Fig. 3: RPLP1 competes with the transcription factor C/EBPβ to bind HIV-1 LTR.

a, b RPLP1 is recruited to the HIV-1 NL4-3 LTR via the C/EBPβ binding sites. Chromatin from HIV-1-infected HEK293T (a) or Jurkat (b) cells was immunoprecipitated with anti-RPLP1 antibody or the negative control IgG and analyzed by RT-qPCR with the indicated primers spanning C/EBPβ (F1/R1) or Sp1 (F2/R2) binding sites in the LTR. c, d C/EBPβ recruitment to the LTR promoter is disrupted by RPLP1. c HEK293T cells were transfected with control or Flag-RPLP1 construct. 48 h later, cells were harvested and nuclear proteins were isolated. Normalized amount of nuclear extracts were subjected for the TransAM C/EBPβ binding assay (ActiveMotif). d HEK293T cells co-transfected with Myc-C/EBPβ plus increasing doses of HA-RPLP1 were infected with HIV-1 NL4-3 and split into two parts, followed by separately subjected to ChIP assays using antibody against HA or Myc with IgG as the negative control, respectively. RT-qPCR was performed with the primers spanning the C/EBPβ binding site. e Schematic of CH167 LTR chimeric mutant. LTR-luciferase reporter construct of HIV-1 CH167 was modified by inserting two C/EBPβ binding sites. f RPLP1 inhibits LTR activity of CH167 harboring C/EBPβ binding sites. HEK293T cells were transfected with indicated constructs, and the cells were collected at 48 h post transfection. The LTR activity was detected with the activity of cells transfected without RPLP1 set as 1. Quantifications in (a–d, f) are shown as means ± SDs from three independent experiments. P values were calculated by the two-tailed Student’s t test (a–d, f). *P < 0.05, **P < 0.01, ***P < 0.001, n.s. denotes no significance. The schematic workflow in (c, d) is generated using BioRender (http://biorender.com/). See also Supplementary Fig. 3. Source data are provided as a Source Data file.