Gene-Targeted Mice Lacking the Trex1 (DNase III) 3′→5′ DNA Exonuclease Develop Inflammatory Myocarditis

Abstract

TREX1, originally designated DNase III, was isolated as a major nuclear DNA-specific 335 exonuclease that is widely distributed in both proliferating and nonproliferating mammalian tissues. The cognate cDNA shows homology to the editing subunit of the Escherichia coli replicative DNA polymerase III holoenzyme and encodes an exonuclease which was able to serve a DNA-editing function in vitro, promoting rejoining of a 3 mismatched residue in a reconstituted DNA base excision repair system. Here we report the generation of genetargeted Trex1؊/؊ mice. The null mice are viable and do not show the increase in spontaneous mutation frequency or cancer incidence that would be predicted if Trex1 served an obligatory role of editing mismatched 3 termini generated during DNA repair or DNA replication in vivo. Unexpectedly, Trex1 ؊/؊ mice exhibit a dramatically reduced survival and develop inflammatory myocarditis leading to progressive, often dilated, cardiomyopathy and circulatory failure.Two distinct nuclear exonucleases account for the major part of the total exonucleolytic activity on DNA observed in mammalian cell extracts (24, 25). They were identified as a 3Ј35Ј exonuclease acting preferentially on single-stranded DNA and a 5Ј33Ј exonuclease specific for double-stranded DNA that could remove a single-stranded 5Ј overhang as an oligonucleotide. These nuclear enzymes were designated DNase III and DNase IV, as they are distinct from the pancreatic and macrophage lysosomal DNA endonucleases DNase I and DNase II. DNase IV was later renamed flap endonuclease 1 (FEN1) (23), and its main function is processing displaced 5Ј single strands that arise during lagging-strand DNA replication, as well as during DNA repair, recombination, and triplet repeat expansion. The elimination of Fen1 activity leads to early embryonic lethality in mice, consistent with an essential role of the enzyme in DNA replication (20). In contrast, DNase III is expressed at similar levels in nonproliferating and proliferating tissues; it was isolated as the major nuclear 3Ј35Ј DNA exonuclease from the adult rabbit liver (14) and also from the calf thymus and human myoblasts, where it was designated TREX1 (37, 38).The human TREX1/DNase III cDNA (14, 29) shares amino acid sequence homology with the Escherichia coli DnaQ/MutD editing subunit of the replicative DNA polymerase III holoenzyme, which can increase the fidelity of an exonuclease-deficient mammalian DNA polymerase in vitro (36). The TREX1/ DNase III cDNA encodes a nonprocessive 3Ј35Ј DNA-specific exonuclease, with a preference for single-stranded DNA or mispaired 3Ј termini; like the native protein isolated from mammalian cells, it forms homodimers (14, 29, 30). Since two of the major mammalian nuclear DNA polymerases, Pol␣ and Pol␤, do not have an intrinsic 3Ј exonuclease function, it was proposed that TREX1/DNase III may serve to edit mismatched deoxyribonucleotides during lagging-strand DNA synthesis or gap filling in DNA base excision repair, which are conducted by Pol␣ and Pol␤, respectively...

Ribonuclease H2 mutations induce a cGAS/STING‐dependent innate immune response

“…The twofold to fourfold induction of ISGs we observed in Rnaseh2b A174T/A174T heart tissue is comparable to the fourfold to sevenfold induction seen in Samhd1 −/− mouse tissue (Rehwinkel et al , 2013). While an overt inflammatory phenotype is seen in Trex1 −/− mice (Morita et al , 2004; Gall et al , 2012), pathological signs of neuroinflammation are not evident, although ISG upregulation in brain tissue can be detected (Pereira‐Lopes et al , 2013). Given that the autoinflammatory process appears to initiate in the Trex1 −/− heart (Gall et al , 2012), there are similarities between the pattern of inflammation in Trex1 −/− mice and the ISG tissue expression pattern in Rnaseh2b A174T/A174T mice.…”

“…Samhd1 −/− mice, like Rnaseh2b A174T/A174T mice, display an ISG response in the absence of detectable pathology (Behrendt et al , 2013; Rehwinkel et al , 2013). In contrast a strong ISG response in Adar1 null or editing‐deficient mice is associated with embryonic lethality (Mannion et al , 2014; Liddicoat et al , 2015; Pestal et al , 2015), and with autoinflammatory cardiomyopathy and multi‐tissue involvement in Trex1 −/− mice (Morita et al , 2004; Stetson et al , 2008; Gall et al , 2012). The variation in severity between different AGS gene mouse models remains unexplained, although it may be meaningful that mutations in human RNASEH2B are associated with the least severe disease course, with AGS onset generally in infancy, in contrast to the prenatal/neonatal onset more commonly seen in TREX1 patients (Crow et al , 2015).…”

The role of DNA exonucleases in protecting genome stability and their impact on ageing

“…As loss-of-function (Zhang et al 2010), as such factors closely resemble 'pathogen-associated molecular patterns' (PAMPS) to which the innate immune system is tuned. Indeed, mice null for Trex1 only live for onefifth as long as wild-type mice and develop inflammatory myocarditis (Morita et al 2004), consistent with autoimmunity and/or premature ageing, since high levels of inflammation are associated with premature ageing (the 'inflamm-aging' hypothesis e.g. Franceschi et al 2007).…”

DNA Degradation and Its Defects

“…Undigested single-stranded DNA that can activate IFN-b gene in Trex1 2/2 cells (Yan et al 2010) is found in the cytoplasm of various cells in Trex1-deficient mice (Stetson et al 2008). The Trex1 2/2 mice develop a lethal, IFN-a-dependent inflammatory myocarditis (Morita et al 2004). These properties associated with the strong inflammation by IFN-b are similar to those in DNase II-null mice.…”

“…Because of its high homology with Escherichia coli DnaQ, Trex1 was originally identified as an enzyme that edits mismatched 3 0 termini generated during DNA repair and DNA synthesis (Höss et al 1999). However, Trex1 is expressed in both proliferating and nonproliferating cells, and its null mutation does not increase spontaneous mutation or tumorigenicity in mice; these observations suggest that Trex1 is either not active in base excision repair, or that it acts redundantly with other 3 0 exonucleases (Morita et al 2004). On the other hand, after studying the origin of DNA fragments accumulated in TREX1-null cells, Stetson et al (2008) proposed that TREX1 cleaves the single-stranded DNA derived from retrotransposons.…”