Transcriptional regulation of the human interleukin-6 gene promoter in human T-cell leukemia virus type I-infected T-cell lines: Evidence for the involvement of NF-κB

Naoki Mori, Fumihiko Shirakawa, Hiroko Shimizu, Shuichi Murakami, Susumu Oda, Ken-ichi Yamamoto, Sumiya Eto

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Abstract

Freshly isolated leukemic cells from patients with adult T-cell leukemia (ATL) and human T-cell leukemia virus type I (HTLV-I)-infected T-cell lines constitutively produce high levels of interleukin-6 (IL-6) protein and mRNA. To clarify the mechanisms that lead to the activation of IL-6 gene in HTLV- I-infected cells, we first studied the regulatory regions in the IL-6 gene transcription by transfection of chloramphenicol acetyltransferase (CAT) reporter plasmids containing the IL-6 promoter. When transfected into HTLV- I-infected T-cell lines MT-2 and HUT-102, IL-6 promoter/CAT plasmids were strongly activated without any stimulation. By deletion analysis of 5' upstream region of IL-6 promoter, the DNA region between -73 and -59 bp from the transcription start site of IL-6 gene was important in the expression of IL-6/CAT activities in HTLV-I-infected cells. This region contains nuclear factor (NF)-κB binding site. The site-directed mutation of the κB motif in IL-6/CAT plasmid resulted in the complete abrogation of IL-6 promoter activity in these cells. Furthermore, when IL-6 promoter/CAT plasmid was introduced into an HTLV-I-uninfected T-cell line, Jurkat, IL-6 promoter activity was silent in the basal level, but strongly increased by the cotransfection with an HTLV-I tax expression plasmid. However, tax expression plasmid showed no transactivation activity, when κB site was mutated in IL- 6 promoter/CAT plasmid. We found that the IL-6 κB site specifically formed a complex with NF-κB-containing nuclear extracts from MT-2 and HUT-102 cells. Finally, transfection of HTLV-I tax into Jurkat cells resulted in induction of specific binding of nuclear extracts to the NF-κB sequence. These results strongly suggest that HTLV-I tax gene may transactivate IL-6 gene through κB site in HTLV-I-positive T-cell lines and activation of NF-κB may be crucial in HTLV-I-induced IL-6 gene activation in ATL.

Original languageEnglish
Pages (from-to)2904-2911
Number of pages8
JournalBlood
Volume84
Issue number9
Publication statusPublished - Nov 1 1994
Externally publishedYes

Fingerprint

Human T-lymphotropic virus 1
T-cells
Viruses
Interleukin-6
Genes
T-Lymphocytes
Cell Line
Chloramphenicol O-Acetyltransferase
Plasmids
Taxation
Adult T Cell Leukemia Lymphoma
Chemical activation
Transcriptional Activation
Transfection
pX Genes
Jurkat Cells
Nucleic Acid Regulatory Sequences
Transcription Initiation Site
Transcription
Genetic Promoter Regions

ASJC Scopus subject areas

  • Hematology

Cite this

Transcriptional regulation of the human interleukin-6 gene promoter in human T-cell leukemia virus type I-infected T-cell lines : Evidence for the involvement of NF-κB. / Mori, Naoki; Shirakawa, Fumihiko; Shimizu, Hiroko; Murakami, Shuichi; Oda, Susumu; Yamamoto, Ken-ichi; Eto, Sumiya.

In: Blood, Vol. 84, No. 9, 01.11.1994, p. 2904-2911.

Research output: Contribution to journalArticle

Mori, Naoki ; Shirakawa, Fumihiko ; Shimizu, Hiroko ; Murakami, Shuichi ; Oda, Susumu ; Yamamoto, Ken-ichi ; Eto, Sumiya. / Transcriptional regulation of the human interleukin-6 gene promoter in human T-cell leukemia virus type I-infected T-cell lines : Evidence for the involvement of NF-κB. In: Blood. 1994 ; Vol. 84, No. 9. pp. 2904-2911.
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abstract = "Freshly isolated leukemic cells from patients with adult T-cell leukemia (ATL) and human T-cell leukemia virus type I (HTLV-I)-infected T-cell lines constitutively produce high levels of interleukin-6 (IL-6) protein and mRNA. To clarify the mechanisms that lead to the activation of IL-6 gene in HTLV- I-infected cells, we first studied the regulatory regions in the IL-6 gene transcription by transfection of chloramphenicol acetyltransferase (CAT) reporter plasmids containing the IL-6 promoter. When transfected into HTLV- I-infected T-cell lines MT-2 and HUT-102, IL-6 promoter/CAT plasmids were strongly activated without any stimulation. By deletion analysis of 5' upstream region of IL-6 promoter, the DNA region between -73 and -59 bp from the transcription start site of IL-6 gene was important in the expression of IL-6/CAT activities in HTLV-I-infected cells. This region contains nuclear factor (NF)-κB binding site. The site-directed mutation of the κB motif in IL-6/CAT plasmid resulted in the complete abrogation of IL-6 promoter activity in these cells. Furthermore, when IL-6 promoter/CAT plasmid was introduced into an HTLV-I-uninfected T-cell line, Jurkat, IL-6 promoter activity was silent in the basal level, but strongly increased by the cotransfection with an HTLV-I tax expression plasmid. However, tax expression plasmid showed no transactivation activity, when κB site was mutated in IL- 6 promoter/CAT plasmid. We found that the IL-6 κB site specifically formed a complex with NF-κB-containing nuclear extracts from MT-2 and HUT-102 cells. Finally, transfection of HTLV-I tax into Jurkat cells resulted in induction of specific binding of nuclear extracts to the NF-κB sequence. These results strongly suggest that HTLV-I tax gene may transactivate IL-6 gene through κB site in HTLV-I-positive T-cell lines and activation of NF-κB may be crucial in HTLV-I-induced IL-6 gene activation in ATL.",
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AU - Mori, Naoki

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AU - Shimizu, Hiroko

AU - Murakami, Shuichi

AU - Oda, Susumu

AU - Yamamoto, Ken-ichi

AU - Eto, Sumiya

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N2 - Freshly isolated leukemic cells from patients with adult T-cell leukemia (ATL) and human T-cell leukemia virus type I (HTLV-I)-infected T-cell lines constitutively produce high levels of interleukin-6 (IL-6) protein and mRNA. To clarify the mechanisms that lead to the activation of IL-6 gene in HTLV- I-infected cells, we first studied the regulatory regions in the IL-6 gene transcription by transfection of chloramphenicol acetyltransferase (CAT) reporter plasmids containing the IL-6 promoter. When transfected into HTLV- I-infected T-cell lines MT-2 and HUT-102, IL-6 promoter/CAT plasmids were strongly activated without any stimulation. By deletion analysis of 5' upstream region of IL-6 promoter, the DNA region between -73 and -59 bp from the transcription start site of IL-6 gene was important in the expression of IL-6/CAT activities in HTLV-I-infected cells. This region contains nuclear factor (NF)-κB binding site. The site-directed mutation of the κB motif in IL-6/CAT plasmid resulted in the complete abrogation of IL-6 promoter activity in these cells. Furthermore, when IL-6 promoter/CAT plasmid was introduced into an HTLV-I-uninfected T-cell line, Jurkat, IL-6 promoter activity was silent in the basal level, but strongly increased by the cotransfection with an HTLV-I tax expression plasmid. However, tax expression plasmid showed no transactivation activity, when κB site was mutated in IL- 6 promoter/CAT plasmid. We found that the IL-6 κB site specifically formed a complex with NF-κB-containing nuclear extracts from MT-2 and HUT-102 cells. Finally, transfection of HTLV-I tax into Jurkat cells resulted in induction of specific binding of nuclear extracts to the NF-κB sequence. These results strongly suggest that HTLV-I tax gene may transactivate IL-6 gene through κB site in HTLV-I-positive T-cell lines and activation of NF-κB may be crucial in HTLV-I-induced IL-6 gene activation in ATL.

AB - Freshly isolated leukemic cells from patients with adult T-cell leukemia (ATL) and human T-cell leukemia virus type I (HTLV-I)-infected T-cell lines constitutively produce high levels of interleukin-6 (IL-6) protein and mRNA. To clarify the mechanisms that lead to the activation of IL-6 gene in HTLV- I-infected cells, we first studied the regulatory regions in the IL-6 gene transcription by transfection of chloramphenicol acetyltransferase (CAT) reporter plasmids containing the IL-6 promoter. When transfected into HTLV- I-infected T-cell lines MT-2 and HUT-102, IL-6 promoter/CAT plasmids were strongly activated without any stimulation. By deletion analysis of 5' upstream region of IL-6 promoter, the DNA region between -73 and -59 bp from the transcription start site of IL-6 gene was important in the expression of IL-6/CAT activities in HTLV-I-infected cells. This region contains nuclear factor (NF)-κB binding site. The site-directed mutation of the κB motif in IL-6/CAT plasmid resulted in the complete abrogation of IL-6 promoter activity in these cells. Furthermore, when IL-6 promoter/CAT plasmid was introduced into an HTLV-I-uninfected T-cell line, Jurkat, IL-6 promoter activity was silent in the basal level, but strongly increased by the cotransfection with an HTLV-I tax expression plasmid. However, tax expression plasmid showed no transactivation activity, when κB site was mutated in IL- 6 promoter/CAT plasmid. We found that the IL-6 κB site specifically formed a complex with NF-κB-containing nuclear extracts from MT-2 and HUT-102 cells. Finally, transfection of HTLV-I tax into Jurkat cells resulted in induction of specific binding of nuclear extracts to the NF-κB sequence. These results strongly suggest that HTLV-I tax gene may transactivate IL-6 gene through κB site in HTLV-I-positive T-cell lines and activation of NF-κB may be crucial in HTLV-I-induced IL-6 gene activation in ATL.

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