No abstract

We demonstrate here the functional reprogramming of a somatic cell using a nuclear and cytoplasmic extract derived from another somatic cell type. Reprogramming of 293T fibroblasts in an extract from primary human T cells or from a transformed T-cell line is evidenced by nuclear uptake and assembly of transcription factors, induction of activity of a chromatin remodeling complex, histone acetylation, and activation of lymphoid cell specific genes. Reprogrammed cells express T cell specific receptors and assemble the interleukin-2 receptor in response to T cell receptor CD3 (TCR CD3) complex stimulation. Reprogrammed primary skin fibroblasts also express T cell specific antigens. After exposure to a neuronal precursor extract, 293T fibroblasts express a neurofilament protein and extend neurite-like outgrowths. In vitro reprogramming of differentiated somatic cells creates possibilities for producing isogenic replacement cells for therapeutic applications.

Differentiation of osteoblasts from mesenchymal stem cells (MSCs) is an integral part of bone development and homeostasis, and may when improperly regulated cause disease such as bone cancer or osteoporosis. Using unbiased high-throughput methods we here characterize the landscape of global changes in gene expression, histone modifications, and DNA methylation upon differentiation of human MSCs to the osteogenic lineage. Furthermore, we provide a first genome-wide characterization of DNA binding sites of the bone master regulatory transcription factor Runt-related transcription factor 2 (RUNX2) in human osteoblasts, revealing target genes associated with regulation of proliferation, migration, apoptosis, and with a significant overlap with p53 regulated genes. These findings expand on emerging evidence of a role for RUNX2 in cancer, including bone metastases, and the p53 regulatory network. We further demonstrate that RUNX2 binds to distant regulatory elements, promoters, and with high frequency to gene 3 0 ends. Finally, we identify TEAD2 and GTF2I as novel regulators of osteogenesis. STEM CELLS

In genetically transformed plants, transgene silencing has been correlated with multiple and complex insertions of foreign DNA, e.g. T-DNA and vector backbone sequences. Occasionally, single-copy transgenes also suffer transgene silencing. We have compared integration patterns and T-DNA/plant DNA junctions in a collection of 37 single-copy T-DNA-transformed Arabidopsis lines, of which 13 displayed silencing. Vector sequences were found integrated in five lines, but only one of these displayed silencing. Truncated T-DNA copies, positioned in inverse orientation to an intact T-DNA copy, were discovered in three lines. The whole nptII gene with pnos promoter was present in the truncated copy of one such line in which heavy silencing has been observed. In the two other lines no silencing has been observed over five generations. Thus, vector sequences and short additional T-DNA sequences are not sufficient or necessary to induce transgene silencing. DNA methylation of selected restriction endonuclease sites could not be correlated with silencing. Our collection of T-DNA/plant DNA junctions has also been used to evaluate current models of T-DNA integration. Data for some of our lines are compatible with T-DNA integration in double-strand breaks, while for others initial invasion of plant DNA by the left or by the right T-DNA end seem important.