Abstract
Aicardi–Goutières syndrome (AGS) is an inflammatory encephalopathy caused by defective nucleic acids metabolism. Over 50% of AGS mutations affect RNase H2 the only enzyme able to remove single ribonucleotidemonophosphates (rNMPs) embedded in DNA. Ribonucleotide triphosphates (rNTPs) are incorporated into genomic DNA with relatively high frequency during normal replication making DNA more susceptible to strand breakage and mutations. Here we demonstrate that human cells depleted of RNase H2 show impaired cell cycle progression associated with chronic activation of post-replication repair (PRR) and genome instability. We identify a similar phenotype in cells derived from AGS patients, which indeed accumulate rNMPs in genomic DNA and exhibit markers of constitutive PRR and checkpoint activation. Our data indicate that in human cells RNase H2 plays a crucial role in correcting rNMPs misincorporation, preventing DNA damage. Such protective function is compromised in AGS patients and may be linked to unscheduled immune responses. These findings may be relevant to shed further light on the mechanisms involved in AGS pathogenesis.
Highlights
Aicardi–Goutieres syndrome (AGS) is a rare and underdiagnosed inflammatory encephalopathy with infancy onset and characterized by high levels of Type I interferon (IFN) production
We identify a similar phenotype in cells derived from Aicardi – Goutieres syndrome (AGS) patients, which accumulate rNMPs in genomic DNA and exhibit markers of constitutive post-replication repair (PRR) and checkpoint activation
To achieve a stable down-regulation of RNase H2, we generated a variety of lentiviral vectors carrying shRNA sequences designed to interfere with the expression of either the catalytic A subunit or the structural B subunit, which are connected, respectively, to the most severe phenotype or the most common mutations found in AGS patients [30]
Summary
Aicardi–Goutieres syndrome (AGS) is a rare and underdiagnosed inflammatory encephalopathy with infancy onset and characterized by high levels of Type I interferon (IFN) production. RNases H are specialized enzymes that process the RNA moiety in RNA : DNA hybrid molecules. These hybrid structures represent physiological intermediates produced during retroviral infection, retroelement mobilization and during genome replication, through the synthesis of Okazaki fragments or when a replication fork collides with the transcriptional machinery [5,6]. RNase H2 is a trimeric complex that, besides being able to process long RNA : DNA hybrid molecules, has the unique property of cleaving single rNMPs embedded in genomic DNA. Recent evidence revealed that ribononucleotide triphosphates (rNTPs) are misincorporated into genomic DNA with high frequency during normal replication [10,11,12].
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