The embryonic mouse lung is a widely used substitute for human lung development. For example, attempts to differentiate human pluripotent stem cells to lung epithelium rely on passing through progenitor states that have only been described in mouse. The tip epithelium of the branching mouse lung is a multipotent progenitor pool that self-renews and produces differentiating descendants. We hypothesized that the human distal tip epithelium is an analogous progenitor population and tested this by examining morphology, gene expression and in vitro self-renewal and differentiation capacity of human tips. These experiments confirm that human and mouse tips are analogous and identify signalling pathways that are sufficient for long-term self-renewal of human tips as differentiation-competent organoids. Moreover, we identify mouse-human differences, including markers that define progenitor states and signalling requirements for long-term self-renewal. Our organoid system provides a genetically-tractable tool that will allow these human-specific features of lung development to be investigated.DOI: http://dx.doi.org/10.7554/eLife.26575.001

Transcription of microRNAs (miRNAs) is thought to be regulated similarly to that of protein-coding genes. However, how miRNAs are regulated during the cell division cycle is not well understood. We have analyzed the transcription profiles of miRNAs in response to mitogenic stimulation in primary fibroblasts. About 33% of the miRNAs expressed in these cells are induced upon exit from quiescence. Many of these miRNAs are specifically induced by E2F1 or E2F3 during the G 1 /S transition and are repressed in E2F1/3-knockout cells. At least four miRNA clusters, let-7a-d, let-7i, mir-15b-16-2, and mir-106b-25, are direct targets of E2F1 and E2F3 during G 1 /S and are repressed in E2F1/3-null cells. Interestingly, these miRNAs do not contribute to E2F-dependent entry into S phase but rather inhibit the G 1 /S transition by targeting multiple cell cycle regulators and E2F targets. In fact, E2F1 expression results in a significant increase in S-phase entry and DNA damage in the absence of these microRNAs. Thus, E2F-induced miRNAs contribute to limiting the cellular responses to E2F activation, thus preventing replicative stress. Given the known function of E2F of inducing other oncogenic miRNAs, control of miRNAs by E2F is likely to play multiple roles in cell proliferation and in proliferative diseases such as cancer.MicroRNAs (miRNAs) are small (ϳ23-nucleotide [nt]) regulatory RNA molecules that exert posttranscriptional control of specific target mRNAs (3). More than half of the human protein-coding genes appear to have been under selective pressure to maintain pairing of their 3Ј untranslated regions (3Ј-UTRs) to miRNAs (5). This interaction is known to induce mRNA degradation or inhibition of translation through specific albeit imperfect base pairing (22,24). Several target genes have been validated, indicating that each individual miRNA can target a few or, possibly, multiple genes. These small miRNAs therefore regulate fundamental cellular processes such as proliferation, differentiation, or apoptosis during development (65). In addition, deregulation of miRNAs is frequently observed in a wide range of diseases, including cancer (19,21). The analysis of miRNA function and regulation in specific cellular processes has proven to be required for a full understanding of malignant transformation and to envision new therapeutic possibilities.Tumor development is accompanied by a variety of genetic and epigenetic alterations in protein-coding genes and small, noncoding RNA genes. By regulating specific oncogenes or tumor suppressor molecules, miRNAs may have profound effects on tumor development. The first report linking miRNAs and cancer showed a frequent deletion of miR-15a and miR-16-1 in patients with B-cell chronic lymphocytic leukemia (13). Further analysis of miRNA expression signatures has suggested a frequent involvement of miRNAs in the initiation and progression of human tumors (12). Although the critical targets for most cancer-associated miRNAs are unknown, some relevant targets have been characterized in sp...

E2F transcription factors control diverse biological processes through regulation of target gene expression. However, the mechanism by which this regulation is established, and the relative contribution of each E2F member are still poorly defined. We have investigated the role of E2F2 in regulating cellular proliferation. We show that E2F2 is required for the normal G 0 /G 1 phase because targeted disruption of the E2F2 gene causes T cells to enter S phase early and to undergo accelerated cell division. A large set of E2F target genes involved in DNA replication and cell cycle progression (such as Mcm's, cyclins and Cdc2a) that are silent in G 0 and typically transcribed late in G 1 phase are already actively expressed in quiescent T cells and MEFs lacking E2F2. The classic E2F activators, E2F1 and E2F3, are largely dispensable for this process because compound loss of E2F1 -/-and E2F2 -/-produces a comparably shortened G 0 /G 1 phase, with early S phase entry. Likewise, shRNA knockdown of E2F3 does not alter significantly the E2F2 -/-phenotype. Chromatin immunoprecipitation analysis indicates that in wild-type cells the promoters of the aberrantly early-transcribed genes are occupied by E2F2 in G 0 , suggesting a direct role for E2F2 in transcriptional repression. We conclude that E2F2 functions to transcriptionally repress cell cycle genes to establish the G 0 state.