Sahai, E. | Search bioRxiv

bioRxiv2017

Actin and Src-family kinases regulate nuclear YAP1 and its export

The transcriptional regulator YAP1 is critical for the pathological activation of fibroblasts. In normal fibroblasts YAP1 is predominantly located in the cytoplasm, while in activated cancer-associated fibroblasts it exhibits nuclear localization and promotes the expression of many genes required for pro-tumorigenic functions. Here, we investigate the dynamics of YAP1 shuttling in normal and activated fibroblasts, using EYFP-YAP1, quantitative photo-bleaching methods, and mathematical modeling. We find that both 14-3-3 and TEAD binding modulate YAP1 shuttling, but neither affects nuclear import. Instead, we find that YAP1 serine phosphorylation is required for nuclear export. Furthermore, YAP1 nuclear accumulation in activated fibroblasts results from Src and actomyosin-dependent suppression of phosphorylated YAP1 export. Finally, we show that nuclear constrained YAP1, upon XPO1 depletion, remains sensitive to blockade of actomyosin function. Together, these data place nuclear export at the center of YAP1 regulation and indicate that the cytoskeleton can regulate YAP1 within the nucleus.

molecular biology↗

bioRxiv2017

Dissecting targeted therapy resistance: Integrating models to quantify environment mediated drug resistance

Drug resistance is the single most important driver of cancer treatment failure for modern targeted therapies. This resistance may be due to the presence of dormant or aggressive tumor cell phenotypes or to context-driven protection. Non-malignant cells and other factors, constituting the microenvironment in which the tumor grows (the stroma), are now thought to play a crucial role in both therapeutic response and resistance. Specifically, the dialogue between the tumor and stroma has been shown to modulate the response to molecularly targeted therapies, through proliferative and survival signaling. The goal of this work is to investigate interactions between a growing tumor and its surrounding stroma in facilitating the emergence of drug resistance. We use mathematical modeling as a theoretical framework to bridge between experimental models and scales, with the aim of separating the intrinsic and extrinsic components of resistance in BRAF mutated melanoma. The model describes tumor-stroma dynamics both with and without treatment. Calibration of our model, through the integration of experimental data, revealed significant variation across animal replicates in either the intensity of stromal promotion or intrinsic tissue carrying capacity. Furthermore our study highlights the need to account for this variation in the design of treatment strategies. Major Findings. Through the integration of a simple mathematical model with in vitro and in vivo experimental growth dynamics of melanoma cell lines (both with and without drug), we were able to dissect the relative contributions of intrinsic versus environmental resistance. Our study revealed significant heterogeneity in vivo, indicating that there is a diversity of either stromal promotion or tumor carrying capacity under targeted therapy. We believe this variation may be one possible explanation for the heterogeneity observed across patients and within individual patients with multiple metastases. Therefore, quantifying this variation both within in vivo model systems and in individual patients could have a significant impact on the design of future treatment strategies that target both the tumor and stroma. Further, we present guidelines for building more effective and longer lasting therapeutic strategies utilizing our experimentally calibrated model. These strategies explicitly consider the protective nature of the stroma and utilize inhibitors that modulate it.\n\nPrecisQuantification of the environmental contribution to drug resistance reveals heterogeneity that significantly alters treatment dynamics that can be exploited for therapeutic gain.\n\nFinancial SupportPicco and Anderson: US National Cancer Institute grant U01CA151924.\n\nPicco: UK Engineering and Physical Sciences Research Council (EPSRC grant number EP/G037280/1).\n\nConflict of Interest DisclosureThe authors declare no potential conflicts of interest.

cancer biology↗