Defining the role of the nuclear lamina in the mechanical regulation of lung fibrosis

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PROJECT SUMMARY / ABSTRACT Fibrosis is the excessive excretion of extracellular matrix (ECM) proteins, which leads to tissue stiffening and decreased organ function. Fibroblasts and myofibroblasts are the primary cell types responsible for ECM deposition. Idiopathic Pulmonary Fibrosis (IPF) is a progressive fibrotic disease of unknown origin. The role of biochemical signaling through the TGFb signaling pathway is a well-established driver of IPF and other fibrotic disease. There is also an emerging and evolving role for mechanical signaling in the regulation of ECM deposition in fibroblasts and the progression of fibrosis. The molecular mechanism by which mechanical inputs are integrated with biochemical signals to regulate fibrosis remains enigmatic. Work from our lab has implicated the Linker of Nucleoskeleton and Cytoskeleton (LINC) complex as a new regulator of fibrosis; ablation of the LINC complex component SUN2 in a Sun2-/- mouse model is protective in the context of hypertrophy-coupled cardiac fibrosis and injury-induced lung fibrosis. Given the established role of LINC complexes in transmitting mechanical force from the cell surface to the nucleus, a compelling hypothesis is that LINC complexes contribute to mechanotransduction during fibrosis progression. The overall objective of this proposal is to define the molecular mechanisms by which loss of SUN2 leads to protection from fibrosis. The proposed research will investigate this objective in two specific aims. Aim 1 will define the integration of biochemical (modulating TGFb and TGFb receptor) and mechanical (modulating cell substrate stiffness) inputs in the regulation of pro-fibrotic gene expression and attainment of pro-fibrotic phenotypes in primary human and mouse lung fibroblasts. The contexts and mechanism by which SUN proteins influence integration of pro-fibrotic signals to regulate gene expression will be interrogated using CRISPR-based gene editing, transcriptome analysis, and imaging techniques. Aim 2 will define the impact of loss of SUN proteins on both chromatin state and the established transducers of TGFb signaling, the Smad family of transcription factors, using a combination of genomic techniques and imaging approaches. This proposal will address fundamental aspects of gene regulation as it pertains specifically to the progression of lung fibrosis, in line with the mission of the National Heart, Lung, and Blood Institute. The trainee will be immersed in a supportive, collaborative and interdisciplinary environment while completing the proposed project under the rich support and mentorship of the sponsor/co-sponsor, ultimately positioning her to approach fundamental questions through a translational lens. She will improve her experimental design skills, obtain training in rigorous analysis of experimental data, and expand her technical skillset to include methods from genomics to imaging of tissue. The trainee will also have ample access to opportunities for improving oral presentation, writing, mentorship, scientific outreach skills.