Abstract P5-11-01: Epithelial-Mesenchymal Plasticity is Regulated by Inflammatory Signaling Networks Coupled to Cell Morphology

Abstract

Abstract Purpose: During development and homeostatic processes such as wound repair, certain cells undergo a remarkable process where they radically transform in cell shape and state, from epithelial to mesenchymal cells. This ability is referred to as ‘Epithelial-Mesenchymal Plasticity’ (EMP), triggered by both mechanical (i.e. loss of cell-cell contact) and soluble cues (i.e. TGFβ), and is absolutely essential in both embryonic and adult organisms. Dysregulation of EMP also occurs in cancer; where tumor cells undergo EMP to become metastatic, stem-like, and drug resistant. Critically, increased EMP correlates with increased cancer severity. However, it is largely a black box as to how EMP is regulated and how epithelial cells sense physical, geometrical, and soluble cues in their environment to assume a mesenchymal fate. This work inferred that cell shape is a determinant of not only fates – but of long-term outcomes. That is, we provide mechanistic explanations between cell context, environment, and cancer severity. In this work, we investigated how mechanical and soluble cues are coupled to the dynamics of signaling pathways that regulate transcriptional and post-transcriptional events that underpin EMP. Especially with regards to mechanical cues, we attempt to unlock the ‘black box’ as to how changes in adhesion, ECM (Extracellular Matrix) stiffness, and environment geometry are coupled to the transcriptional events that drive EMP. Results: We show that changes in cell and nuclear shape result from the actions of the cytoskeleton and important drivers of EMP in upregulating ‘interlocking’ networks that promote EMP-driving inflammation and suppressing insulin signaling. Using a combination of cell biology, proteomics, and new statistical methods, we provide a systems biology model demonstrating: Cell shape → MT bound Kinesin-1 activity and nuclear shape → inflammation (IKK, JNK), insulin signaling (IRS), and YAP/TAZ → EMP Our work connects observable changes in phenotype to causal changes in signaling network architecture and cell fates. We used an integrative -omic approach to analyze tumors from breast cancer patients. We identified a novel tumor suppressor – JAM3 – whose loss is associated with altered nuclear shape in vivo, inhibiting JAM3 in cells, or stimulation with canonical EMP inducer TGFβ to promote EMP, and thus changes the organization of microtubules and alters nuclear shape. During EMP we observe there is upregulation of a pro-inflammatory, insulin resistant, signaling network that is predictive of mesenchymal states across cancer. EMP following JAM3 depletion and/or TGFβ stimulation is rescued by inhibition of Kinesin-1 motors. This rescue is explained by changes in inflammatory and insulin signaling. We show that while Kinesin-1 activity is responsible for upregulation in canonical signalling and network ‘hubs’, changes in nuclear shape upregulate ‘effectors’ of these hubs. Thus, microtubules and nuclei differentially regulate different parts of ‘interlocking’ networks. Conclusions: This work has integrated image-omics, comprehensive global proteomics, and quantitative cell biology to provide a mechanistic ¬and systems-level understanding of how epithelial cells differentiate into mesenchymal forms during disease development and progression. This work is of major significance for three reasons. First, it shows how cell shape can mechanistically regulate cell fates on an unprecedented systems-level. Second, we identify an EMP network that is conserved across cancers and may indeed be conserved across both normal and diseased mesenchymal cells. Indeed, we speculate that different types of diseased cells may all share the same network. Finally, we introduce the concept of interlocking networks – where hubs and effectors are regulated by different cellular components. Our work has been extensively validated, using chemical and genetic approaches and in vivo model of in human breast cancer. Citation Format: Zheng Yin. Epithelial-Mesenchymal Plasticity is Regulated by Inflammatory Signaling Networks Coupled to Cell Morphology [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr P5-11-01.