A cell's mechanical environment can have many effects, but whether it impacts a cell's DNA sequence has remained unclear. To investigate this, we developed a live-cell method to measure changes in chromosome numbers. We edited constitutive genes with GFP/RFP-tags on single alleles and discovered that cells that lose Chromosome-reporters (ChReporters) become non-fluorescent. We applied our new tools to confined mitosis and to inhibition of the putative tumor suppressor Myosin-II. We quantified compression of mitotic chromatin in vivo and demonstrated that similar compression in vitro resulted in cell death, but also rare and heritable ChReptorter loss. Myosin-II suppression rescued lethal multipolar divisions and maximized ChReporter loss in 3D-compression and 2D-confinement, but not in standard 2D-culture. ChReporter loss associated with chromosome mis-segregation, rather than just the number of divisions, and loss in vitro and in mice was selected against in subsequent 2D-cultures. Inhibition of the spindle assembly checkpoint (SAC) caused ChReporter loss in 2D, as expected, but not in 3D-compression, suggesting a SAC perturbation. Thus, confinement and myosin-II affect DNA sequence and mechano-evolution, and ChReporters enable diverse studies of viable genetic changes.
The molecular mechanisms leading to the establishment of durable immunological memory are inadequately understood, limiting the development of effective vaccines and durable anti-tumor immune therapies. Using a T cell-conditional knockout mouse model and a viral pathogen, we show that expression of the transcription cofactor OCA-B (Pou2af1/Bob.1/OBF-1) within T cells is necessary for proper CD4+ memory T cell formation. We also show that ectopic OCA-B expression is sufficient to drive T cells towards a memory fate, while having minimal effects on primary antiviral effector response. Bulk and single-cell gene expression profiling comparing cells transduced with OCA-B and empty vector at primary effector response identifies changes in gene expression consistent with later memory formation, including genes increased (Tbx21, Il7r, Gadd45b, Socs2) in specific subpopulations by ectopic OCA-B expression. Short-lived effector T cell compartments are expanded but show increased expression of Gadd45b and Socs2, while clusters of effector cells with memory potential show increased expression of Bcl2, Il7r, Tcf7 and Slamf6. We also describe an OCA-B-mCherry reporter mouse allele that selectively labels B and T lymphocytes, and shows high reporter expression in CD4+ TCM cells. We show that elevated OCA-B expression prospectively identifies cells with increased survival capability and memory recall potential. Cumulatively, the results demonstrate that OCA-B is necessary and sufficient to promote CD4 T cell memory in vivo.
Cell proliferation and migration play important parts in ovarian cancer progression. BMP9, as one of the members of the TGF-ant array of biological roles, including cell differentiation, proliferation, apoptosis, tumorigenesis, and metabolism. However, the role and mechanism of BMP9 in ovarian cancer progression remains uncertain. We found that the expression of BMP9 was increased in human ovarian cancer cell lines, which induced Notch1 intracellular domain (NICD1) accumulation. And we also found the expression abundance of BMP9 is low in ovarian cancer cells. Thus, we generated recombinant adenoviruses overexpressing BMP9 to perform the research. We found that overexpression of BMP9 promoted ovarian cancer cell proliferative viability, cell cycle progression, cell migration in vitro, and accelerated subcutaneous tumor growth in vivo, which was inhibited by dominant-negative mutant Notch1 recombinant adenoviruses. Besides, we also demonstrated that silencing BMP9 by recombinant adenoviruses inhibited ovarian cancer cell viability and migration in vitro. Additionally, BMP9-induced ovarian cancer cell progression also involved the e levation of HES2, c-Myc, MMP9, and Cyclin D1, as well as repressed expression of p27. Together, these results revealed that BMP9 acts as a promoting factor in ovarian cancer progression, and overexpression of BMP9 promotes ovarian cancer progression and growth via Notch1 signaling.Thereby our research may provide new insight into the pathogenesis of ovarian cancer and BMP9-Notch1 signaling may serve as a novel therapeutic target axis for ovarian cancer treatment.
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