Background
HIV-1 subtype C (HIV-1C) has a unique genetic characteristic of incorporating additional NF-kB binding motifs within its long terminal repeat (LTR) region, which enhances its transcriptional strength and replication fitness. Unlike other HIV-1 subtypes, HIV-1C demonstrates a subtype-specific interaction between its central NF-kB and Sp1III motifs. Our recent research identified nine emerging HIV-1C LTR variants from India, exhibiting co-duplication of NF-kB motifs along with upstream LEF and RBEIII sites, known to suppress transcription. This co-evolution of transcription-enhancing (NF-kB) and suppressive (LEF, RBEIII) motifs is novel and significant for understanding viral latency and transcription regulation.
We hypothesize that the presence of both classical H-kB and unique C-kB motifs in HIV-1C LTR provides an advantage by optimizing receptivity to classical and alternative NF-kB signals. This adaptability is specific to HIV-1C, distinguishing it from other subtypes. Our study explores whether NF-kB motif polymorphisms in HIV-1C LTR allow efficient integration of these pathways and enhance latency reversal compared to other subtypes.
Methods
Jurkat cells were treated with AZD5582 to selectively activate the alternative NF-kB pathway. Activation was confirmed by p100 processing into p52, while IκBα levels confirmed no classical pathway activation. Jurkat cells infected with HIV-1B or HIV-1C LTR reporter viruses were treated with AZD5582 (Figure1-A). Additionally, primary CD4+ T cells infected with a dual-reporter HIV-1 virus were treated with AZD5582, TNF-α, or PMA.
Results
Flow cytometry showed a higher response in HIV-1C LTR to alternative NF-kB signaling than HIV-1B (Figure-1B). Immunoblotting confirmed p52 processing in CD4+ T cells treated with AZD5582, with increased reactivated cells observed via flow cytometry, indicating enhanced transcription and latency reversal potential in HIV-1C (Figure-1C).
Conclusions
In conclusion, our results suggest that HIV-1C LTR integrates both classical and alternative NF-kB signals more effectively than HIV-1B, contributing to its transcriptional superiority and replication fitness. Future research will assess clinical samples to further compare HIV-1B and HIV-1C LTR responses, potentially informing novel strategies for HIV-1 latency reversal and cure.