• Asian-Australasian Journal of Animal Sciences
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• v.33 no.6
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• pp.1012-1022
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• 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.

• Journal of Life Science
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• v.27 no.10
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• pp.1191-1198
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• 2017

Vesicles and organelles are transported along microtubule and delivered to appropriate compartments in cells. The intracellular transport process is mediated by molecular motor proteins, kinesin, and dynein. Kinesin is a plus-end-directed molecular motor protein that moves the various cargoes along microtubule tracks. Kinesin 1 is first isolated from squid axoplasm is a dimer of two heavy chains (KHCs, also called KIF5s), each of which is associated with the light chain (KLC). KIF5s interact with many different binding proteins through their carboxyl (C)-terminal tail region, but their binding proteins have yet to be specified. To identify the interacting proteins for KIF5A, we performed the yeast two-hybrid screening and found a specific interaction with Ras-GTPase-activating protein (GAP) Src homology3 (SH3)-domain-binding protein 2 (G3BP2), which is involved in stress granule formation and mRNA-protein (mRNP) localization. G3BP2 bound to the C-terminal 73 amino acids of KIF5A but did not interact with the KIF5B, nor the KIF5C in the yeast two-hybrid assay. The arginine-glycine-glycine (RGG)/Gly-rich region domain of G3BP2 is a minimal binding domain for interaction with KIF5A. However, G3BP1 did not interact with KIF5A. When co-expressed in HEK-293T cells, G3BP2 co-localized with KIF5A and was co-immunoprecipitated with KIF5A. These results indicate that G3BP2, which was originally identified as a Ras-GAP SH3 domain-binding protein, is a protein that interacts with KIF5A.

• Journal of Life Science
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• v.12 no.1
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• pp.14-18
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• 2002

Subpopulations of nucleotides that bind specifically to a variety of proteins have been isolated from a population of random sequence RNA/DNA molecules. Roughly one in $10^{13}$ random sequence RNA/DNA molecules folds in such a way as to create a specific binding site for small ligands. Since the development of in vitro selection procedure, more than 50 nucleic acid ligands (aptamers) have been isolated. These molecules are very useful for the study of molecular recognition between nucleic acid and protein/organic compound. In addition to these basic studies this method gives us a dream to produce new drugs against several diseases. We focused on several aptamers which specifically binds to trypsin-like serine proteases (thrombin, human neutrophil elastase, activated protein C and NS3 protease of human hepatitis C virus) and want to introduce their structural characteristics and some functions.

• Development and Reproduction
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• v.25 no.4
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• pp.313-319
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• 2021

Hub cells comprise a niche for germline stem cells and cyst stem cells in the Drosophila testis. Hub cells arise from common somatic gonadal precursors in embryos, but the mechanism of their specification is still poorly understood. Here we find that RNA binding proteins Lin28 and Imp mediate transcript stability of Bowl, a known hub specification factor; Bowl transcripts were reduced in the testis of Lin28 and Imp mutants, and also when RNA-mediated interference against Lin28 or Imp was expressed in hub cells. In tissue culture Luciferase assays involving the Bowl 3'UTR, stability of Luc reporter transcripts depended on the Bowl 3'UTR and required Lin28 and Imp. Our findings suggest that proper Bowl function during hub cell specification requires Lin28 and Imp in the testis hub cells.

• BMB Reports
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• v.57 no.7
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• pp.318-323
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• 2024

Regulation of cell fate and lung cell differentiation is associated with Aminoacyl-tRNA synthetases (ARS)-interacting multifunctional protein 2 (AIMP2), which acts as a non-enzymatic component required for the multi-tRNA synthetase complex. In response to DNA damage, a component of AIMP2 separates from the multi-tRNA synthetase complex, binds to p53, and prevents its degradation by MDM2, inducing apoptosis. Additionally, AIMP2 reduces proliferation in TGF-β and Wnt pathways, while enhancing apoptotic signaling induced by tumor necrosis factor-α. Given the crucial role of these pathways in tumorigenesis, AIMP2 is expected to function as a broad-spectrum tumor suppressor. The full-length AIMP2 transcript consists of four exons, with a small section of the pre-mRNA undergoing alternative splicing to produce a variant (AIMP2-DX2) lacking the second exon. AIMP2-DX2 binds to FBP, TRAF2, and p53 similarly to AIMP2, but competes with AIMP2 for binding to these target proteins, thereby impairing its tumor-suppressive activity. AIMP2-DX2 is specifically expressed in a diverse range of cancer cells, including breast cancer, liver cancer, bone cancer, and stomach cancer. There is growing interest in AIMP2-DX2 as a promising biomarker for prognosis and diagnosis, with AIMP2-DX2 inhibition attracting significant interest as a potentially effective therapeutic approach for the treatment of lung, ovarian, prostate, and nasopharyngeal cancers.

• Journal of Life Science
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• v.24 no.10
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• pp.1144-1150
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• 2014

The eIF4E protein is the key regulator of translation initiation. The interaction of eIF4E with eIF4G triggers the translation of mRNA, and several proteins interrupt this association to modulate translation. Human 4E-T is one of the eIF4E-binding partners that represses the translation of bound mRNAs, and it is involved in the transport of eIF4E to processing bodies (P-bodies). Although Clast4, the mouse homolog of human 4E-T, might play critical roles in the regulation of translation, its properties are not well known. In this report, we deciphered the properties of Clast4 by determining its phosphorylation state, binding to eIF4E, and effects of overexpression on cell proliferation. Clast4 was phosphorylated by protein kinase A (PKA) in vivo on several residues of its amino terminus. Nevertheless, the PKA phosphorylation of Clast4 appeared to have no effect on either its eIF4E-binding ability or localization. Clast4 interacted with eIF4E1 and CPEB. The conserved eIF4E-binding sequence in Clast4, $YXXXXL_{\phi}$, was important for binding eIF4E1A but not eIF4E1B. Similar to that of another well-known eIF4E regulator, the eIF4E binding protein (4E-BP), the overexpression of Clast4 decreased cell proliferation. These results suggest that Clast4 acts as a global translation regulator in cells.

• Journal of Life Science
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• v.17 no.5 s.85
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• pp.687-693
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• 2007

Insulin-like growth factor-I(IGF-I) has significant insulin-like anabolic effects which include the stimulation of glucose and amino acid uptake, as well as protein and glycogen synthesis. IGFs exist in serum and other biological fluids as complexes bound to a family of structurally related insulin-like growth factor binding proteins(IGFBPs). Six human IGFBPs can modulate the effects of IGFs on target tissues by several mechanisms, including altering the serum's half-life and the transcapillary transport of IGFs, as well as changing the availability of IGFs to specific cell surface receptors. Human fibroblasts secrete IGFBPs that can modify IGF-I action. Previous to our study using either Northern blotting, and Western blotting have shown that fibroblasts express mRNA IGFBP-3, -4, and -5, and synthesize these proteins. In addition, fibroblast cell lysates revealed that the IGFBP-3 was most abundant. For these reasons, we undertook to gain further insight into the effects of high and low glucose incubation condition on metabolism and IGFBP-3 expression. In results of metabolites and IGFBP-3 expression in GM10 cells cultivated with various glucose concentration, the consumption of glucose and accumulation of triglyceride were increased in condition of high glucose, and total protein level was decreased. in the course of time. After 5 days incubation, levels of free amino acid in medium containing glucose of high concentration glucose were higher than in conditions of low glucose. Although the levels of IGFBP-3 protein and mRNA levels were increased in low glucose, and IGFBP-3 was not affected by any pretense. Taken together, we suggest that the study of growth factors, like IGFs, might be a possible model of diabetes militus in cell, although the results in cell models were not in accord with in vivo.

• Development and Reproduction
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• v.14 no.4
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• pp.287-295
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• 2010

It was recently reported that nesfatin-1/NUCB2, which is secreted from the brain, controls appetite and energy metabolism. The purpose of this research was to confirm whether or not the protein and its binding site should have been expressed in the mouse reproductive organs and to know the possible effects of nesfatin-1 on the reproductive function. Using the ICR female mouse ovary and uterus, the expression of NUCB2 mRNA was confirmed with the conventional PCR and the relative amount of NUCB2 mRNA in the tissues was analyzed with real-time PCR. Immunohistochemical staining was performed using the nesfatin-1 antibody to investigate the nesfatin-1 protein expression and the biotin conjugated nesfatin-1 to confirm the binding site for nesfatin-1 in the ovary. Furthermore, in order to examine if the expression of NUCB2 mRNA in the ovary and uterus is affected by gonadotropin, its mRNA expression was analyzed after PMSG administration into mice. As a result, the expression level of NUCB2 mRNA in the ovary and the uterus was as much as the expression level in hypothalamus. As a result of the immunohistochemical staining, nesfatin-1 proteins were localized at the theca cells, the interstitial cells, and some of the luteal cells. However, the granulosa cells in the follicles did not stain. Interestingly, the oocytes in the some follicles were stained with nesfatin-1. On the other hand, nesfatin-1 protein binding sites were displayed at the theca cells and the interstitial cells near the tunica albuginea. After PMSG administration the expression level of NUCB2 mRNA was increased in the ovary and the uterus. These results demonstrate that for the first time the nesfatin-1 and its binding site were expressed in the ovary and NUCB2 mRNA expression was controlled by gonadotropin, suggesting an important role in the reproductive organs as a local regulator. Therefore, further study is needed to elucidate the functions of nesfatin-1 on the reproductive organs.

• Journal of Life Science
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• v.27 no.7
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• pp.840-848
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• 2017

Proper intracellular transport is essential for normal cell function. Intracellular transport is mediated by microtubule-dependent molecular motor proteins, as well as kinesin and cytoplasmic dynein, which move their cargo along long, microtubule tracks in cells. Kinesins are ATP-dependent plus-end-directed motor proteins in the intracellular transport of organelles, vesicles, RNA complexes, and protein complexes. The mislocalization of these different types of cargo has been linked to cell dysfunction and degeneration. The cargo transport of kinesins can be described by the following steps: binding to the appropriate cargo and/or adaptor proteins, activation of the kinesin's motility and movement along the microtubule, and the release of the cargo at the correct destination. Recently, several studies have revealed the mechanisms for the regulation of kinesin motor activity, including cargo loading and unloading. Intracellular cargo transport is also modulated by adaptor proteins, which link the kinesins to their cargo. The regulatory proteins, which include protein kinases and phosphatases, regulate kinesin motor activity directly through the phosphorylation or dephosphorylation of kinesins and indirectly through the modification of adaptor proteins, such as c-Jun NH-terminal kinase-interacting proteins, or of the microtubule network. These findings lay the groundwork for understanding how kinesins are differentially engaged in intracellular cargo transport. In addition, understanding the regulatory mechanisms of each kinesin is an area of key interest within cell biology and neurophysiology. In this study, we reviewed kinesins' regulation proteins and discuss how their regulation affects cargo recognition and transport.

• Journal of Life Science
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• v.13 no.4
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• pp.541-550
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• 2003

E2 glycoprotein of hepatitis C virus (HCV) comprises a surface of viral particle together with E1 glycoprotein, and is thought to be involved in the attachment of HCV viral particle to receptor (s) on the permissible cells including hepatocytes, B cells, T cells, and monocytes. We constructed a phage library expressing cellular proteins of hepatocytes on the phage surface, which turned out to be 8.8${\times}$$10^5$ cfu of diversity and carried inserts in 95% of library. We screened both cDNA phage library and 12-mer peptide library to identify the cellular proteins binding to E2 protein. Some intracellular proteins including tensin and membrane band 4.1 which are involved in signal transduction of survival and cytoskeleton organization, were selected from cDNA phage library through several rounds of panning and screening. On the contrary, membrane proteins such as CCR7, CKR-L2, and insulin-like growth factor-1 receptor were identified through screening of peptide library. Phages expressing peptides corresponding to those membrane proteins were bound to E2 protein specifically as determined by neutralization of binding assay. Since it is well known that HCV can infect T cells as well as hepatocytes, we examined to see if E2 protein can bind to CCR7, a member of C-protein coupled receptor family expressed on T cells, using CCR7 transfected tells. Human CCR7 cDNA was cloned into pcDNA3.1(-) vector and transfected into human embryonic kidney cell, 293T, and expressed on the surface of the cell as shown by flow cytometer. Binding assay of E2 protein using CCR7 transfected cells indicated that E2 protein bound to CCR7 by dose-dependent mode, giving rise to the possibility that CCR7 might be a putative cellular receptor for HCV.