RNA-Seq, NChip-Seq and RIP-Seq analysis of FUBP3 in T cells
Description
The Far Upstream Element Binding Protein 3 (FUBP3) enhances HIV-1 transcription and regulates immune responses pathways in T cells: FUBP3 (Far Upstream Element-Binding Protein 3) has emerged as a hit factor in HIV-1 transcription, yet its molecular function and mechanisms remained unexplored. In this study, we investigate the role of FUBP3 in HIV-1 transcriptional activation, focusing on its interactions with the viral transactivator protein Tat and TAR RNA. Using a combination of transcriptomic analyses, chromatin immunoprecipitation, and biochemical assays, we demonstrate that FUBP3 enhances HIV-1 transcription in a Tat-dependent manner. FUBP3 directly binds and stabilizes TAR RNA, exhibiting a strong preference for its secondary structure, which is essential for efficient Tat function. Knockdown of FUBP3 results in significant downregulation of HIV-1 transcription and alters the expression of numerous host genes involved in T cell activation and inflammation, underscoring its role in regulating immune responses. Our findings position FUBP3 as a key player in the HIV-1 lifecycle, enhancing our understanding of the interplay between viral and host factors. This study opens avenues for future research aimed at targeting FUBP3 for therapeutic intervention in HIV-1 infection, particularly in the context of latency and reactivation. By elucidating the multifaceted roles of FUBP3, we contribute valuable insights into its potential as a therapeutic target and its implications in oncogenic and inflammatory pathways. RNA-Sequencing analysis of total RNA extracted from J-Lat 10.6, Jurkat and primary CD4+T cells upon FUBP3 depletion by shRNAmirs (shFUBP3 and shCD8B). RIP-Seq analysis of FUBP3-IP using the EZ-Magna RIP Kit (Millipore, #17-701) in Jurkat-D6 cells. Native ChIP-Seq (NChIP) analysis of RNAPII and FUBP3 in J-Lat 10.6 cells with TNFa stimulation or without stimulation (MOCK).