TREX1 acts in concert with the SET complex in granzyme A-mediated apoptosis, and mutations in TREX1 cause Aicardi-Goutières syndrome and familial chilblain lupus. Here, we report monoallelic frameshift or missense mutations and one 3' UTR variant of TREX1 present in 9/417 individuals with systemic lupus erythematosus but absent in 1,712 controls (P = 4.1 x 10(-7)). We demonstrate that two mutant TREX1 alleles alter subcellular targeting. Our findings implicate TREX1 in the pathogenesis of SLE.

TREX1 constitutes the major 3'-->5' DNA exonuclease activity measured in mammalian cells. Recently, biallelic mutations in TREX1 have been shown to cause Aicardi-Goutieres syndrome at the AGS1 locus. Interestingly, Aicardi-Goutieres syndrome shows overlap with systemic lupus erythematosus at both clinical and pathological levels. Here, we report a heterozygous TREX1 mutation causing familial chilblain lupus. Additionally, we describe a de novo heterozygous mutation, affecting a critical catalytic residue in TREX1, that results in typical Aicardi-Goutieres syndrome.

The TREX1 enzyme processes DNA ends as the major 3 3 5 exonuclease activity in human cells. Mutations in the TREX1 gene are an underlying cause of the neurological brain disease Aicardi-Goutières syndrome implicating TREX1 dysfunction in an aberrant immune response. TREX1 action during apoptosis likely prevents autoimmune reaction to DNA that would otherwise persist. To understand the impact of TREX1 mutations identified in patients with Aicardi-Goutières syndrome on structure and activity we determined the x-ray crystal structure of the dimeric mouse TREX1 protein in substrate and product complexes containing single-stranded DNA and deoxyadenosine monophosphate, respectively. The structures show the specific interactions between the bound nucleotides and the residues lining the binding pocket of the 3 terminal nucleotide within the enzyme active site that account for specificity, and provide the molecular basis for understanding mutations that lead to disease. Three mutant forms of TREX1 protein identified in patients with AicardiGoutières syndrome were prepared and the measured activities show that these specific mutations reduce enzyme activity by 4 -35,000-fold. The structure also reveals an 8-amino acid polyproline II helix within the TREX1 enzyme that suggests a mechanism for interactions of this exonuclease with other protein complexes.Processing of DNA ends is an important step in many DNA metabolic pathways such as replication, repair, and recombination. The 3Ј 3 5Ј exonucleases play a critical role in correcting fragmented, modified, mispaired, or even normal nucleotides to generate 3Ј termini suitable for downstream events. The drastic consequences that result from impaired 3Ј exonuclease activities underscore the importance of these enzymes for cell survival. Proofreading of DNA synthesis by 3Ј exonucleases is one of the major determinants of mutagenesis and genome stability and cells lacking this ability show a high incidence of cancers (1-3) (for review, see Ref. 4). Cells with defects in proteins containing 3Ј exonuclease activity, such as the Werner syndrome protein, MRE11, APE1, and p53 proteins display chromosomal instability, cell cycle checkpoint defects, and sensitivity to ionizing radiation (5-9).The major 3Ј 3 5Ј exonuclease activity detected in human cell extracts is catalyzed by the TREX1 enzyme. The genes encoding the TREX1 and closely related TREX2 proteins have been identified and cloned (10, 11), and the recombinant proteins confirm the robust catalytic nature of these enzymes (12, 13). Amino acid sequence analysis reveals the TREX proteins belong to the DnaQ family of 3Ј 3 5Ј exonucleases; a structurally conserved group of exonucleases that span Archaea and bacteria to humans and includes such proteins as the exonuclease domains of Werner syndrome protein, the bacterial ⑀ subunit of DNA polymerase III (⑀ subunit), and exonuclease I (Exo I) 2 (14 -17). A hallmark of the DnaQ family exonucleases is three conserved sequence motifs known as Exo I, II, and III. These motifs contain four ...

We recently described a novel autosomal-dominant genodermatosis, termed familial chilblain lupus, and mapped its genetic locus to chromosome 3p21. Familial chilblain lupus manifests in early childhood with ulcerating acral skin lesions and is associated with arthralgias and circulating antinuclear antibodies. In this study, we report the identification of a heterozygous missense mutation (D18N) in TREX1 encoding the 3'-5'repair exonuclease 1 in affected individuals of the family with chilblain lupus. The homodimeric TREX1 is the most abundant intracellular DNase in mammalian cells. We have recently shown that TREX1 plays a role in apoptotic single-stranded DNA damage induced by the killer lymphocyte protease granzyme A. D18N affects a highly conserved amino acid residue critical for catalytic activity. Recombinant mutant TREX1 homodimers are enzymatically inactive, while wild type/mutant heterodimers show residual exonucleolytic activity, suggesting a heterozygous loss of function. Lymphoblastoid cells carrying the D18N mutation are significantly less sensitive to granzyme A-mediated cell death, suggesting a novel role for this caspase-independent form of apoptosis in the pathogenesis of familial chilblain lupus. Our findings also warrant further investigation of TREX1 in common forms of lupus erythematosus.

The TREX1 gene encodes a potent DNA exonuclease, and mutations in TREX1 cause a spectrum of lupus-like autoimmune diseases. Most lupus patients develop autoantibodies to double-stranded DNA (dsDNA), but the source of DNA antigen is unknown. The TREX1 D18N mutation causes a monogenic, cutaneous form of lupus called familial chilblain lupus, and the TREX1 D18N enzyme exhibits dysfunctional dsDNA-degrading activity, providing a link between dsDNA degradation and nucleic acid-mediated autoimmune disease. We determined the structure of the TREX1 D18N protein in complex with dsDNA, revealing how this exonuclease uses a novel DNA-unwinding mechanism to separate the polynucleotide strands for single-stranded DNA (ssDNA) loading into the active site. The TREX1 D18N dsDNA interactions coupled with catalytic deficiency explain how this mutant nuclease prevents dsDNA degradation. We tested the effects of TREX1 D18N in vivo by replacing the TREX1 WT gene in mice with the TREX1 D18N allele. The TREX1 D18N mice exhibit systemic inflammation, lymphoid hyperplasia, vasculitis, and kidney disease. The observed lupus-like inflammatory disease is associated with immune activation, production of autoantibodies to dsDNA, and deposition of immune complexes in the kidney. Thus, dysfunctional dsDNA degradation by TREX1 D18N induces disease in mice that recapitulates many characteristics of human lupus. Failure to clear DNA has long been linked to lupus in humans, and these data point to dsDNA as a key substrate for TREX1 and a major antigen source in mice with dysfunctional TREX1 enzyme.