SUMMARY Exosomes are secreted by all cell types and contain proteins and nucleic acids. Here, we report that breast cancer associated exosomes contain microRNAs (miRNAs) associated with the RISC Loading Complex (RLC) and display cell-independent capacity to process precursor microRNAs (pre-miRNAs) into mature miRNAs. Pre-miRNAs, along with Dicer, AGO2, and TRBP, are present in exosomes of cancer cells. CD43 mediates the accumulation of Dicer specifically in cancer exosomes. Cancer exosomes mediate an efficient and rapid silencing of mRNAs to reprogram the target cell transcriptome. Exosomes derived from cells and sera of patients with breast cancer instigate non-tumorigenic epithelial cells to form tumors in a Dicer-dependent manner. These findings offer opportunities for the development of exosomes based biomarkers and therapies.

Binding within or nearby target genes involved in cell proliferation and survival enables the p53 tumor suppressor gene to regulate their transcription and cell-cycle progression. Using genome-wide chromatin-binding profiles, we describe binding of p53 also to regions located distantly from any known p53 target gene. Interestingly, many of these regions possess conserved p53-binding sites and all known hallmarks of enhancer regions. We demonstrate that these p53-bound enhancer regions (p53BERs) indeed contain enhancer activity and interact intrachromosomally with multiple neighboring genes to convey long-distance p53-dependent transcription regulation. Furthermore, p53BERs produce, in a p53-dependent manner, enhancer RNAs (eRNAs) that are required for efficient transcriptional enhancement of interacting target genes and induction of a p53-dependent cell-cycle arrest. Thus, our results ascribe transcription enhancement activity to p53 with the capacity to regulate multiple genes from a single genomic binding site. Moreover, eRNA production from p53BERs is required for efficient p53 transcription enhancement.

EDA is a facultative type III homology of human fibronectin encoded by an alternative spliced exon. The EDA+ and EDA- mRNA forms show a cell type specific distribution with their relative proportion varying during development, aging and oncogenic transformation. We have previously demonstrated that an 81 bp nucleotide sequence within the exon itself is essential for differential RNA processing. Fine mapping of cis acting elements within this region has been carried out to identify possible target sites for the modulation of alternative splicing. There are at least two short nucleotide sequences involved. Element A (GAAGAAGA) is a positive modulator for the recognition of the exon, its deletion results in constitutive exclusion of the EDA exon. Element B (CAAGG) is a negative modulator for exon recognition, its deletion results in constitutive inclusion of the EDA exon. This bipartite structure of the splicing enhancer is a novel feature of the mammalian exons.