• 한국동물위생학회지
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• 제27권4호
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• pp.355-369
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• 2004

The equine herpesvirus type 1 (EHV-1) immediate-early (IE) protein is a potent transactivator responsible for the activation of both early and late genes during the course of infection and is comprised of discrete functional domains that mediate its many functions. Interaction between trans activators such as the IE protein and various components of the RNA polymerase II transcription initiation machinery has been demonstrated to be critical for transactivation. In the present report, it is addressed the hypothesis that the IE protein interacts with various components of transcription machinery to mediate transactivation of target viral genes. In these studies, it is demonstrated that in vitro transcribed and translated IE protein interacts with TFIIB-agarose conjugate but not with TFIID-agarose conjugate. Additional immunoprecipitation studies using nuclear extracts derived from EHV-1 infected RK-13 cells confirmed that the IE protein interacts strongly with TFIIB, but fails to interact with TFIID. IR2, a truncated form of the IE protein lacking the potent transactivation domain and involved in the down-regulation of the IE gene, also interacted with TFIIB but not with TFIID. Studies were also performed to ascertain if particular TBP-associated factors (TAFs) could mediate IE or IR2 binding to TFIID. In vitro transcribed and translated TAF250 added to nuclear extracts generated from EHV-1 infected cells also failed to mediate an interaction between the IE protein or the IR2 protein and TFIID. This study demonstrated that the IE protein mediates transactivation of target viral genes by a mechanism that involves TFIIB. This is in contrast to mechanisms that have been proposed for both the herpes simplex virus ICP4 and VP16 protein which have been proposed to transactivate viral genes through interactions involving both TFIIB and TFIID. This study also intimates that IR2 mediate its repressive effects during the course of EHV-1 infection by a mechanism that involves sequestration of various transcription factors.

• 한국동물위생학회지
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• 제25권4호
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• pp.333-346
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• 2002

We previously reported that the equine herpesvirus type 1(EHV-1) immediate-early protein(IE protein) physically interacts with the general transcription factor human TFIIB(Jang et al, J Virol 75：10219-10230, 2001). The interaction between the IE protein and TFIIB is necessary for the IE protein to efficiently transactivate the early TK and late IR5 EHV-1 promoters. A panel of deletion and truncation mutants of the TFIIB gene was constructed and employed in protein-binding assays to map the IE protein-binding domain within TFIIB. Evidence is presented that the first direct repeat of TFIIB interacts specifically with the EHV-1 IE protein.

• Korean Circulation Journal
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• 제48권11호
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• pp.1014-1024
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• 2018

Background and Objectives: Intense exercise (IE) induced myocardial fibrosis (MF) showed contradictory findings in human studies, making the relationship between IE and the development of MF unclear. This study aims to demonstrate exercise induced MF is associated with cardiac damage, and inflammation is essential to the development of exercise induced MF. Methods: Sprague-Dawley rats were submitted to daily 60-minutes treadmill exercise sessions at vigorous or moderate intensity, with 8-, 12-, and 16-week durations; time-matched sedentary rats served as controls. Enzyme-linked immunosorbent assay (ELISA) was used to measure serum cardiac troponin I (cTnI) concentration. After completion of the exercise protocol rats were euthanized. Biventricular morphology, ultrastructure, and collagen deposition were then examined. Protein expression of interleukin $(IL)-1{\beta}$ and monocyte chemotactic protein (MCP)-1 was evaluated in both ventricles. Results: After IE, right but not left ventricle (LV) MF occurred. Serum cTnI levels increased and right ventricular damage was observed at the ultrastructure level in rats that were subjected to long-term IE. Leukocyte infiltration into the right ventricle (RV) rather than LV was observed after long-term IE. Long-term IE also increased protein expression of proinflammation factors including $IL-1{\beta}$ and MCP-1 in the RV. Conclusions: Right ventricular damage induced by long-term IE is pathological and the following inflammatory response is essential to the development of exercise induced MF.

• BMB Reports
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• 제35권6호
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• pp.553-561
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• 2002

A gene that encodes a protein homologue to baculoviral IE-1 was identified and sequenced in the genome of the Choristoneura fumiferana granulovirus (ChfuGV). The gene has an 1278 nucleotide (nt) open-reading frame (ORF) that encodes 426 amino acids with an estimated molecular weight of 50.33 kDa. At the nucleotide level, several cis-acting regulatory elements were detected within the promoter region of the ie-1 gene of ChfuGV along with other studied granuloviruses (GVs). Two putative CCAAT elements were detected within the noncoding leader region of this gene; one was located on the opposite strand at -92 and the other at -420 nt from the putative start triplet. Two baculoviral late promoter motifs (TAAG) were also detected within the promoter region of the ie-1 gene of ChfuGV. A single polyadenylation signal, AATAAA, was located 18nt downstream of the putative translational stop codon of ie-1 from ChfuGV. At the protein level, the amino acid sequence data that was derived from the nucleotide sequence in ChfuGV IE-1 was compared to those of the Cydia pomonella granulovirus (CpGV), Xestia c-nigrum granulovirus (XcGV) and Plutella xylostella granulovirus (PxGV). The C-terminal regions of the granuloviral IE-1 sequences appeared to be more conserved when compared to the N-terminal regions. A domain, similar to the basic helix-loop-helix like (bHLH-like) domain in NPVs, was detected at the C-terminal region of IE-1 from ChfuGV (residues 387 to 414). A phylogenetic tree for baculoviral IE-1 was constructed using a maximum parsimony analysis. A phylogenetic estimation demonstrates that ChfuGV IE-1 is most closely related to that of CpGV.

• Journal of Microbiology and Biotechnology
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• 제24권12호
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• pp.1719-1727
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• 2014

In the present study, whispovirus immediate early 1 promoter (ie-1) was used to initiate surface expression of the hemagglutinin (HA) protein of Egyptian H5N1 avian influenza virus (AIV) by using the baculovirus expression vector system. The HA gene and whispovirus ie-1 promoter sequence were synthesized as a fused expression cassette (ie1-HA) and successfully cloned into the pFastBac-1 transfer vector. The recombinant vector was transformed into DH10Bac competent cells, and the recombinant bacmid was generated via site-specific transposition. The recombinant bacmid was used for transfection of Spodoptera frugiperda (Sf-9) insect cells to construct the recombinant baculovirus and to induce expression of the HA protein of H5N1 AIV. The recombinant glycoprotein expressed in Sf-9 cells showed hemadsorption activity. Hemagglutination activity was also detected in both extra- and intracellular recombinant HAs. Both the HA and hemadsorption activities were inhibited by reference polyclonal anti-H5 sera. Significant expression of the recombinant protein was observed on the surface of infected insect cells by using immunofluorescence. SDS-PAGE analysis of the expressed protein revealed the presence of a visually distinguishable band of ~63 kDa in size, which was absent in the non-infected cell control. Western blot analysis confirmed that the distinct 63 kDa band corresponded to the recombinant HA glycoprotein of H5N1 AIV. This study reports the successful expression of the HA protein of H5N1 AIV. The expressed protein was displayed on the plasma membrane of infected insect cells under the control of whispovirus ie-1 promoter by using the baculovirus expression vector system.

• Journal of Microbiology and Biotechnology
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• 제16권8호
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• pp.1169-1173
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• 2006

Imaging ellipsometry (IE) for detection of binding of Escherichia coli O157:H7 (E. coli O157:H7) to an immunosensor is reported. A protein G layer, chemically bound to a self-assembled layer of 11-mercaptoundecanoic acid (11-MUA), was adopted for immobilization of monoclonal antibody against E. coli O157:H7 (Mab). The immobilization of antibody was investigated using surface plasmon resonance. To fabricate antibody spots on a gold surface, protein G solution was spotted onto the gold surface modified with an 11-MUA layer, followed by immobilizing Mab on the protein G spot. Ellipsometric images of the protein G spot, the Mab spot, and Mab spots with binding of E. coli O157:H7 in various concentrations were acquired using the IE system. The change of mean optical intensity of the Mab spots in the ellipsometric images indicated that the lowest detection limit was $10^3$CFU/ml for E. coli O157:H7. Thus, IE can be applied to an immunosensor for detection of E. coli O157:H7 as a detection method with the advantages of allowing label-free detection, high sensitivity, and operational simplicity.

• 대한바이러스학회지
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• 제28권3호
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• pp.267-274
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• 1998

Human cytomegalovirus (HCMV) has the ability to activate the expression of many viral and cellular genes. Among various viral proteins, the immediate early proteins (IE1-72kDa, IE2-86kDa) have been known to be potent transactivators. The product of c-jun proto-oncogene is important in cell activation and differentiation. Here, we tried to find out if the IE could activate the c-jun promoter and also tried to identify the responsible sequence elements in the c-jun activation by IE1-72kDa. We found HCMV IE expression transactivated the c-jun promoter in human embryonal lung fibroblasts (HEL). The activation fold by IE1-72kDa, IE2-86kDa and IE2-55kDa was 23, 35, and 5, respectively. When the expression of each IE was combined, it showed synergism. Expression of (IE1-72kDa + IE2-86kDa) and (IE1-72kDa + IE2-86kDa + IE2-55kDa) resulted in 131 and 162 fold increase, respectively. The c-jun promoter region between -117 and -59 contains binding sites for the transcription factors Spl, CAAT, AP-l like (ATF/CREB), and MEF2. Transient expression assays were performed using various reporter plasmids containing the c-jun promoter-regulatory region linked to the luciferase gene and a plasmid expressing HCMV IE1 gene. Deletional and point mutational analysis showed that the sequence between -225 to -160 and the CTF binding site were involved in the up-regulation of c-jun promoter.

• International Journal of Industrial Entomology and Biomaterials
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• 제3권1호
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• pp.43-49
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• 2001

We have cloned and characterized an immediate early-1 gene, iel, which is activated immediately upon entrance of the viral genome into the cell nucleus, from Bombyx mori nuclear polyhedrosis virus (BmNPV) K1 strain. This gene encodes a protein 584 amino acids with a predicted molecular weight of 67 kDa. The promoter and coding regions of BmNPV-K1 ie1 showed high homology with Autographa californica nuclear polyhedrosis virus and BmNPV T3 strain. The BmNPV-K1 ie1 was different from amino acid sequence at 4 positions in BmNPV T3. The location of ie1 gene in the BmNPV-K1 genome was confirmed by Southern blot analysis and its expression patterns at the transcriptional level in the infected cells were confirmed by Nerthern hybridization analysis.

• International Journal of Industrial Entomology and Biomaterials
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• 제15권2호
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• pp.131-135
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• 2007

Open reading frame 4 of Bombyx mori nucleopolyhedrovirus (BmNPV), designated as Bm4, is a gene whose function is completely unknown. With the recently developed BmNPV bacmid and a modified pFastBac1 whose polyhedrin promoter was replaced with ie1 promoter, a recombinant bacmid expressing Bm4-EGFP fusion protein under the control of ie1 promoter in BmN cells was successfully constructed. The result not only showed that the polyhedrin promoter can be replaced efficiently with other promoters to direct the expression of foreign gene in BmN cells by using Bac-to-Bac/BmNPV baculovirus expression system but also laid the foundation for rescue experiment of Bm4 deletion mutant due to the ability of ie1 promoter to direct gene expression throughout the infection cycle.

• 한국미생물학회:학술대회논문집
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• 한국미생물학회 2005년도 International Meeting of the Microbiological Society of Korea
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• pp.219.3-219
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• 2005