The sialoglycoprotein podocalyxin is absent in normal pancreas but is overexpressed in pancreatic cancer and is associated with poor clinical outcome. Here, we investigate the role of podocalyxin in migration and metastasis of pancreatic adenocarcinomas using SW1990 and Pa03c as cell models. Although ezrin is regarded as a cytoplasmic binding partner of podocalyxin that regulates actin polymerization via Rac1 or RhoA, we did not detect podocalyxin-ezrin association in pancreatic cancer cells. Moreover, depletion of podocalyxin did not alter actin dynamics or modulate Rac1 and RhoA activities in pancreatic cancer cells. Using mass spectrometry, bioinformatics analysis, coimmunoprecipitation, and pull-down assays, we discovered a novel, direct binding interaction between the cytoplasmic tail of podocalyxin and the large GTPase dynamin-2 at its GTPase, middle, and pleckstrin homology domains. This podocalyxin-dynamin-2 interaction regulated microtubule growth rate, which in turn modulated focal adhesion dynamics and ultimately promoted efficient pancreatic cancer cell migration via microtubule-and Src-dependent pathways. Depletion of podocalyxin in a hemispleen mouse model of pancreatic cancer diminished liver metastasis without altering primary tumor size. Collectively, these findings reveal a novel mechanism by which podocalyxin facilitates pancreatic cancer cell migration and metastasis. Significance: These findings reveal that a novel interaction between podocalyxin and dynamin-2 promotes migration and metastasis of pancreatic cancer cells by regulating microtubule and focal adhesion dynamics.
Highlights d Rhabdoid cell lines and tumors have few mutations yet highly express a range of RTKs d RTKs and SHP2 are vulnerabilities in small-molecule and CRISPR-knockout screens d RTK inhibitors are effective against a xenografted rhabdoid mouse model in vivo d Perturbational screens may identify vulnerabilities not detectable in genomic analyses
Renewing tissues have the remarkable ability to maintain both proliferative progenitor and specialized mature cell types at consistent ratios. How are complex milieus of microenvironmental signals interpreted to coordinate tissue cell-type composition? Here, we develop a high-throughput approach, combining organoid technology, combinatorial perturbations, and quantitative imaging to address these questions in the context of the intestinal epithelium. We find that changes in proliferation of transit-amplifying (TA) cells, but not Lgr5 + stem cells, alters the composition of mature secretory and absorptive cell-types in organoids and in vivo. The link between TA proliferation and mature fate bias arises from differential amplification of secretory and absorptive progenitor cells. Further, TA cells have a distinct pattern of regulation from other epithelial celltypes that stems, in part, from signal integration via the MEK-Erk pathway and paracrine BMP production. These results demonstrate that TA cells, a feature of many renewing tissues, play a critical role in converting complex microenvironmental signals into changes in cell-type composition.
Gastrointestinal toxicity is a major concern in the development of drugs. Here, we establish the ability to use murine small and large intestine-derived monolayers to screen drugs for toxicity. As a proof-of-concept, we applied this system to assess gastrointestinal toxicity of ~50 clinically used oncology drugs, encompassing diverse mechanisms of action. Nearly all tested drugs had a deleterious effect on the gut, with increased sensitivity in the small intestine. The identification of differential toxicity between the small and large intestine enabled us to pinpoint differences in drug uptake (antifolates), drug metabolism (cyclophosphamide) and cell signaling (EGFR inhibitors) across the gut. These results highlight an under-appreciated distinction between small and large intestine toxicity and suggest distinct tissue properties important for modulating drug-induced gastrointestinal toxicity. The ability to accurately predict where and how drugs affect the murine gut will accelerate preclinical drug development.
Cancer persister cells are able to survive otherwise lethal doses of drugs through nongenetic mechanisms, which can lead to cancer regrowth and drug resistance. The broad spectrum of molecular differences observed between persisters and their treatment-naïve counterparts makes it challenging to identify causal mechanisms underlying persistence. Here, we modulate environmental signals to identify cellular mechanisms that promote the emergence of persisters and to pinpoint actionable vulnerabilities that eliminate them. We found that interferon-γ (IFNγ) can induce a pro-persistence signal that can be specifically eliminated by inhibition of type I protein arginine methyltransferase (PRMT) (PRMTi). Mechanistic investigation revealed that signal transducer and activator of transcription 1 (STAT1) is a key component connecting IFNγ’s pro-persistence and PRMTi’s antipersistence effects, suggesting a previously unknown application of PRMTi to target persisters in settings with high STAT1 expression. Modulating environmental signals can accelerate the identification of mechanisms that promote and eliminate cancer persistence.
<p>PODXL knockdown suppresses chemotactic migration of pancreatic cancer cells inside PDMS-based microchannels of prescribed widths. Representative scramble control (top) and PODXL-KD (bottom) cells migrating inside microchannels of 200 µm in length, 10µm in height, and either 6, 10, 20 or 50 µm in width. Scale bar represents 50µm. Related to Fig. 1E.</p>
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