Multiscale light-sheet organoid imaging framework.

Organoids provide an accessible in vitro system to mimic the dynamics of tissue regeneration and development. However, long-term live-imaging of organoids remains challenging. Here we present an experimental and image-processing framework capable of turning long-term light-sheet imaging of intestinal organoids into digital organoids.

Phenotypic landscape of intestinal organoid regeneration.

The development of intestinal organoids from single adult intestinal stem cells in vitro recapitulates the regenerative capacity of the intestinal epithelium. Here we unravel the mechanisms that orchestrate both organoid formation and the regeneration of intestinal tissue, using an image-based screen to assay an annotated library of compounds. We generate multivariate feature profiles for hundreds of thousands of organoids to quantitatively describe their phenotypic landscape.

Memo has a novel role in S1P signaling and is [corrected] crucial for vascular development.

Memo is a conserved protein that was identified as an essential mediator of tumor cell motility induced by receptor tyrosine kinase activation. Here we show that Memo null mouse embryonic fibroblasts (MEFs) are impaired in PDGF-induced migration and this is due to a defect in sphingosine-1-phosphate (S1P) signaling. S1P is a bioactive phospholipid produced in response to multiple stimuli, which regulates many cellular processes.

Memo is a cofilin-interacting protein that influences PLCgamma1 and cofilin activities, and is essential for maintaining directionality during ErbB2-induced tumor-cell migration.

Heregulin (HRG) activates ErbB2-ErbB3 heterodimers thereby stimulating many cellular responses, including motility. Memo and PLCgamma1 interact with ErbB2 autophosphorylation sites and are essential for HRG-induced chemotaxis. By tracing HRG-stimulated cell migration in Dunn chambers, we found that Memo- or PLCgamma1 knockdown (KD) strongly impairs cell directionality.

Novel c-MYC target genes mediate differential effects on cell proliferation and migration.

The developmental and oncogenic roles of MYC proteins are well established, but the transcriptional targets mediating their functions remain elusive. Using small interfering RNA-mediated knockdown in breast and cervix carcinoma cell lines, which overexpress c-MYC, we show that c-MYC independently controls metabolism and cell proliferation, and can, depending on the cells, promote or inhibit migration. We identified new c-MYC target genes in these cell lines, and show that selective regulation of some targets correlates with the phenotypic responses of these different cell lines to c-MYC depletion.

Beta1 integrins regulate mammary gland proliferation and maintain the integrity of mammary alveoli.

Integrin-extracellular matrix interactions play important roles in the coordinated integration of external and internal cues that are essential for proper development. To study the role of beta1 integrin in the mammary gland, Itgbeta1(flox/flox) mice were crossed with WAPiCre transgenic mice, which led to specific ablation of beta1 integrin in luminal alveolar epithelial cells. In the beta1 integrin mutant mammary gland, individual alveoli were disorganized resulting from alterations in cell-basement membrane associations.

The ErbB2/ErbB3 heterodimer functions as an oncogenic unit: ErbB2 requires ErbB3 to drive breast tumor cell proliferation.

ErbB2 is a receptor tyrosine kinase whose activity in normal cells depends on dimerization with another ligand-binding ErbB receptor. In contrast, amplification of c-erbB2 in tumors results in dramatic overexpression and constitutive activation of the receptor. Breast cancer cells overexpressing ErbB2 depend on its activity for proliferation, because treatment of these cells with ErbB2-specific antagonistic antibodies or kinase inhibitors blocks tumor cells in the G1 phase of the cell cycle.