Fibrosis is a risk factor for cancer in many tissues and accompanies tumor progression. Fibrosis-associated stiffening of breast tumors is linked with increased tumor growth, invasion, and metastasis. However, the mechanisms through which tissue stiffness imparts these phenotypes on a tumor are still poorly understood, due in part to limitations in existing in vitro culture systems and methods to discern mechanically soft and stiff regions within tumors. In this dissertation, I sought to better identify physiologically-relevant mechanisms through which fibrosis drives tumor aggression by emphasizing unbiased screening approaches and utilizing recently available clinical transcriptomic datasets. These projects identified metabolic state, glycocalyx content and bulkiness, and immune cell infiltration and phenotypes as a network of changes in the tumor microenvironment driven by and feeding back into tissue fibrosis. Moreover, my collaborators and I developed a novel method for characterizing tissue rigidity and identified shortcomings in existing cell culture models, thus facilitating future research into the impact of tissue mechanics in breast cancer. Altogether, these works apply state-of-the-art sequencing and machine learning methods to shed light on the impact of tissue fibrosis and mechanics in breast cancer development and aggression.

Cell behavior is regulated by both internal and external signals. In early biological research, studies into cell signaling focused on biochemical cues. However, in recent decades, the scientific community has come to recognize the importance of biophysical cues in determining cell fate. The extracellular matrix (ECM), in particular, has emerged as a major regulator of cell behavior and is a source of both chemical and physical cues. Extracellular matrix makeup, organization, and presentation have been implicated in influencing cell growth, migration, differentiation, and more. My doctoral work focused on dissecting the molecular mechanisms underlying biophysical regulation of cell signaling. Specifically, I examined the effect of two biophysical properties of the extracellular matrix—dimensionality and rigidity—on mammary epithelial cell growth and survival signaling. My thesis details work connecting altered ECM properties to changes in the biophysical properties of the cell, which directly influence the context and dynamics of canonical growth factor receptor and GTPase signaling to reprogram cell behavior.

Engineering functional tissues and organs from stem cells requires simultaneous control over both cell fate and organization of tissue architecture. Cell-generated forces, such as cytoskeletal contractility, traction stresses, and cell-cell tension have long been recognized as critical drivers of tissue morphogenesis; nevertheless, it has remained unclear whether these physical forces also directly regulate cell fate decisions by modulating biochemical signaling pathways. In this work, we demonstrated that human embryonic stem cells (hESCs) cultured on compliant engineered substrates self-organize to form discrete gastrulation nodes that foster mesoderm specification. We identified a mechanism whereby localized high cell-adhesion tension induces mesoderm specification by enhancing Wnt/β-catenin signaling, and furthermore, we showed that application of mechanical tension via stretching is sufficient to induce mesoderm specification. Additionally, we demonstrated that cell adhesion proteins and regulators of cytoskeletal contractility control pattern formation in human induced pluripotent stem cells (hiPSCs), independent of cell fate specification. Further, we discovered that a specific Wnt ligand from the niche regulates muscle stem cell (SC) quiescence via Rho signaling that promotes SC contractility. Finally, we developed a novel protocol for performing atomic force microscopy (AFM) on mouse bone marrow to enable characterization of the biophysical properties of the hematopoietic stem cell (HSC) niche. Together, these findings underscore the interplay between tissue organization, cell-generated forces, and morphogen-dependent differentiation. This body of work demonstrates that mechanical forces play a direct role in cell fate specification during development and regulate stem cell function in the adult organism.