• Molecules and Cells
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• v.43 no.12
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• pp.975-988
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• 2020

Hypoxia plays important roles in cancer progression by inducing angiogenesis, metastasis, and drug resistance. However, the effects of hypoxia on long noncoding RNA (lncRNA) expression have not been clarified. Herein, we evaluated alterations in lncRNA expression in lung cancer cells under hypoxic conditions using lncRNA microarray analyses. Among 40,173 lncRNAs, 211 and 113 lncRNAs were up- and downregulated, respectively, in both A549 and NCI-H460 cells. Uroplakin 1A (UPK1A) and UPK1A-antisense RNA 1 (AS1), which showed the highest upregulation under hypoxic conditions, were selected to investigate the effects of UPK1A-AS1 on the expression of UPK1A and the mechanisms of hypoxia-inducible expression. Following transfection of cells with small interfering RNA (siRNA) targeting hypoxia-inducible factor 1α (HIF-1α), the hypoxia-induced expression of UPK1A and UPK1A-AS1 was significantly reduced, indicating that HIF-1α played important roles in the hypoxia-induced expression of these targets. After transfection of cells with UPK1A siRNA, UPK1A and UPK1A-AS1 levels were reduced. Moreover, transfection of cells with UPK1A-AS1 siRNA downregulated both UPK1A-AS1 and UPK1A. RNase protection assays demonstrated that UPK1A and UPK1A-AS1 formed a duplex; thus, transfection with UPK1A-AS1 siRNA decreased the RNA stability of UPK1A. Overall, these results indicated that UPK1A and UPK1A-AS1 expression increased under hypoxic conditions in a HIF-1α-dependent manner and that formation of a UPK1A/UPK1A-AS1 duplex affected RNA stability, enabling each molecule to regulate the expression of the other.

• Proceedings of the Korean Institute of Information and Commucation Sciences Conference
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• 2018.10a
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• pp.444-447
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• 2018

Baculovirus expression systems have many known advantages including fast and cost-effective methods to generate large amounts of recombinant proteins in comparison to bacterial expression systems, particularly those requiring complex post-translational modifications. Especially, recombinant baculoviruses can transfer their vectors and express their recombinant proteins in a wide range of mammalian cell types. In this study, baculoviral vectors which were reconstructed from pcDNA3.1 vector, were recombined with cytomegalovirus (CMV) promoter,uroplakin II promoter, polyhedron promoter, vesicular stomatitis virus G (VSVG), enhanced green fluorescent protein (EGFP), and protein transduction domain (PTD). These recombinant vectors were infected with various cells and cell lines. The baculovirus vector thus developed was analyzed by comparing the metastasis and expression of the recombinant genes with conventional vectors. These results suggest that the baculovirus vector has higher efficiency in metastasis and expression than the control vector.

• Proceedings of the Korean Institute of Information and Commucation Sciences Conference
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• 2018.05a
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• pp.588-591
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• 2018

Baculovirus was originally isolated from the alfalfa looper and contains a 134-kbp genome with 154 open reading frames (ORF). The major capsid protein VP39 together with some minor proteins forms the nucleocapsid ($21nm{\times}260nm$) that encloses the DNA with p6.9 protein. They are double-stranded, circular, supercoiled DNA molecules in a rod-shaped capsid. Wild-type baculoviruses exhibit both lytic and occluded life cycles that develop independently throughout the three phases of virus replication. Recombinant baculoviruses can transfer their vectors and express their recombinant proteins in a wide range of mammalian cell types. Especially, inclusion of a dominant selectable marker in these baculoviral vectors can express diverse recombinant genes in many cells. Baculoviral vectors were reconstructed with cytomegalovirus (CMV) promoter,uroplakin II promoter, polyhedron promoter, vesicular stomatitis virus G (VSVG), enhanced green fluorescent protein (EGFP), protein transduction domain (PTD) gene and so on. These reconstructed vectors were infected into various cell and cell lines. We performed transfection and expression of these recombinant vectors comparison with other control vectors. From this study, we knew that transfection and expression of these recombinant vectors have higher efficacy than any control vector. This work was supported by a grant from Mid-Career Researcher Program(NRF-2016R1A2B4016552) through the National Research Foundation of Korea(NRF) funded by the Ministry of Science, ICT & Future Planning(MSIP).