The nucleus is the organelle which must properly transduce or resist biophysical forces to dictate the spatial organization of the genome, which in turn determines the expression profile of the cell. Initial force experiments have revealed that lamins are a major structural and resistive component of the nucleus and are located inside the inner nuclear envelope. Unlike lamins, it is unknown if heterochromatin, which is tethered to the nuclear periphery and lamins, or chromatin in general contributes to the mechanical or structural properties of nuclei. Mutation or altered amounts of lamins and heterochromatin accompany aberrant nuclear morphology in many major human diseases. These diseases include laminopathies, based in mutations in lamins, which are focused in cells or tissues that incur constant mechanical strain, such as the cardiovascular system or muscles. It is unclear how mutant lamin and altered chromatin in these diseases or other diseases each contribute to alterations in nuclear mechanics and structure. I propose to use microdissection, micromanipulation, and nanonewton-level force measurement in conjunction with live cell imaging to probe the physical properties of mouse and human nuclei. First, I propose to provide a comprehensive analysis of nuclear properties in wild-type cells by depleting or altering each component (chromatin or lamin protein) and analyzing force and structural response. Preliminary evidence suggests that chromatin contributes to the resistive properties of the nucleus: increasing the amount of decompact euchromatin using a histone deacetylase (HDAC) inhibitor results in a more compliant nucleus as quantified by micromanipulation. Second, I plan to use cells of patients with a premature aging laminopathy disease Hutchinson-Gilford progeria syndrome (HGPS) with a common mutation in lamin A called progerin to investigate changes in the physical properties of nuclei with nuclear blebs. Nuclear blebs are a phenotype common to laminopathies, cancer and aging. In all of these pathologies the nucleus becomes disfigured with multiple lobes that extend away from the nucleus. The mechanics underlying this phenotype are unknown. I will determine the contributions of accumulation of mutant lamin A progerin and decreased heterochromatin to the physical properties of nuclei with nuclear blebs. Through analyzing the contribution of each component as well as the interplay between them I aim to uncover the mechanistic basis behind changes in nuclear properties relevant to human disease.
The nucleus protects the organization of the genome which in turn determines genome expression and cellular behavior/function. Mutations or alterations in chromatin and lamins, the major components of the nucleus, result in the nucleus becoming deformed in many human diseases including laminopathies (premature aging, muscular dystrophy, heart disease and diabetes) and cancer. I will determine the contribution of chromatin and lamins to the mechanical and structural properties of the nucleus and how each is altered in laminopathies to provide significant insight into human diseases.
|Stephens, Andrew D; Banigan, Edward J; Adam, Stephen A et al. (2017) Chromatin and lamin A determine two different mechanical response regimes of the cell nucleus. Mol Biol Cell 28:1984-1996|
|Banigan, Edward J; Stephens, Andrew D; Marko, John F (2017) Mechanics and Buckling of Biopolymeric Shells and Cell Nuclei. Biophys J 113:1654-1663|