Influence of intron and exon splicing enhancers on mammalian cell expression of a truncated spike protein of SARS-CoV and its implication for subunit vaccine development.
Identifieur interne : 002520 ( PubMed/Corpus ); précédent : 002519; suivant : 002521Influence of intron and exon splicing enhancers on mammalian cell expression of a truncated spike protein of SARS-CoV and its implication for subunit vaccine development.
Auteurs : Chia-Yin Chang ; Willy W L. Hong ; Pele Chong ; Suh-Chin WuSource :
- Vaccine [ 0264-410X ] ; 2006.
English descriptors
- KwdEn :
- Animals, CHO Cells, Cricetinae, Enhancer Elements, Genetic, Exons, Glycosylation, Introns, Membrane Glycoproteins (genetics), Membrane Glycoproteins (immunology), SARS Virus (immunology), Spike Glycoprotein, Coronavirus, Vaccines, Subunit (immunology), Vaccines, Synthetic (immunology), Viral Envelope Proteins (genetics), Viral Envelope Proteins (immunology), Viral Vaccines (immunology).
- MESH :
- chemical , genetics : Membrane Glycoproteins, Viral Envelope Proteins.
- chemical , immunology : Membrane Glycoproteins, Vaccines, Subunit, Vaccines, Synthetic, Viral Envelope Proteins, Viral Vaccines.
- immunology : SARS Virus.
- Animals, CHO Cells, Cricetinae, Enhancer Elements, Genetic, Exons, Glycosylation, Introns, Spike Glycoprotein, Coronavirus.
Abstract
The spike (S) protein of severe acute respiratory syndrome coronavirus (SARS-CoV) is important for vaccine development. A truncated S protein of the TW1 strain, STR2 (88 kDa), carrying three S fragments (S74-253, S294-739, and S1129-1255) was investigated to study the influences of intron and exon splicing enhancers to improve STR2 protein expression in mammalian cells. Our results showed that STR2 protein expression with the use of an 138 base-pair intron addition increased by 1.9-, 2.5-, and 4.1-fold in Vero E6, QBI-293A cells, and CHO/dhFr- cells (dihydrofolate reductase [dhfr] gene deficient CHO cells), respectively. Using the exon splicing enhancers, including a bidirectional splicing enhancer (BSE) or an exon splicing enhancer derived from the EDA alternative exon of the fibronectin gene (EDA ESE), were also found to increase STR2 protein expression in CHO/dhFr- cells by 1.7- and 2.6-fold. Nevertheless, combination of the intron and the exon splicing enhancers resulted in suppressing the intron-enhancing e STR2 protein expression in in CHO/dhFr- cells. Our studies also demonstrated the STR2 protein was mainly as the Endo H-sensitive glycoprotein (115 kDa) expressed in Vero E6, QBI-293A, and CHO/dhFr- cells. However, only a minor form of the Endo H-resistant glycoproteins ( approximately 130 kDa) was detected in CHO/dhFr- cells. Taken together, our results indicated that intron had a better enhancing effect on STR2 protein expression than exon splicing enhancers, and the expression of approximately 130 kDa STR2 glycoprotein was enhanced by the intron addition into the expression vector construct. Results of the present study can provide an optimal strategy to enhance SARS-CoV S protein expression in mammalian cells and may contribute to the development of SARS-CoV subunit vaccine.
DOI: 10.1016/j.vaccine.2005.09.011
PubMed: 16194584
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pubmed:16194584Le document en format XML
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Hong</name></author><author><name sortKey="Chong, Pele" sort="Chong, Pele" uniqKey="Chong P" first="Pele" last="Chong">Pele Chong</name></author><author><name sortKey="Wu, Suh Chin" sort="Wu, Suh Chin" uniqKey="Wu S" first="Suh-Chin" last="Wu">Suh-Chin Wu</name></author></titleStmt><publicationStmt><idno type="wicri:source">PubMed</idno><date when="2006">2006</date><idno type="RBID">pubmed:16194584</idno><idno type="pmid">16194584</idno><idno type="doi">10.1016/j.vaccine.2005.09.011</idno><idno type="wicri:Area/PubMed/Corpus">002520</idno><idno type="wicri:explorRef" wicri:stream="PubMed" wicri:step="Corpus" wicri:corpus="PubMed">002520</idno></publicationStmt><sourceDesc><biblStruct><analytic><title xml:lang="en">Influence of intron and exon splicing enhancers on mammalian cell expression of a truncated spike protein of SARS-CoV and its implication for subunit vaccine development.</title><author><name sortKey="Chang, Chia Yin" sort="Chang, Chia Yin" uniqKey="Chang C" first="Chia-Yin" last="Chang">Chia-Yin Chang</name><affiliation><nlm:affiliation>Institute of Biotechnology, Department of Life Science, National Tsing Hua University, Hsinchu 30013, Taiwan.</nlm:affiliation></affiliation></author><author><name sortKey="Hong, Willy W L" sort="Hong, Willy W L" uniqKey="Hong W" first="Willy W L" last="Hong">Willy W L. Hong</name></author><author><name sortKey="Chong, Pele" sort="Chong, Pele" uniqKey="Chong P" first="Pele" last="Chong">Pele Chong</name></author><author><name sortKey="Wu, Suh Chin" sort="Wu, Suh Chin" uniqKey="Wu S" first="Suh-Chin" last="Wu">Suh-Chin Wu</name></author></analytic><series><title level="j">Vaccine</title><idno type="ISSN">0264-410X</idno><imprint><date when="2006" type="published">2006</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass><keywords scheme="KwdEn" xml:lang="en"><term>Animals</term><term>CHO Cells</term><term>Cricetinae</term><term>Enhancer Elements, Genetic</term><term>Exons</term><term>Glycosylation</term><term>Introns</term><term>Membrane Glycoproteins (genetics)</term><term>Membrane Glycoproteins (immunology)</term><term>SARS Virus (immunology)</term><term>Spike Glycoprotein, Coronavirus</term><term>Vaccines, Subunit (immunology)</term><term>Vaccines, Synthetic (immunology)</term><term>Viral Envelope Proteins (genetics)</term><term>Viral Envelope Proteins (immunology)</term><term>Viral Vaccines (immunology)</term></keywords><keywords scheme="MESH" type="chemical" qualifier="genetics" xml:lang="en"><term>Membrane Glycoproteins</term><term>Viral Envelope Proteins</term></keywords><keywords scheme="MESH" type="chemical" qualifier="immunology" xml:lang="en"><term>Membrane Glycoproteins</term><term>Vaccines, Subunit</term><term>Vaccines, Synthetic</term><term>Viral Envelope Proteins</term><term>Viral Vaccines</term></keywords><keywords scheme="MESH" qualifier="immunology" xml:lang="en"><term>SARS Virus</term></keywords><keywords scheme="MESH" xml:lang="en"><term>Animals</term><term>CHO Cells</term><term>Cricetinae</term><term>Enhancer Elements, Genetic</term><term>Exons</term><term>Glycosylation</term><term>Introns</term><term>Spike Glycoprotein, Coronavirus</term></keywords></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en">The spike (S) protein of severe acute respiratory syndrome coronavirus (SARS-CoV) is important for vaccine development. A truncated S protein of the TW1 strain, STR2 (88 kDa), carrying three S fragments (S74-253, S294-739, and S1129-1255) was investigated to study the influences of intron and exon splicing enhancers to improve STR2 protein expression in mammalian cells. Our results showed that STR2 protein expression with the use of an 138 base-pair intron addition increased by 1.9-, 2.5-, and 4.1-fold in Vero E6, QBI-293A cells, and CHO/dhFr- cells (dihydrofolate reductase [dhfr] gene deficient CHO cells), respectively. Using the exon splicing enhancers, including a bidirectional splicing enhancer (BSE) or an exon splicing enhancer derived from the EDA alternative exon of the fibronectin gene (EDA ESE), were also found to increase STR2 protein expression in CHO/dhFr- cells by 1.7- and 2.6-fold. Nevertheless, combination of the intron and the exon splicing enhancers resulted in suppressing the intron-enhancing e STR2 protein expression in in CHO/dhFr- cells. Our studies also demonstrated the STR2 protein was mainly as the Endo H-sensitive glycoprotein (115 kDa) expressed in Vero E6, QBI-293A, and CHO/dhFr- cells. However, only a minor form of the Endo H-resistant glycoproteins ( approximately 130 kDa) was detected in CHO/dhFr- cells. Taken together, our results indicated that intron had a better enhancing effect on STR2 protein expression than exon splicing enhancers, and the expression of approximately 130 kDa STR2 glycoprotein was enhanced by the intron addition into the expression vector construct. Results of the present study can provide an optimal strategy to enhance SARS-CoV S protein expression in mammalian cells and may contribute to the development of SARS-CoV subunit vaccine.</div></front></TEI><pubmed><MedlineCitation Status="MEDLINE" Owner="NLM"><PMID Version="1">16194584</PMID><DateCompleted><Year>2006</Year><Month>03</Month><Day>23</Day></DateCompleted><DateRevised><Year>2020</Year><Month>04</Month><Day>15</Day></DateRevised><Article PubModel="Print-Electronic"><Journal><ISSN IssnType="Print">0264-410X</ISSN><JournalIssue CitedMedium="Print"><Volume>24</Volume><Issue>8</Issue><PubDate><Year>2006</Year><Month>Feb</Month><Day>20</Day></PubDate></JournalIssue><Title>Vaccine</Title><ISOAbbreviation>Vaccine</ISOAbbreviation></Journal><ArticleTitle>Influence of intron and exon splicing enhancers on mammalian cell expression of a truncated spike protein of SARS-CoV and its implication for subunit vaccine development.</ArticleTitle><Pagination><MedlinePgn>1132-41</MedlinePgn></Pagination><Abstract><AbstractText>The spike (S) protein of severe acute respiratory syndrome coronavirus (SARS-CoV) is important for vaccine development. A truncated S protein of the TW1 strain, STR2 (88 kDa), carrying three S fragments (S74-253, S294-739, and S1129-1255) was investigated to study the influences of intron and exon splicing enhancers to improve STR2 protein expression in mammalian cells. Our results showed that STR2 protein expression with the use of an 138 base-pair intron addition increased by 1.9-, 2.5-, and 4.1-fold in Vero E6, QBI-293A cells, and CHO/dhFr- cells (dihydrofolate reductase [dhfr] gene deficient CHO cells), respectively. Using the exon splicing enhancers, including a bidirectional splicing enhancer (BSE) or an exon splicing enhancer derived from the EDA alternative exon of the fibronectin gene (EDA ESE), were also found to increase STR2 protein expression in CHO/dhFr- cells by 1.7- and 2.6-fold. Nevertheless, combination of the intron and the exon splicing enhancers resulted in suppressing the intron-enhancing e STR2 protein expression in in CHO/dhFr- cells. Our studies also demonstrated the STR2 protein was mainly as the Endo H-sensitive glycoprotein (115 kDa) expressed in Vero E6, QBI-293A, and CHO/dhFr- cells. However, only a minor form of the Endo H-resistant glycoproteins ( approximately 130 kDa) was detected in CHO/dhFr- cells. Taken together, our results indicated that intron had a better enhancing effect on STR2 protein expression than exon splicing enhancers, and the expression of approximately 130 kDa STR2 glycoprotein was enhanced by the intron addition into the expression vector construct. Results of the present study can provide an optimal strategy to enhance SARS-CoV S protein expression in mammalian cells and may contribute to the development of SARS-CoV subunit vaccine.</AbstractText></Abstract><AuthorList CompleteYN="Y"><Author ValidYN="Y"><LastName>Chang</LastName><ForeName>Chia-Yin</ForeName><Initials>CY</Initials><AffiliationInfo><Affiliation>Institute of Biotechnology, Department of Life Science, National Tsing Hua University, Hsinchu 30013, Taiwan.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Hong</LastName><ForeName>Willy W L</ForeName><Initials>WW</Initials></Author><Author ValidYN="Y"><LastName>Chong</LastName><ForeName>Pele</ForeName><Initials>P</Initials></Author><Author ValidYN="Y"><LastName>Wu</LastName><ForeName>Suh-Chin</ForeName><Initials>SC</Initials></Author></AuthorList><Language>eng</Language><PublicationTypeList><PublicationType UI="D016428">Journal Article</PublicationType><PublicationType UI="D013485">Research Support, 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Synthetic</NameOfSubstance></Chemical><Chemical><RegistryNumber>0</RegistryNumber><NameOfSubstance UI="D014759">Viral Envelope Proteins</NameOfSubstance></Chemical><Chemical><RegistryNumber>0</RegistryNumber><NameOfSubstance UI="D014765">Viral Vaccines</NameOfSubstance></Chemical><Chemical><RegistryNumber>0</RegistryNumber><NameOfSubstance UI="C578557">spike glycoprotein, SARS-CoV</NameOfSubstance></Chemical></ChemicalList><CitationSubset>IM</CitationSubset><MeshHeadingList><MeshHeading><DescriptorName UI="D000818" MajorTopicYN="N">Animals</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D016466" MajorTopicYN="N">CHO Cells</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D006224" MajorTopicYN="N">Cricetinae</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D004742" MajorTopicYN="N">Enhancer Elements, Genetic</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D005091" MajorTopicYN="Y">Exons</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D006031" MajorTopicYN="N">Glycosylation</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D007438" MajorTopicYN="Y">Introns</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D008562" MajorTopicYN="N">Membrane Glycoproteins</DescriptorName><QualifierName UI="Q000235" MajorTopicYN="Y">genetics</QualifierName><QualifierName UI="Q000276" MajorTopicYN="Y">immunology</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D045473" MajorTopicYN="N">SARS Virus</DescriptorName><QualifierName UI="Q000276" MajorTopicYN="Y">immunology</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D064370" MajorTopicYN="N">Spike Glycoprotein, Coronavirus</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D022223" MajorTopicYN="N">Vaccines, Subunit</DescriptorName><QualifierName UI="Q000276" 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