Fig. 6. TASOR interacts and cooperates with nuclear RNA degradation factors.
a TASOR is a nuclear protein. Cytoplasmic and nuclear protein extracts from HeLa HIV-1 LTR-ΔTAR-Luc cells were loaded on a SDS-PAGE gel. GAPDH and MATR3 are markers of the cytoplasmic and nuclear fractions respectively (n > 3). b Endogenous TASOR interacts with the endogenous, nuclear CNOT1, and its partners CNOT7, YTHDF2, the endogenous TRAMP-like/NEXT/PAXT component MTR4, and the endogenous RNA exosome factor EXOSC10. HeLa HIV-1 LTR-ΔTAR-Luc cells were fractionated and endogenous TASOR immunoprecipitation was performed in the nuclear fraction. Lamin B1 is negative control (n > 3). c Reverse immunoprecipitation confirmed the interaction between endogenous CNOT1 and MTR4 proteins with endogenous TASOR in the nucleus. HeLa HIV-1 LTR-ΔTAR-Luc cells were fractionated and endogenous CNOT1 and MTR4 immunoprecipitation was performed in the nuclear fraction. The asterisk shows the SUPT6H band (n = 3). d TASOR interacts with the known CNOT1 partners in an RNA-dependent manner. DDK and TASOR-DDK vectors were transfected in HeLa HIV-1 LTR-ΔTAR-Luc cells. After 48 h, lysates were treated or not with RNase A for 30 min at room temperature. Anti-DDK immunoprecipitation was then performed. All lanes are from the same gel (n > 3). e TASOR cooperates with the nuclear RNA destabilization/degradation factors. After 72 h of siRNA transfections in HeLa HIV-1 LTR-ΔTAR-Luc, cells were lysed and luciferase activity was measured and normalized on protein concentration (n = 5 for the siRNA TASOR and siRNA CNOT1 conditions; n = 3 for other siRNA transfections; each color represents one different independent experiment, mean and SEM are shown). f Schematic representation of RNA-mediated interactions between TASOR and CNOT1 and its partners: the TRAMP-like/NEXT/PAXT component MTR4, the Nuclear Exosome, and the m6A reader YTHDF2. Source data are provided as a Source data file.