TREX1 is a potent 3' → 5' exonuclease that degrades single- and double-stranded 
DNA (ssDNA and dsDNA). TREX1 mutations at amino acid positions Asp-18 and 
Asp-200 in familial chilblain lupus and Aicardi-Goutières syndrome elicit 
dominant immune dysfunction phenotypes. Failure to appropriately disassemble 
genomic DNA during normal cell death processes could lead to persistent DNA 
signals that trigger the innate immune response and autoimmunity. We tested this 
concept using dsDNA plasmid and chromatin and show that the TREX1 exonuclease 
locates 3' termini generated by endonucleases and degrades the nicked DNA 
polynucleotide. A competition assay was designed using TREX1 dominant mutants 
and variants to demonstrate that an intact DNA binding process, coupled with 
dysfunctional chemistry in the active sites, explains the dominant phenotypes in 
TREX1 D18N, D200N, and D200H alleles. The TREX1 residues Arg-174 and Lys-175 
positioned adjacent to the active sites act with the Arg-128 residues positioned 
in the catalytic cores to facilitate melting of dsDNA and generate ssDNA for 
entry into the active sites. Metal-dependent ssDNA binding in the active sites 
of the catalytically inactive dominant TREX1 mutants contributes to DNA 
retention and precludes access to DNA 3' termini by active TREX1 enzyme. Thus, 
the dominant disease genetics exhibited by the TREX1 D18N, D200N, and D200H 
alleles parallel precisely the biochemical properties of these TREX1 dimers 
during dsDNA degradation of plasmid and chromatin DNA in vitro. These results 
support the concept that failure to degrade genomic dsDNA is a principal pathway 
of immune activation in TREX1-mediated autoimmune disease.