ena-STUDY-WTCMP-03-08-2017-14:39:51:409-1611
Trypanosoma brucei ribonuclease H2A is an essential enzyme that resolves R-loops associated with transcription initiation and antigenic variation
In every cell ribonucleotides inserted into DNA genomes represent a threat to the stability and transmission of the genetic material. Two types of Ribonuclease H (RNase H) enzyme are found in most organisms and play key roles tackling ribonucleotides in DNA, either through excision, when the ribonucleotides form part of the DNA strand, or by hydrolysing RNA when it base-pairs with DNA, in structures termed R-loops. Surprisingly, whereas loss of either RNase H1 or RNase H2 is embryonically lethal in mammals, yeast can prosper in the absence of both enzymes. In the protozoan parasite Trypanosoma brucei, loss of RNase H1 is also tolerated but no work has examined the function of RNase H2. Here we show that loss of the catalytic subunit of T. brucei RNase H2 (TbRH2A) leads to growth and cell cycle arrest, which is concomitant with pronounced accumulation of damage in the parasite cell nucleus. Genome-wide mapping reveals that TbRH2A loss results in the combined accumulation of DNA damage and loss of R-loops specifically at sites of RNA polymerase (Pol) II transcription initiation, revealing a novel and critical role for RNase H2. In addition, we detail differential gene expression of both RNA Pol I and II transcribed genes after TbRH2A loss, including patterns that may relate to cytosolic DNA accumulation in humans with autoimmunity caused by RNase H2 mutation. Finally, we show that TbRH2A loss causes R-loop and DNA damage accumulation in telomeric RNA Pol I transcription sites used to express variant surface glycoprotein, undermining immune evasion by antigenic variation. Thus, we demonstrate a separation of function between the two nuclear T. brucei RNase H enzymes during RNA Pol II transcription, but overlap in function during RNA Pol I-mediated gene expression in a critical strategy for parasite-host interaction.
In every cell ribonucleotides inserted into DNA genomes represent a threat to the stability and transmission of the genetic material. Two types of Ribonuclease H (RNase H) enzyme are found in most organisms and play key roles tackling ribonucleotides in DNA, either through excision, when the ribonucleotides form part of the DNA strand, or by hydrolysing RNA when it base-pairs with DNA, in structures termed R-loops. Surprisingly, whereas loss of either RNase H1 or RNase H2 is embryonically lethal in mammals, yeast can prosper in the absence of both enzymes. In the protozoan parasite Trypanosoma brucei, loss of RNase H1 is also tolerated but no work has examined the function of RNase H2. Here we show that loss of the catalytic subunit of T. brucei RNase H2 (TbRH2A) leads to growth and cell cycle arrest, which is concomitant with pronounced accumulation of damage in the parasite cell nucleus. Genome-wide mapping reveals that TbRH2A loss results in the combined accumulation of DNA damage and loss of R-loops specifically at sites of RNA polymerase (Pol) II transcription initiation, revealing a novel and critical role for RNase H2. In addition, we detail differential gene expression of both RNA Pol I and II transcribed genes after TbRH2A loss, including patterns that may relate to cytosolic DNA accumulation in humans with autoimmunity caused by RNase H2 mutation. Finally, we show that TbRH2A loss causes R-loop and DNA damage accumulation in telomeric RNA Pol I transcription sites used to express variant surface glycoprotein, undermining immune evasion by antigenic variation. Thus, we demonstrate a separation of function between the two nuclear T. brucei RNase H enzymes during RNA Pol II transcription, but overlap in function during RNA Pol I-mediated gene expression in a critical strategy for parasite-host interaction.