The JAK1^A634D mutant, but not JAK1^A634D/Y107A, mediates constitutive expression of type I IFN–regulated genes and promotes constitutive phosphorylation of the components of the type I IFN signal transduction pathway in BW5147 and BaF3 cells. Total RNA was extracted from 10⁶ BW5147 (A) and BaF3 (B) cells stably transduced by JAK1^WT or JAK^A634D with intact or mutated Y107A FERM domain (JAK1^A634D/Y107A). To quantify basal expression of type I IFN–induced genes, such as mIRF-7 (left) and mOasl-2 (right), quantitative RT-PCR was performed. Transcripts were normalized to the level of the murine β-actin transcripts. Results are mean ± SD of biologic triplicates. Similar results were obtained in 3 independent experiments. (C) BaF3 cells (10⁶) stably transduced with JAK1^WT or JAK^A634D, with intact or mutated Y107A FERM domain (JAK1^A634D/Y107A), were starved for 4 hours, lysed, and subjected to Western blot analysis. Basal phosphorylation of JAK1, TYK2, STAT1, and STAT2 was detected with specific anti-pY1022/1023 JAK1, anti-pY1054/1055 TYK2, anti-pY701 STAT1, and anti-pY689 STAT2 antibodies. Membranes were reprobed with anti-JAK1, anti-STAT1, anti-STAT2, and anti–β-actin antibodies as control. Similar results were obtained in 3 independent experiments for the BaF3 and BW5147 cell lines. WT indicates wild-type.