Fig 2: Bmal1 deficiency augments the PAI-1-tPA/uPA-plasmin axis. A) The stromal vascular fraction (SVF) of CRC tumors grown in Bmal1+/+ and Bmal1-/- mice were isolated by FACS sorting. RNAs were extracted and subjected for sequencing. B) Volcano plot presenting differential gene expression in isolated SVF from Bmal1+/+ versus Bmal1-/- mice. Orange indicates significantly increased genes and blue indicates significantly decreased genes. C) Genome-wide expression profiling of SVF from Bmal1+/+ and Bmal1-/- tumor tissues (n = 4 samples per group). Serpine1 was identified as one of the highly down regulated genes. D) Quantification of Serpine1 mRNA level in CRC tumors, SVF, and FACS-isolated FAP+ CAFs from Bmal1+/+ and Bmal1-/- tumors (n = 3 samples per group). E) Schematic diagram of Serpine1 transcription. The Serpine1 is a direct transcriptional target of BMAL1-CLOCK heterodimers and the Serpine1 gene promoter contained canonical E-boxes of BMAL1-CLOCK binding sites. F–H) Relative PAI-1, uPA, and tPA protein levels and quantification in CRC tumors, SVF, and CAFs from Bmal1+/+ and Bmal1-/- mice (n = 4 samples per group). I) ELISA quantification of active PAI-1 in CRC tumors, SVF, CAF supernatant and plasma from tumor-bearing mice (n = 5 samples per group). J,K) Quantification of uPA and tPA activity in CRC tumors, SVF, and CAF supernatant from Bmal1+/+ and Bmal1-/- mice (n = 4 samples per group). L) Quantification of plasmin activity in CRC tumor and plasma from tumor-bearing mice (n = 6 samples per group). Data presented as mean ± S.E.M. *P < 0.05; **P < 0.01; ***P < 0.001; two-tailed student t-test.