• BMB Reports
• /
• 제50권4호
• /
• pp.194-200
• /
• 2017

Eukaryotic gene expression is precisely regulated at all points between transcription and translation. In this review, we focus on translational control mediated by the 3'-untranslated regions (UTRs) of mRNAs. mRNA 3'-UTRs contain cis-acting elements that function in the regulation of protein translation or mRNA decay. Each RNA binding protein that binds to these cis-acting elements regulates mRNA translation via various mechanisms targeting the mRNA cap structure, the eukaryotic initiation factor 4E (eIF4E)-eIF4G complex, ribosomes, and the poly (A) tail. We also discuss translation-mediated regulation of mRNA fate.

• Journal of Microbiology and Biotechnology
• /
• 제34권6호
• /
• pp.1348-1355
• /
• 2024

The eukaryotic translation initiation factor eIF5B is a bacterial IF2 ortholog that plays an important role in ribosome joining and stabilization of the initiator tRNA on the AUG start codon during the initiation of translation. We identified the fluorophenyl oxazole derivative 2,2-dibromo-1-(2-(4-fluorophenyl)benzo[d]oxazol-5-yl)ethanone quinolinol as an inhibitor of fungal protein synthesis using an in vitro translation assay in a fungal system. Mutants resistant to this compound were isolated in Saccharomyces cerevisiae and were demonstrated to contain amino acid substitutions in eIF5B that conferred the resistance. These results suggest that eIF5B is a target of potential antifungal compound and that mutation of eIF5B can confer resistance. Subsequent identification of 16 other mutants revealed that primary mutations clustered mainly on domain 2 of eIF5B and secondarily mainly on domain 4. Domain 2 has been implicated in the interaction with the small ribosomal subunit during initiation of translation. The tested translation inhibitor could act by weakening the functional contact between eIF5B and the ribosome complex. This data provides the basis for the development of a new family of antifungals.

• 식물조직배양학회지
• /
• 제28권5호
• /
• pp.283-287
• /
• 2001

감자 (Solanum tuberosum L. cv. Irish Cobbler)의 괴경형성과정 (tuberization) 동안에 발현하는 유전자들의 발현양상을 조사하고자 differential display법을 실시하였다. Differential display를 이용하여 분리된 eIF5A DNA단편을 probe로 사용하여 감자의 cDNA library screening을 통하여 eIF5A full-length cDNA를 감자에서 처음으로 분리하였다. 감자의 eIF5A, clone은 토마토의 eIF5A cDNA 염기서열과 94.8%. 아미노산 서열에서는 97.5%로 매우 높은 유사성을 나타내었다. 감자의 eIF5A 유전자는 길이가 716 bp로 하나의 단백질 code영역 (ORF)을 포함하고 있었다. 이 영역은 분자량 17.4 kD, pI 5.5로 추정되는 160개의 아미노산으로 구성된 eIF5A단백질을 code하고 있었다. eIF5A 단백질들에서 12개의 아미노산 서열 (STSKTGKHGHAK)은 효모에서 사람에 이르기까지 완벽하게 보존되어 있는 것으로 알려져 있는데, 감자에서도 또한 잘 보존되어 있었다. 이 영역은 eIF5A 단백질의 활성을 나타내는 데 있어서 필수적인 hypusine을 생성하는 전사 후 수식 부위가 들어 있는 아주 중요한 곳이다. 감자에서 eIF5A 유전자의 발현양상을 조사한 결과 감자의 전조직에서 발현을 보였는데, 성숙잎이나 괴경보다는 세포분열 및 물질축적이 활발히 일어나고 있는 꽃기관들 (stamen, ovary, petal. sepal), 과실 (fruit)과 stolen 등의 조직들에서 비교적 활발히 발현되고 있었다.

• 생명과학회지
• /
• 제23권7호
• /
• pp.941-946
• /
• 2013

신경세포의 가지돌기 내 단백질합성은 필요한 단백질을 실시간으로 제공할 수 있는 이점을 제공한다. 본 연구에서는 단백질합성인자 eIF4E와 그 억제 단백질인 eIF4EBP1의 발생단계별 표현을 배양한 해마신경세포를 면역 염색하여 조사하였다. eIF4E는 가지돌기에 점박이 모양으로 표현되었으며, 핵에는 표현되지 않았다. 그러나 eIF4EBP1는 가지돌기 뿐 아니라 발생초기(DIV 0.5)부터 핵에서 표현되었으며 성숙한 세포에서 핵에 더욱 뚜렷이 표현되었다. eIF4E 혹은 eIF4EBP1의 PSD95과의 colocalization은 $39.1{\pm}9.6%$ 및 $70.5{\pm}5.2%$ (DIV 7), $57.7{\pm}8.2%$ 및 $36.0{\pm}3.1%$ (DIV 10), $29.9{\pm}2.9%$ 및 $40.2{\pm}11.7%$ (DIV 20)이었다. eIF4E와 eIF4EBP1의 colocalizatin은 $18.5{\pm}2.6%$ (DIV 7), $11.1{\pm}3.9%$ (DIV 10), $38.6{\pm}5.6%$ (DIV 20)이었다. 이 결과는 eIF4E 및 eIF4EBP1의 많은 부분이 연접후에 위치하며, 발생초기에는 eIF4E가 활동적인 형태로 존재하지만, 성숙 신경세포에서는 eIF4EBP1과 결합하여 비활성적인 형태로 존재함을 의미한다.

• 한국미생물·생명공학회지
• /
• 제36권4호
• /
• pp.353-359
• /
• 2008

Hepatitis C virus (HCV) is known as the causative agent of blood transmitted hepatitis. Two viral proteins, E2 and NS5A, are known to exert interferon resistance of HCV via PKR pathway. Here, we report a third protein, the RNA-dependent RNA polymerase (NS5B) of HCV, induced interferon resistance inhibiting p56 pathway. p56 was shown to interact with p48 subunit of eukaryotic initiation factor 3 (eIF3). This interaction inhibited formation of ternary complex in translation initiation. Using dual reporter assay system, we observed that the translation decreased when interferon alpha was added to the culture. But, in the presence of HCV NS5B, the translation partly recovered. NS5B and p48 subunit of eIF3 were shown to interact. This interaction seems to inhibit the interaction between p48 and p56. This is the first report that a virus exerts interferon resistance via p56 pathway.

• 통합자연과학논문집
• /
• 제12권4호
• /
• pp.127-132
• /
• 2019

Programmed cell death 4 (PDCD4) is a novel tumor suppressor that function in the nucleus and the cytoplasm and appears to be involved in the regulation of transcription and translation. Stress granules (SGs) are cytoplasmic foci at which untranslated mRNAs accumulate when cells exposed to environmental stresses. Since PDCD4 has implicated in translation repression through direct interaction with eukaryotic translation initiation factor 4A (eIF4A), we here investigated if PDCD4 has a functional role in the process of SG assembly under oxidative stresses. Using immunofluorescence microscopy, we found that PDCD4 is localized to SGs under oxidative stresses. Next, we tested if knockdown of PDCD4 has an effect on the assembly of SG using PDCD4-specific siRNA. Interestingly, SG assembly was accelerated and this effect was caused by sensitization of phosphorylation of eIF2α and dephosphorylation of eIF4E binding protein (4E-BP). These results suggest that PDCD4 has an effect on SG dynamics and possibly involved in cap-dependent translation repression under stress conditions.

• Asian-Australasian Journal of Animal Sciences
• /
• 제33권6호
• /
• pp.1012-1022
• /
• 2020

Objective: Caseins and fatty acids of milk are synthesized and secreted by the epithelial cells of the mammary gland. All-trans retinoic acid (ATRA), an active metabolite of vitamin A, has been shown to promote mammary development. This study was conducted to determine the effect of ATRA on casein synthesis and fatty acid composition in MAC-T cells. Methods: MAC-T cells were allowed to differentiate for 4 d, treated with ATRA (0, 1.0, 1.5, and 2.0 μM), and incubated for 3 d. We analyzed the fatty acid composition, the mRNA expression of casein and fatty acid synthesis-related genes, and the phosphorylation of casein synthesis-related proteins of MAC-T cells by gas chromatography, quantitative polymerase chain reaction, and western blotting, respectively. Results: In MAC-T cells, ATRA increased the mRNA levels of α_S1-casein and β-casein, janus kinase 2 (JAK2) and E74-like factor 5 of the signal transducer and activator of transcription 5 β (STAT5-β) pathway, ribosomal protein S6 kinase beta-1 (S6K1) and eukaryotic translation initiation factor 4E binding protein 1 of the mammalian target of rapamycin (mTOR) pathway, inhibited the mRNA expression of phosphoinositide 3-kinase and eukaryotic initiation factor 4E of the mTOR pathway, and promoted the phosphorylation of STAT5-β and S6K1 proteins. Additionally, ATRA increased the de novo synthesis of fatty acids, reduced the content of long-chain fatty acids, the ratio of monounsaturated fatty acids to saturated fatty acids (SFA), the ratio of polyunsaturated fatty acids (PUFA) to SFA, and the ratio of ω-6 to ω-3 PUFA. The mRNA levels of acetyl-CoA carboxylase 1, fatty acid synthase, lipoprotein lipase, stearoyl-CoA desaturase, peroxisome proliferator-activated receptor gamma, and sterol regulatory element-binding protein 1 (SREBP1) were enhanced by ATRA. Conclusion: ATRA promotes the synthesis of casein by regulating JAK2/STAT5 pathway and downstream mTOR signaling pathway, and it improves the fatty acid composition of MAC-T cells by regulating SREBP1-related genes.

• Toxicological Research
• /
• 제8권2호
• /
• pp.343-357
• /
• 1992

Tumor necrosis factor-${\alpha}$(TNF), a polypeptide hormone secreted primarily by activated macrophages, was originally identified on the basis of its ability to cause hemorrhagic necrosis and tumor regression in vivo. Subsequently, TNF has been shown to be an important component of the host responses to infection and cancer and may mediate the wasting syndrome known as cachexia. These systemic actions of TNF are reflected in its diverse effects on target cells in vitro. TNF initiates its diverse cellular actions by binding to specific cell surface receptors. Although TNF receptors have been identified on most of animal cells, regulation of these receptors and the mechanisms which transduce TNF receptor binding into cellular responses are not well understood. Therefore, in the present study, the mechanisms how TNF receptors are being regulated and how TNF receptor binding is being transduced into cellular responses were investigated in rat liver plasma membranes (PM) and ME-180 human cervical carcinoma cell lines. $^{125}I$-TNF bound to high ($K_d=1.51{\pm}0.35nM$)affinity receptors in rat liver PM. Solubilization of PM with 1% Triton X-100 increased both high affinity (from $0.33{\pm}0.04\;to\;1.67{\pm}0.05$ pmoles/mg protein) and low affinity (from $1.92{\pm}0.16\;to\;7.57{\pm}0.50$ pmoles/mg protein) TNF binding without affecting the affinities for TNF, suggesting the presence of a large latent pool of TNF receptors. Affinity labeling of receptors whether from PM or solubilized PM resulted in cross-linking of $^{125}I$-TNF into $M_r$ 130 kDa, 90 kDa and 66kDa complexes. Thus, the properties of the latent TNF receptors were similar to those initially accessible to TNF. To determine if exposure of latent receptors is regulated by TNF, $^{125}I$-TNF binding to control and TNF-pretreated membranes were assayed. Specific binding was increased by pretreatment with TNF (P<0.05), demonstrating that hepatic PM contains latent TNF receptors whose exposure is promoted by TNF. Homologous up-regulation of TNF receptors may, in part, be responsible for sustained hepatic responsiveness during chronic exposure to TNF. As a next step, the post-receptor events induced by TNF were examined. Although the signal transduction pathways for TNF have not been delineated clearly, the actions of many other hormones are mediated by the reversible phosphorylation of specific enzymes or target proteins. The present study demonstrated that TNF induces phosphorylation of 28 kDa protein (p28). Two dimensional soidum dodecyl sulfate-polyacrylamide gel electrophoresis(SDS-PAGE) resolved the 28kDa phosphoprotein into two isoforms having pIs of 6.2 and 6.1. The pIs and relative molecular weight of p28 were consistent with those of a previously characterized mRNA cap binding protein. mRNA cap binding proteins are a class of translation initiation factors that recognize the 7-methylguanosine cap structure found on the 5' end of eukaryotic mRNAs. In vitro, these proteins are defined by their specific elution from affinity columns composed of 7-methylguanosine 5'-triphosphate($m^7$GTP)-Sepharose. Affinity purification of mRNA cap binding proteins from control and TNF treated ME-180 cells proved that TNF rapidly stimulates phosphorylation of an mRNA cap binding protein. Phosphorylation occurred in several cell types that are important in vitro models of TNF action. The mRNA cap binding protein phosphorylated in response to TNF treatment was purifice, sequenced, and identified as the proto-oncogene product eukaryotic initiation factor-4E(eIF-4E). These data show that phosphorylation of a key component of the cellular translational machinery is a common early event in the diverse cellular actions of TNF.

• Molecules and Cells
• /
• 제42권6호
• /
• pp.495-500
• /
• 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).

• The Plant Pathology Journal
• /
• 제37권1호
• /
• pp.47-56
• /
• 2021

Plants protect against viruses through passive and active resistance mechanisms, and in most cases characterized thus far, natural recessive resistance to potyviruses has been mapped to mutations in the eukaryotic initiation factor eIF4E or eIF(iso)4E genes. Five eIF4E copies and three eIF(iso)4E copies were detected in Brassica rapa. The eIF4E and eIF(iso)4E genes could interact with turnip mosaic virus (TuMV) viral protein linked to the genome (VPg) to initiate virus translation. From the yeast two-hybrid system (Y2H) and bimolecular fluorescence complementation (BiFC) assays, the TuMV-CHN2/CHN3 VPgs could not interact with BraA.eIF4E.a/c or BraA.eIF(iso)4E.c, but they could interact with BraA.eIF(iso)4E.a in B. rapa. Further analysis indicated that the amino acid substitution L186F (nt T556C) in TuMV-UK1 VPg was important for the interaction networks between the TuMV VPg and eIF(iso)4E proteins. An interaction model of the BraA. eIF(iso)4E protein with TuMV VPg was constructed to infer the effect of the significant amino acids on the interaction of TuMV VPgs-eIF(iso)4Es, particularly whether the L186F in TuMV-UK1 VPg could change the structure of the TuMV-UK1 VPg protein, which may terminate the interaction of the BraA.eIF(iso)4E and TuMV VPg protein. This study provides new insights into the interactions between plant viruses and translation initiation factors to reveal the working of key amino acids.