34 DIFFERENTIALLY EXPRESSED PROTEINS OF EARLY-STAGE PLACENTA DERIVED FROM CLONED CAT EMBRYOS USING PROTEOMICS ANALYSIS

2012 ◽  
Vol 24 (1) ◽  
pp. 129
Author(s):  
J. I. Bang ◽  
D. W. Bae ◽  
Y. S. Kwon ◽  
G. K. Deb ◽  
B. H. Choi ◽  
...  
Keyword(s):  
Heat Shock ◽  
Heat Shock Protein ◽  
Early Pregnancy ◽  
Protein Disulfide Isomerase ◽  
Triosephosphate Isomerase ◽  
Early Stage ◽  
Differentially Expressed ◽  
Differentially Expressed Proteins ◽  
Disulfide Isomerase ◽  
Protein Disulfide

We have previously demonstrated that differentially expressed proteins affect abnormal development and function of cloned term placenta. This is associated with cloned fetus morbidity and mortality. We also frequently observed loss of the cloned fetus and failed development during early pregnancy periods. To confirm the pattern of important gene expression in cloned placenta during pre- and post-implantation, we investigated expression pattern of proteins in early stage (21 days) domestic cat placentas of cloned embryo transfer (CEP; n = 2) and artificial insemination (CP; n = 4) derived pregnancy. The differentially expressed proteins were investigated by 2-DE and MALDI-TOF/MS. Twenty-three proteins were up- and down-regulated at least 1.5-fold in the CEP (P < 0.05) compared with the CP. Differentially expressed proteins were analysed using PDQest program and statistically analysed by 1-way ANOVA using the SPSS software. In CEP, 13 proteins were up-regulated, such as 78-kDa glucose-regulated protein (GRP78), annexin A2 (ANXA2), protein DJ-1 (DJ1), adenylate kinase isoenzyme 1 (AK1), protein disulfide-isomerase A3 (PDIA3), heat shock protein β-1 (HSPB1), actin, cytoplasmic 1 (ACTB), serum albumin (ALB), protein disulfide-isomerase A6 (PDIA6), G protein-regulated inducer of neurite outgrowth 1 (GRIN1) and triosephosphate isomerase (TIM). In contrast, 10 proteins were down-regulated, such as vinculin (VCL), triosephosphate isomerase (TIM), heterogeneous nuclear ribonucleoprotein H (hnRNPH), tropomyosin α-4 (TPM4), 60-kDa heat shock protein, mitochondrial (Hsp60), serum albumin (ALB), calumenin (CALU), keratin type 1 (CK1) and prohibitin (PHB). To validate the identified proteins in the CEP compared with the CP, we investigate a peptide sequences using MALDI-TOF/TOF tandem mass spectrometry. The sequence information obtained a high ions score from NCBI and Swiss-Prot databases. In conclusion, we did identify abnormal expression of proteins that might be associated with impaired development of CEP, which may endanger the cloned fetus during early pregnancy. This work was partly supported by the BK21 program and the KOSEF (10525010001-05N2501-00110) and the Next-generation BioGreen21 program (No. PJ007990012011).

Proteomic and mRNA Expression Chip Analysis of Acquired TRAIL-Resistance in Human HL60 Myeloid Leukemia Cells.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 4155-4155
Author(s):  
Jan Zivny ◽  
Pavel Klener ◽  
Ondrej Toman ◽  
Petr Halada ◽  
Robert Ivanek ◽  
...  
Keyword(s):  
Gene Expression ◽  
Protein Disulfide Isomerase ◽  
Expression Profiles ◽  
Differentially Expressed ◽  
Differentially Expressed Proteins ◽  
Disulfide Isomerase ◽  
Mrna Gene ◽  
Trail Resistance ◽  
Protein Disulfide ◽  
Mrna Gene Expression

Abstract Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL or Apo2L) is a proapoptotic cytokine implicated in cancer cell surveillance. TRAIL induces apoptosis of target cells by the extrinsic, receptor-mediated, apoptotic pathway. A potential of TRAIL as cancer-specific therapeutic agent has been proposed, either as a single agent or in combination with chemotherapy agents. Development of TRAIL-resistant clones in the originally TRAIL-sensitive tumor cell population may be a critical complication of TRAIL based cancer therapy. Prolonged exposure of TRAIL-sensitive leukemia cell line, “wild-type” (WT) HL60 cells, to recombinant soluble TRAIL resulted in the establishment of resistant subclones. To get deeper insight into the molecular mechanism of acquired TRAIL resistance we compared expression profiles of TRAIL-sensitive HL-60 cells and TRAIL-resistant HL-60 subclones using microarray mRNA gene expression chips and 2D-PAGE followed by MS and MS/MS analysis. Using cluster analysis of microarray mRNA gene expression profiles of 6 individual HL60 resistant subclones we identified two molecular signatures of TRAIL resistance. When compared to WT HL60 TRAIL-sensitive mRNA expression the group of HL60 subclones designed as phenotype 1 (P1, n=3) had 1122 significantly upregulated and 985 significantly downregulated mRNAs and subclones designed as phenotype 2 (P2, n=3) had 1583 significantly upregulated and 1798 significantly downregulated mRNAs (p<0.05). The subclones of P1 and P2 phenotype shared 533 upregulated and 422 downregulated genes compared to HL60 WT cells. To identify differentially expressed proteins we performed 2D-PAGE of two representative subclones of P1 and P2 phenotypes (in five replicates) and compared them with HL60 WT cells. We detected and identified differentially expressed proteins involved in various aspects of cellular metabolism. Except for downregulation of chromobox protein homolog 5, a protein involved in the regulation of gene expression, and downregulation of Annexin A6 and protein disulfide-isomerase A3 precursor the TRAIL-resistant P1 and P2 subclones showed different protein expression profile. In P1 TRAIL-resistant cells we identified downregulation of proteins involved in energy metabolism, including pyruvate kinase, enolase, cytochrome c-reductase, NADH dehydrogenase and downregulation of proteins involved in proteasome activity, such as proteasome activator complex subunits 1 and 2. In P2 TRAIL-resistant cells we detected downregulation of important regulatory proteins, such as DNA replication licensing factor MCM7, proliferation-associated protein 2G4, replication protein A 32 kDa subunit. In both, P1 and P2, subclones we identified significant changes in cytoskeletal rearrangement proteins. In addition, several chaperones were differentially expressed, namely protein-disulfide isomerase, GrpE, HSC71 and HSP27, 78 kDa glucose-regulated protein precursor. Our data show two HL60 cell line-derived TRAIL resistant subclones, P1 and P2, have specific molecular signatures suggesting distinct mechanisms of TRAIL resistance. These distinct pathways for development of resistance to TRAIL-induced apoptosis are relevant for design of more effective strategies for leukemia therapy.

scholarly journals Protein Disulfide Isomerase Inhibitor Suppresses Viral Replication and Production during Antibody-Dependent Enhancement of Dengue Virus Infection in Human Monocytic Cells

Viruses ◽  
2019 ◽  
Vol 11 (2) ◽  
pp. 155 ◽  
Author(s):  
Nantapon Rawarak ◽  
Aroonroong Suttitheptumrong ◽  
Onrapak Reamtong ◽  
Kobporn Boonnak ◽  
Sa-nga Pattanakitsakul
Keyword(s):  
Dengue Virus ◽  
Protein Disulfide Isomerase ◽  
Early Stage ◽  
Secondary Infection ◽  
Protein Translation ◽  
Virus Infectivity ◽  
Disulfide Isomerase ◽  
Antibody Dependent Enhancement ◽  
Altered Proteins ◽  
Protein Disulfide

One of several mechanisms that leads to the development of dengue hemorrhagic fever (DHF) and dengue shock syndrome (DSS) is called antibody-dependent enhancement (ADE). Monocytes can be infected by the ADE phenomenon, which occurs in dengue secondary infection. This study aimed to investigate the proteins involved in ADE of DENV infection in the human monocytic cell line U937. The phosphoproteins were used to perform and analyze for protein expression using mass spectrometry (GeLC-MS/MS). The differential phosphoproteins revealed 1131 altered proteins compared between isotype- and DENV-specific antibody-treated monocytes. The altered proteins revealed 558 upregulated proteins and 573 downregulated proteins. Protein disulfide isomerase (PDI), which is an enzyme that had a high-ranking fold change and that catalyzes the formation, breakage, and rearrangement of disulfide bonds within a protein molecule, was selected for further study. PDI was found to be important for dengue virus infectivity during the ADE model. The effect of PDI inhibition was also shown to be involved in the early stage of life cycle by time-of-drug-addition assay. These results suggest that PDI is important for protein translation and virion assembly of dengue virus during infection in human monocytes, and it may play a significant role as a chaperone to stabilize dengue protein synthesis.

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