• Title/Summary/Keyword: translation initiation

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Transcription, Translation, and Immunolocalization of ODVP-6E/ODV-E56 and p74 Proteins: Two Highly Conserved ODV-associated Envelope Proteins of Choristoneura fumiferana Granulovirus

  • Rashidan, Kianoush Khajeh;Nassoury, Nasha;Giannopoulos, Paresa N.;Guertin, Claude
    • BMB Reports
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    • v.38 no.1
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    • pp.65-70
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    • 2005
  • Choristoneura fumiferana granulovirus (ChfuGV) infection results two types of enveloped virions: Occlusion-derived virus (ODV) and budded virus (BV). Structural proteins ODVP-6E/ODV-E56 and p74 are two major conserved ODV-associated proteins that may be involved in the initiation of viral infection cycle in susceptible host insect larvae. This study presents the characterization of ChfuGV odvp-6e/odv-e56 and p74 transcription and translation as well as immunolocalization of these proteins in the occluded ChfuGV virion. Our results revealed that the transcription of odvp-6e/odv-e56 and p74 genes, both, start at 24 hours post infection (h p.i.). Using monospecific polyclonal antibodies made against ODVP-6E/ODV-E56 and p74 we demonstrated that these proteins are both expressed late in infection (24 h p.i.). Immunogold labeling using antisera against ODVP-6E/ODV-E56 and p74 proteins demonstrated that ODVP-6E/ODV-E56 and p74 proteins are both associated with the ODV envelop of ChfuGV.

Heterogeneous Sequences of Brain Cytoplasmic 200 RNA Formed by Multiple Adenine Nucleotide Insertions

  • Shin, Heegwon;Lee, Jungmin;Kim, Youngmi;Jang, Seonghui;Kim, Meehyein;Lee, Younghoon
    • Molecules and Cells
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    • v.42 no.6
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    • pp.495-500
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    • 2019
  • Brain cytoplasmic 200 RNA (BC200 RNA), originally identified as a neuron-specific non-coding RNA, is also observed in various cancer cells that originate from non-neural cells. Studies have revealed diverse functions of BC200 RNA in cancer cells. Accordingly, we hypothesized that BC200 RNA might be modified in cancer cells to generate cancerous BC200 RNA responsible for its cancer-specific functions. Here, we report that BC200 RNA sequences are highly heterogeneous in cancer cells by virtue of multiple adenine nucleotide insertions in the internal A-rich region. The insertion of adenine nucleotides enhances BC200 RNA-mediated translation inhibition, possibly by increasing the binding affinity of BC200 RNA for eIF4A (eukaryotic translation initiation factor 4A).

A Theoretical Investigation on the Generation of Strength in Staple Yarns

  • Ghosh Anindya
    • Fibers and Polymers
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    • v.7 no.3
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    • pp.310-316
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    • 2006
  • In this article, an attempt has been made to explain the failure mechanism of spun yams. The mechanism includes the aspects of generation and distribution of forces on a fibre under the tensile loading of a yam, the free body diagram of forces, the conditions for gripping and slipping of a fibre, and the initiation, propagation, and ultimate yam rupture in its weakest link. A simple mathematical model for the tenacity of spun yams has been proposed. The model is based on the translation of fibre bundle tenacity into the yam tenacity.

Expression of Recombinant Korean Mistletoe(KM) Lectin and B genes in Saccharomyces cerevisiae (Saccharomyces cerevisae에서 한국산 겨우살이 유래 lectin A 및 B 유전자의 발현)

  • 최윤혁;김종배;양웅석;황철원
    • Journal of Life Science
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    • v.14 no.5
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    • pp.840-846
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    • 2004
  • A study for expression of Korean Mistletoe (KM) lectin gene (A,B) in Saccharomyces cerevisiae was done using transforming system of yeast. In order to overexpress the genes efficiently in yeast, two lectin genes (A,B) were re-cloned and modified including Kozak translation initiation sequence using PCR amplification. The constructed plasmids containing modified lectin A and B genes were transformed to S. cerevisea INVSc (MAT G, his3 $\Delta$1, leu2, trpl-289, ura3-52). The transformed cells were identified by DNA sequencing with ABI3700 system and induced with 2% of galactose for recombinant KM lectin (rKM lectin) protein. The rKM lectin A and B proteins were determinated about 29kDa size of protein by SOS-P AGE and western blotting analysis. The expressed recombinant lectin was determinated 1.24∼1.75 $\mu\textrm{g}$ per 1 mg of cytosolic soluble protein by sandwich ELISA method. Moreover the lectin genes were expressed as maximum level at 36 h after galactose induction and lectin A gene was were repressed after 48 h.

Salubrinal-Mediated Upregulation of eIF2α Phosphorylation Increases Doxorubicin Sensitivity in MCF-7/ADR Cells

  • Jeon, Yong-Joon;Kim, Jin Hyun;Shin, Jong-Il;Jeong, Mini;Cho, Jaewook;Lee, Kyungho
    • Molecules and Cells
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    • v.39 no.2
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    • pp.129-135
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    • 2016
  • Eukaryotic translation initiation factor 2 alpha ($eIF2{\alpha}$), which is a component of the eukaryotic translation initiation complex, functions in cell death and survival under various stress conditions. In this study, we investigated the roles of $eIF2{\alpha}$ phosphorylation in cell death using the breast cancer cell lines MCF-7 and MCF-7/ADR. MCF-7/ADR cells are MCF-7-driven cells that have acquired resistance to doxorubicin (ADR). Treatment of doxorubicin reduced the viability and induced apoptosis in both cell lines, although susceptibility to the drug was very different. Treatment with doxorubicin induced phosphorylation of $eIF2{\alpha}$ in MCF-7 cells but not in MCF-7/ADR cells. Basal expression levels of Growth Arrest and DNA Damage 34 (GADD34), a regulator of $eIF2{\alpha}$, were higher in MCF-7/ADR cells compared to MCF-7 cells. Indeed, treatment with salubrinal, an inhibitor of GADD34, resulted in the upregulation of $eIF2{\alpha}$ phosphorylation and enhanced doxorubicin-mediated apoptosis in MCF-7/ADR cells. However, MCF-7 cells did not show such synergic effects. These results suggest that dephosphorylation of $eIF2{\alpha}$ by GADD34 plays an important role in doxorubicin resistance in MCF-7/ADR cells.

Interaction Study of Soybean mosaic virus Proteins with Soybean Proteins using the Yeast-Two Hybrid System

  • Seo, Jang-Kyun;Hwang, Sung-Hyun;Kang, Sung-Hwan;Choi, Hong-Soo;Lee, Su-Heon;Sohn, Seong-Han;Kim, Kook-Hyung
    • The Plant Pathology Journal
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    • v.23 no.4
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    • pp.281-286
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    • 2007
  • Interactions between viral proteins and host proteins are essential for virus replication. Especially, translation of viral genes completely depends on the host machinery. In potyviruses, interactions of genome-linked viral protein (VPg) with host translation factors including eIF4E, eIF(iso)4E, and poly(A)-binding protein (PABP) has previously been characterized. In this study, we investigated interactions between Soybean mosaic virus (SMV) viral proteins and host translation factors by yeast two-hybrid system. SMV VPg interacted with eIF4E, eIF(iso)4E, and PABP in yeast two-hybrid system, while SMV helper component proteinase (HC-pro) interacted with neither of those proteins. The interaction between SMV NIb and PABP was also detected. These results are consistent with those reported previously in other potyviruses. Interestingly, we found reproducible and specific interactions between SMV coat protein (CP) and PABP. Deletion analysis showed that the region of CP comprising amino acids 116 to 206 and the region of PABP comprising amino acids 520 to 580 are involved in CP/PABP interactions. Soybean library screening with SMV NIb by yeast two-hybrid assay also identified several soybean proteins including chlorophyll a/b binding preprotein, photo-system I-N subunit, ribulose 1,5-biphosphate carboxylase, ST-LSI protein, translation initiation factor 1, TIR-NBS type R protein, RNA binding protein, ubiquitin, and LRR protein kinase. Altogether, these results suggest that potyviral replicase may comprise a multi-protein complex with PABP, CP, and other host factors.

Construction and Characterization of Vector Expressing Low Level of Translation Factor eIF5B (단백질합성인자 eIF5B의 저 발현 효모벡터의 제조 및 특성)

  • 최상기;송진희;이준행;이병욱;성치남
    • Korean Journal of Microbiology
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    • v.40 no.1
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    • pp.7-11
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    • 2004
  • eIF5B is a translation initiation factor that delivers Met-$tRNA^{Met}$ to AUG start codon and subsequently joins the small and large ribosomes. In order to study the function of eIF5B encoded by FUN12, we constructed FUN12 which lacked 5' end of its sequence. We found that this construct lacking almost all of its promoter in pRS plasmid partially complemented slow growth phenotype of fun12 deletion strain. Interestingly, this construct expressed N-terminally truncated eIF5B and its expression level was about 5% of that of wild type eIF5B. Low amount of the eIF5B expressed additionally in fun12 deletion strain played a direct role as a limiting factor for its growth. This limiting factor eIF5B in those strains also modulates activities of overall translation in vitro.

Effects of different target sites on antisense RNA-mediated regulation of gene expression

  • Park, Hongmarn;Yoon, Yeongseong;Suk, Shinae;Lee, Ji Young;Lee, Younghoon
    • BMB Reports
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    • v.47 no.11
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    • pp.619-624
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    • 2014
  • Antisense RNA is a type of noncoding RNA (ncRNA) that binds to complementary mRNA sequences and induces gene repression by inhibiting translation or degrading mRNA. Recently, several small ncRNAs (sRNAs) have been identified in Escherichia coli that act as antisense RNA mainly via base pairing with mRNA. The base pairing predominantly leads to gene repression, and in some cases, gene activation. In the current study, we examined how the location of target sites affects sRNA-mediated gene regulation. An efficient antisense RNA expression system was developed, and the effects of antisense RNAs on various target sites in a model mRNA were examined. The target sites of antisense RNAs suppressing gene expression were identified, not only in the translation initiation region (TIR) of mRNA, but also at the junction between the coding region and 3' untranslated region. Surprisingly, an antisense RNA recognizing the upstream region of TIR enhanced gene expression through increasing mRNA stability.

Site-Specific Mutagenesis of the gshI Gene for Increasing the Activity of ${\gamma}$-Glutamylcysteine Synthetase in Escherichia coli K-12

  • Kwak, Joon-Hyeok;Nam, Yong-Suk;Lee, Se-Yong
    • BMB Reports
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    • v.31 no.3
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    • pp.254-257
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    • 1998
  • The gshI gene from the Escherichia coli K-12 strain codes for ${\gamma}-glutamylcysteine$ synthetase which mediates the rate-limiting step of glutathione biosynthesis. The isolated gshI gene from E. coli K-12 has an unusual translation initiation codon, UUG. The 494th amino acid is Ala rather than Gly which was found in a mutant strain E. coli B. In order to improve the translational rate of the gshI gene of E. coli K-12, the initiation codon, UUG, was changed to the usual AUG codon by the site-specific mutagenesis. This change has resulted in a 53% increase of ${\gamma}-glutamylcysteine$ synthetase activity. The enzyme activity was also improved by replacing $Ala^{494}$ with Val (A494V) or Leu (A494L). The replacement of $Ser^{495}$ with Thr (S495T) also resulted in a 62% increase of the enzyme activity. Therefore, the specific activity of ${\gamma}-glutamylcysteine$ synthetase was increased with the increasing chain length of the aliphathic amino acid at the site of the 494th amino acid (Ala<$Val{\leq}Leu$).

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