Comparison of Fnip1^−/− and tamoxifen-induced Flcn deficient mice. (A) BM B-cell analysis in Flcn^f/+ERCre⁺ and Flcn^f/−ERCre⁺ mice 4 weeks posttamoxifen (TM) injection. Cell samples were stained with CD43 (FITC) and B220 (PE). (B) Flcn^f/−ERCre⁺ mice treated with tamoxifen (right micrograph) show hyperplastic cystic lesions (denoted with arrowheads) in kidney. Conversely, Fnip^−/− mice rarely display cysts (left micrograph). Scale bar = 200 μm. (C) Western blot analysis of kidney cell lysates from Fnip1^+/−, Fnip1^−/−, and Flcn^−/− mice. mTOR activation was assessed via phosphorylation of mTOR serines 2481 and 2448, as well as phosphorylation of ribosomal protein S6, serines 240/244 and 235/236. GAPDH expression was used as a loading control. (D) Failure to rescue peripheral CD19⁺ B lymphocytes in Fnip1^−/− mice treated with buffer or rapamycin (2 mg/kg) for 3 weeks. Each fluorescence-activated cell sorter (FACS) analysis is representative of at least 3 independent experiments.