NFAT5 regulates HIV-1 in primary monocytes via a highly conserved long terminal repeat site.
To replicate, HIV-1 capitalizes on endogenous cellular activation pathways resulting in recruitment of key host transcription factors to its viral enhancer. RNA interference has been a powerful tool for blocking key checkpoints in HIV-1 entry into cells. Here we apply RNA interference to HIV-1 transcription in primary macrophages, a major reservoir of the virus, and specifically target the transcription factor NFAT5 (nuclear factor of activated T cells 5), which is the most evolutionarily divergent NFAT protein. By molecularly cloning and sequencing isolates from multiple viral subtypes, and performing DNase I footprinting, electrophoretic mobility shift, and promoter mutagenesis transfection assays, we demonstrate that NFAT5 functionally interacts with a specific enhancer binding site conserved in HIV-1, HIV-2, and multiple simian immunodeficiency viruses. Using small interfering RNA to ablate expression of endogenous NFAT5 protein, we show that the replication of three major HIV-1 viral subtypes (B, C, and E) is dependent upon NFAT5 in human primary differentiated macrophages. Our results define a novel host factor-viral enhancer interaction that reveals a new regulatory role for NFAT5 and defines a functional DNA motif conserved across HIV-1 subtypes and representative simian immunodeficiency viruses. Inhibition of the NFAT5-LTR interaction may thus present a novel therapeutic target to suppress HIV-1 replication and progression of AIDS.
Ranjbar
Shahin
S
CBR Institute for Biomedical Research, Harvard Medical School, Boston, Massachusetts, United States of America.
Tsytsykova
Alla V
AV
Lee
Sang-Kyung
SK
Rajsbaum
Ricardo
R
Falvo
James V
JV
Lieberman
Judy
J
Shankar
Premlata
P
Goldfeld
Anne E
AE
D000163:Acquired Immunodeficiency Syndrome / Q000503:physiopathology; D002460:Cell Line; D002478:Cells, Cultured; D004279:DNA, Viral; D018450:Disease Progression; D005786:Gene Expression Regulation; D015497:HIV-1 / Q000235:genetics / Q000472:pathogenicity / Q000502:physiology*; D006367:HeLa Cells; D006801:Humans; D008264:Macrophages / Q000502:physiology / Q000821:virology*; D011485:Protein Binding; D034622:RNA Interference; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D020079:Terminal Repeat Sequences / Q000235:genetics / Q000502:physiology*; D014157:Transcription Factors / Q000235:genetics / Q000502:physiology*; D014779:Virus Replication / Q000235:genetics / Q000502:physiology*
D004279:DNA, Viral; C118125:NFAT5 protein, human; D012333:RNA, Messenger; D014157:Transcription Factors