Differential effect of the Pol III inhibitor ML-60218 in the transcription of distinct ncRNAs

GIMENEZ, MACARENA; FERNÁNDEZ ALVAREZ, ANA JULIA; BOCCACCIO, GRACIELA L

Congreso; Riboclub; 2021

Pol III is the largest RNA polymerase complex and transcribes non-coding RNAs. Two paralogs for one of the Pol III subunits, termed POLR3G and POLR3GL, exist in mammalian cells. In proliferating cells POLR3G and POLR3GL are believed to be redundant and when both POLR3G and POLR3GL are available, Pol III can incorporate either of these subunits to transcribe the majority of Pol III genes (Pietri et al., 2019). POLR3G but not POLR3GL is inhibited by ML-60218, a poorly described Pol III inhibitor. Upon exposure of mammalian cells to ML-60218, POLR3G is downregulated and POLR3GL shows increased association with the core Pol III subunit (Pietri et al., 2019). More recently, mapping studies showed that ML-60218 disrupts the occupancy of both POLR3G and POLR3GL in opposite manner. POLR3G presence is reduced in most Pol III-transcribed genes whereas POLR3GL occupancy is largely unaffected, or even increased in a number of genes (Van Bortle et al., BioRxiv 2021). Based on these observations, we speculate that transcripts that depends on POLR3G and that cannot be transcribed by POLR3GL will be downregulated by ML-60218. Genes that can be transcribed either by POLR3G or POLR3GL would show a moderate or null effect, and transcripts that are transcribed mostly by POLR3GL would show no effect, or would be upregulated by ML-60218. Here we show that ML-60218 elicits differential effects on Vault RNAs, Y4 and U6 RNAs. None of these ncRNAs showed reduced levels upon POLR3G inhibition. In contrast, Vault 1-2 and Y4 levels increased 4X after exposure to ML-60218. U6 and Vault 1-3 were not affected and Vault 1-1 levels moderately increased. These observations are compatible with a differential dependence of POLR3G and POLR3GL for the transcription of Y4 and Vault 1-2 in comparison with U6 and Vault 1-3 RNA. The immediate upregulation of Y4 and Vault 1-2 RNAs upon exposure to ML-60218 suggests that their promoters are actively transcribed by POLR3GL. We propose that the response to ML-60218 indirectly informs on the balance POLR3G/POLR3GL in the transcription of a given POL III gene.