LEDGF

The transcriptional co-activator lens epithelium-derived growth factor/p75 (LEDGF/p75) plays an important role in the biology of the cell and in several human diseases, including MLL-rearranged acute leukemia, autoimmunity, and HIV-1 infection. In both health and disease, LEDGF/p75 functions as a chromatin tether that interacts with proteins such as MLL1 and HIV-1 integrase via its integrase-binding domain (IBD) and with chromatin through its N-terminal PWWP domain. Recently, dimerization of LEDGF/p75 was shown, mediated by a network of electrostatic contacts between amino acids from the IBD and the C-terminal α₆-helix. Here, we investigated the functional impact of LEDGF/p75 variants on the dimerization using biochemical and cellular interaction assays. The data demonstrate that the C-terminal α₆-helix folds back in cis on the IBD of monomeric LEDGF/p75. We discovered that the presence of DNA stimulates LEDGF/p75 dimerization. LEDGF/p75 dimerization enhances binding to MLL1 but not to HIV-1 integrase, a finding that was observed in vitro and validated in cell culture. Whereas HIV-1 replication was not dependent on LEDGF/p75 dimerization, colony formation of MLLr-dependent human leukemic THP-1 cells was. In conclusion, our data indicate that intricate changes in the quaternary structure of LEDGF/p75 modulate its tethering function.

Osteosarcoma is generally considered a cold tumor and is characterized by epigenetic alterations. Although tumor cells are surrounded by many immune cells such as macrophages, T cells may be suppressed, be inactivated, or not be presented due to various mechanisms, which usually results in poor prognosis and insensitivity to immunotherapy. Immunotherapy is considered a promising anti-cancer therapy in osteosarcoma but requires more research, but osteosarcoma does not currently respond well to this therapy. The cancer immunity cycle (CIC) is essential for anti-tumor immunity, and is epigenetically regulated. Therefore, it is possible to modulate the immune microenvironment of osteosarcoma by targeting epigenetic factors. In this study, we explored the correlation between epigenetic modulation and CIC in osteosarcoma through bioinformatic methods. Based on the RNA data from TARGET and GSE21257 cohorts, we identified epigenetic related subtypes by NMF clustering and constructed a clinical prognostic model by the LASSO algorithm. ESTIMATE, Cibersort, and xCell algorithms were applied to analyze the tumor microenvironment. Based on eight epigenetic biomarkers (SFMBT2, SP140, CBX5, HMGN2, SMARCA4, PSIP1, ACTR6, and CHD2), two subtypes were identified, and they are mainly distinguished by immune response and cell cycle regulation. After excluding ACTR6 by LASSO regression, the prognostic model was established and it exhibited good predictive efficacy. The risk score showed a strong correlation with the tumor microenvironment, drug sensitivity and many immune checkpoints. In summary, our study sheds a new light on the CIC-related epigenetic modulation mechanism of osteosarcoma and helps search for potential drugs for osteosarcoma treatment.

R-loops that accumulate at transcription sites pose a persistent threat to genome integrity. PSIP1 is a chromatin protein associated with transcriptional elongation complex, possesses histone chaperone activity, and is implicated in recruiting RNA processing and DNA repair factors to transcription sites. Here, we show that PSIP1 interacts with R-loops and other proteins involved in R-loop homeostasis, including PARP1. Genome-wide mapping of PSIP1, R-loops and γ-H2AX in PSIP1-depleted human and mouse cell lines revealed an accumulation of R-loops and DNA damage at gene promoters in the absence of PSIP1. R-loop accumulation causes local transcriptional arrest and transcription-replication conflict, leading to DNA damage. PSIP1 depletion increases 53BP1 foci and reduces RAD51 foci, suggesting altered DNA repair choice. Furthermore, PSIP1 depletion increases the sensitivity of cancer cells to PARP1 inhibitors and DNA-damaging agents that induce R-loop-induced DNA damage. These findings provide insights into the mechanism through which PSIP1 maintains genome integrity at the site of transcription.