The role of nuclear mechano-transduction in regulating gene expression and shaping the spatial landscape in glioblastoma - Glioblastoma (GBM) is the most common malignant primary brain tumor, with a median survival of 12-15 months. Despite surgical resection and adjuvant therapy, the tumor inevitably recurs. GBM transcriptional states, which are diverse and driven by distinct gene expression programs, are critical determinants of the response to therapy and prognosis. To find new treatments to block GBM progression, we need a better understanding of gene expression regulation which underlies the diverse transcriptional states in GBM. In the proposed work, we will investigate how physical force regulates GBM transcriptional states, which is an understudied area of gene expression regulation in GBM. Physical force in the form of tissue stiffness is transmitted from the cytoplasm to the nucleus and leads to changes in gene expression, among other biological effects, in a process called nuclear mechano-transduction (NMT). We found that the level of Lamin A/C, a key structural component of the nuclear lamina, controls the threshold for NMT. Our overarching hypothesis is that changing NMT will alter GBM transcriptional states and GBM progression. Several facts support our hypothesis: 1- GBM cells exhibit significant nuclear membrane irregularities, which is a sign of abnormal force transmission on the nucleus. 2- Specific GBM transcriptional states are differentially spatially localized across anatomic locations with varying degrees of tissue stiffness – and thereby force transmitted onto GBM nuclei. And 3- The Astrocyte-like/mesenchymal GBM transcriptional state, which is associated with treatment resistance, can be induced by signaling via hippo (YAP1) signaling, which can be activated by NMT. The goal of the study is to dissect the roles of NMT in GBM gene expression and prognosis. In aim #1, we will leverage the diverse landscape of tissue stiffness in the infiltrated human brain, including the GBM core and periphery, to define the correlation between NMT and GBM transcriptional states. We will further use mouse xenograft models of de-identified patient-derived GBM cell lines to quantify the effects of tissue anatomy – and thereby stiffness – on GBM transcriptional states. In aim #2, we will control NMT levels in human GBM cell lines by increasing or decreasing Lamin A/C and measure the changes in GBM transcriptional states across the landscape of the infiltrated brain in mouse xenografts GBM models. Using LMNA mutants that do not increase NMT, we will control for the NMT- independent effects of Lamin A/C on gene expression. A dominant-negative KASH construct that abolishes NMT will serve as a negative control. We will monitor the activity of NMT using a YAP1 reporter and measure the changes in animal survival, and multiple transcriptional and histopathologic readouts of tumor progression. This study will open a new avenue of investigation into the gene expression regulation in GBM and identify novel mechanisms that can guide the development of therapeutic strategies against GBM.
