TY - JOUR
T1 - Cell cycle regulation of human interleukin-8 gene expression by the human immunodeficiency virus type 1 Tat protein
AU - Mahieux, R.
AU - Lambert, P. F.
AU - Agbottah, E.
AU - Halanski, M. A.
AU - Deng, L.
AU - Kashanchi, F.
AU - Brady, J. N.
N1 - Copyright:
Copyright 2007 Elsevier B.V., All rights reserved.
PY - 2001
Y1 - 2001
N2 - The human immunodeficiency virus type 1 (HIV-1) Tat protein has been reported to transactivate several cellular genes, including the potent chemotactic factor interleukin-8 (IL-8). Consistent with these in vitro assays, elevated levels of IL-8 protein are found in the serum of HIV-infected individuals. We now extend these observations by demonstrating that Tat induction of IL-8 is linked to the cell cycle. Cells that constitutively express the Tat(1-86) protein (eTat) and control cells (pCEP) were reversibly blocked at the G1/S border with hydroxyurea or thymidine. The cells were subsequently released, and IL-8 expression was monitored by RNase protection assays and enzyme-linked immunosorbent assay (ELISA). RNase protection assays demonstrated that IL-8 mRNA expression is transiently induced, approximately fourfold, as the Tat-expressing cells enter S phase. Consistent with the RNase protection assay, an increase in IL-8 protein was observed in the cell supernatant using an IL-8 ELISA. Similar experiments were performed following a reversible block at the G2/M border with nocodazole and release into G1. Using the RNase protection assay and ELISA, little or no increase in IL-8 expression was observed during G1. Using gel shift as well as an immobilized DNA binding assay, we demonstrate that the increase in IL-8 gene expression correlates with a specific increase in p65 NF-κB binding activity only in the nucleus of the Tat-expressing cells. Moreover, the CREB-binding protein coactivator is present in the complex in the Tat cell line. Finally, we demonstrate that the presence of the proteasome inhibitor MG-132 inhibits the induction of NF-κB binding, as well as IL-8 expression, supporting the role of NF-κB.
AB - The human immunodeficiency virus type 1 (HIV-1) Tat protein has been reported to transactivate several cellular genes, including the potent chemotactic factor interleukin-8 (IL-8). Consistent with these in vitro assays, elevated levels of IL-8 protein are found in the serum of HIV-infected individuals. We now extend these observations by demonstrating that Tat induction of IL-8 is linked to the cell cycle. Cells that constitutively express the Tat(1-86) protein (eTat) and control cells (pCEP) were reversibly blocked at the G1/S border with hydroxyurea or thymidine. The cells were subsequently released, and IL-8 expression was monitored by RNase protection assays and enzyme-linked immunosorbent assay (ELISA). RNase protection assays demonstrated that IL-8 mRNA expression is transiently induced, approximately fourfold, as the Tat-expressing cells enter S phase. Consistent with the RNase protection assay, an increase in IL-8 protein was observed in the cell supernatant using an IL-8 ELISA. Similar experiments were performed following a reversible block at the G2/M border with nocodazole and release into G1. Using the RNase protection assay and ELISA, little or no increase in IL-8 expression was observed during G1. Using gel shift as well as an immobilized DNA binding assay, we demonstrate that the increase in IL-8 gene expression correlates with a specific increase in p65 NF-κB binding activity only in the nucleus of the Tat-expressing cells. Moreover, the CREB-binding protein coactivator is present in the complex in the Tat cell line. Finally, we demonstrate that the presence of the proteasome inhibitor MG-132 inhibits the induction of NF-κB binding, as well as IL-8 expression, supporting the role of NF-κB.
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U2 - 10.1128/JVI.75.4.1736-1743.2001
DO - 10.1128/JVI.75.4.1736-1743.2001
M3 - Article
C2 - 11160671
AN - SCOPUS:0035136286
VL - 75
SP - 1736
EP - 1743
JO - Journal of Virology
JF - Journal of Virology
SN - 0022-538X
IS - 4
ER -