation, one of post-translation modifications, refers to the addition of saturated 16-carbon palmitic acid to cysteine residues via the thioester bond. It plays key roles in various functional activities, such as the interaction, stability and location of proteins. Heat shock protein 90 (Hsp90), an important molecular chaperone, has been reported to be involved in sperm capacitation. However, it remains unclear whether protein palmitoylation exists in sperm and whether Hsp90 in sperm is palmitoylated under different physiological conditions. In this study, we examined whether the protein palmitoylation is present in mouse cauda epididymis sperm using acyl-biotin exchange method, predicted the potential palmitoylated sites of Hsp90 by the software CSS-Palm 4.0 and detected the palmitoylated Hsp90 in the mouse sperm from caput epididymis and cauda epididymis by immunoprecipitation. We found that some proteins, approximately 50, 65, 72, 85 and 130 kDa, were palmitoylated in mouse cauda epididymis sperm. Five sites in two Hsp90 isoforms were predicted to be palmitoylated. The results also showed that Hsp90 in mouse sperm was palmitoylated and its palmitoylation level was involved in different physiological conditions: the palmitoylation level of cauda epididymis sperm was higher than that of caput epididymis sperm; and the palmitoylation level after capacitation was much higher than that before capacitation. In conclusion, this study reveals that protein palmitoylation is present in mouse sperm and the palmitoylated Hsp90 is associated with different physiological conditions in sperm.
Animals ; Epididymis ; HSP90 Heat-Shock Proteins ; metabolism ; Lipoylation ; Male ; Mice ; Palmitic Acid ; chemistry ; Sperm Capacitation ; Spermatozoa ; metabolism

Proteins fold into their functional 3-dimensional structures from a linear amino acid sequence. In vitro this process is spontaneous; while in vivo it is orchestrated by a specialized set of proteins, called chaperones. Protein folding is an ongoing cellular process, as cellular proteins constantly undergo synthesis and degradation. Here emerging links between this process and cancer are reviewed. This perspective both yields insights into the current struggle to develop novel cancer chemotherapeutics and has implications for future chemotherapy discovery.
Amino Acid Sequence ; Heat-Shock Proteins ; metabolism ; Homeostasis ; Humans ; Molecular Chaperones ; chemistry ; metabolism ; Molecular Targeted Therapy ; Neoplasms ; drug therapy ; metabolism ; Protein Folding ; Proteins ; chemistry ; metabolism ; Proteolysis

This article is to summarize the molecular and functional analysis of the gene "suppression of tumorigenicity 13" (ST13). ST13 is in fact the gene encoding Hsp70 interacting protein (Hip), a co-factor (co-chaperone) of the 70-kDa heat shock proteins (Hsc/Hsp70). By collaborating with other positive co-factors such as Hsp40 and the Hsp70-Hsp90 organizing protein (Hop), or competing with negative co-factors such as Bcl2-associated athanogen 1 (Bag1), Hip facilitates may facilitate the chaperone function of Hsc/Hsp70 in protein folding and repair, and in controlling the activity of regulatory proteins such as steroid receptors and regulators of proliferation or apoptosis. Although the nomenclature of ST13 implies a role in the suppression of tumorigenicity (ST), to date available experimental data are not sufficient to support its role in cancer development, except for the possible down-regulation of ST13 in gastric and colorectal cancers. Further investigation of this gene at the physiological level would benefit our understanding of diseases such as endocrinological disorders, cancer, and neurodegeneration commonly associated with protein misfolding.
Adenosine Triphosphate ; metabolism ; Animals ; Carrier Proteins ; chemistry ; genetics ; physiology ; Cloning, Molecular ; HSP70 Heat-Shock Proteins ; metabolism ; Humans ; Protein Folding ; Tumor Suppressor Proteins ; chemistry ; genetics ; physiology

OBJECTIVETo investigate HSP70 gene expression from Dendrobium officinale under low temperature stress, which will provide the molecular biological foundation for breeding the low temperature resistant strain.

METHODHSP70 gene full length cDNA was cloned by rapid amplification of cDNA ends (RACE) on the basis of HSP70 gene fragment sequences, and the structure and function of HSP70 gene were deduced. The expression of HSP70 under low temperature stress was detected by RT-PCR.

RESULTThe full length of HSP70 gene cDNA was 2 296 bp containing a 1 944 bp open reading frame (ORF) that encoded a protein of 647 amino acids. Its amino acids sequence had typical HSP70 characteristics and high homology with other plant's HSP70. Cold stress expression analysis showed that expression of the HSP70 gene could be induced by low temperature.

CONCLUSIONThe HSP70 gene of D. officinale was successfully cloned and reported for the first time which proved that the expression could be induced by low temperature. The cloning of HSP70 gene provides a stable foundation for further study of D. officinale cultivation and the breeding of the cold resistance strains.

Amino Acid Sequence ; Cloning, Molecular ; Cold Temperature ; Dendrobium ; classification ; genetics ; metabolism ; HSP70 Heat-Shock Proteins ; chemistry ; genetics ; metabolism ; Molecular Sequence Data ; Plant Proteins ; chemistry ; genetics ; metabolism ; Sequence Alignment

OBJECTIVETo study the role of a BTB/POZ domain protein in the expression of hsp90alpha gene.

METHODSThe eukaryotic expression plasmids of sense- and antisense-GAGA related protein (GRP) or empty vector were transfected into Jurkat cells with pREP4 episomal vector plasmids carrying the hsp90alpha promoter sequence from -1756 to +37 and control plasmids pMCAT. Total RNA was extracted. The relative promoter activity of hsp90alpha-CAT reporter gene was determined by competitive RT-PCR assay.

RESULTSGRP markly increased the relative promoter activity of hsp90alpha-CAT reporter gene during heat shock.

CONCLUSIONGRP may promote the expression of hsp90alpha gene by participating in chromatin remolding.

Amino Acid Motifs ; Animals ; Checkpoint Kinase 1 ; Cloning, Molecular ; DNA ; metabolism ; DNA, Complementary ; metabolism ; DNA-Binding Proteins ; metabolism ; Drosophila ; genetics ; Drosophila Proteins ; genetics ; Gene Expression Regulation ; HSP90 Heat-Shock Proteins ; biosynthesis ; genetics ; Heat-Shock Response ; genetics ; Homeodomain Proteins ; chemistry ; genetics ; metabolism ; Humans ; Repressor Proteins ; genetics ; Transcription Factors ; genetics ; Two-Hybrid System Techniques

In response to viral infection, RIG-I-like RNA helicases detect viral RNA and signal through the mitochondrial adapter protein VISA. VISA activation leads to rapid activation of transcription factors IRF3 and NF-κB, which collaborate to induce transcription of type I interferon (IFN) genes and cellular antiviral response. It has been demonstrated that VISA is activated by forming prion-like aggregates. However, how this process is regulated remains unknown. Here we show that overexpression of HSC71 resulted in potent inhibition of virus-triggered transcription of IFNB1 gene and cellular antiviral response. Consistently, knockdown of HSC71 had opposite effects. HSC71 interacted with VISA, and negatively regulated virus-triggered VISA aggregation. These findings suggest that HSC71 functions as a check against VISA-mediated antiviral response.
Adaptor Proteins, Signal Transducing ; biosynthesis ; chemistry ; genetics ; metabolism ; Cell Aggregation ; genetics ; GPI-Linked Proteins ; metabolism ; Gene Knockdown Techniques ; HEK293 Cells ; HSC70 Heat-Shock Proteins ; genetics ; metabolism ; Heat-Shock Response ; genetics ; Humans ; Interferon Regulatory Factor-3 ; genetics ; metabolism ; Interferon-beta ; genetics ; NF-kappa B ; genetics ; Prions ; metabolism ; Receptors, Retinoic Acid ; metabolism ; Viruses ; drug effects ; metabolism ; pathogenicity

Although two chain transfering separately could be used to overcome the volume limitation of adeno-associated virus vectors (AAV) in coagulation factor VIII (FVIII) gene delivery, it leads to chain imbalance for inefficient heavy chain secretion. In this study we aimed to improve the efficacy of two chain strategy in FVIII gene delivery through the degradation of glucose-regulated protein 78 (GRP78) known as a protein chaperone in endoplasmic reticulum (ER) by deoxynivalenol (DON) to decrease GRP78-bound FVIII heavy chain. By treating the two-chain gene transduced 293 cells with DON, the heavy chain (HC) secretion and FVIII bioactivity were observed. Data showed that 293 cells after three hours post-treatment with DON at a concentration of 500 ng mL(-1) resulted in obvious decrease the level of GRP78 but no effect on the cell proliferation. The HC secreted from DON-treated cells transfected with HC gene alone was 59 +/- 11 ng mL(-1), higher than that secreted by control cells (15 +/- 4 ng mL(-1)), and the HC secretion was further increasing to 146 +/- 34 ng mL(-1) in light chain (LC) gene co-transfected cells with an activity measured up to 0.66 +/- 0.15 U mL(-1), also greater than control cells (76 +/- 17 ng mL(-1) and 0.35 +/- 0.09 U mL(-1)). Taken together, these data suggest that DON-mediated GRP78 down-regulation could improve the efficacy of two-chain FVIII gene transfering by facilitating HC secretion, providing an experimental basis for in vivo dual-AAV application in FVIII gene delivery.
Cell Proliferation ; Down-Regulation ; Factor VIII ; chemistry ; genetics ; secretion ; Gene Transfer Techniques ; HEK293 Cells ; Heat-Shock Proteins ; metabolism ; Humans ; Transfection ; Trichothecenes ; pharmacology