Role of chromatin mechanics in nuclear shape and integrity - Project Summary Abnormal nuclear shape is a diagnostic marker of disease progression and contributes to cellular dysfunction. Perturbations to major nuclear rigidity components, chromatin or lamins, results in a weaker nucleus that is deformed by external and/or internal forces to produce nuclear blebs and ruptures causing cellular dysfunction. Previously, we revealed that, independent of lamins, chromatin compaction via histone modification state (eu- /heterochromatin) determines nuclear mechanics, shape, rupture, and function. Changes in heterochromatin subtypes, transcription activity, and mitotic segregation are well-known to occur in diseases presenting abnormal nuclear morphology. The overall goal of my research program is to identify how nuclear rigidity, shape, and rupture are controlled to maintain proper genomic and cellular function. The central hypothesis is that chromatin rigidity via heterochromatin subtypes, chromatin motion via transcription activity, and chromatin organization during mitotic segregation drive the mechanisms underlying nuclear physical properties. This hypothesis addresses three knowledge gaps. First, what are the specific components that provide heterochromatin rigidity? Heterochromatin is composed of subtypes, constitutive vs. facultative, and their specific histone modifications that facilitate chromatin compaction and may have distinct impacts on rigidity. Second, is there another contributor to nuclear deformation? While actin compression and contraction provide an external nuclear deformation, transcription-driven chromatin motion represents a potential new internal nuclear deformation. Third, is there an alternative mechanism to cause abnormal nuclear shape? Mitotic segregation error represents a potential new mechanism of abnormal nuclear shape and rupture through chromatin disorganization during nuclear reformation. We are qualified to determine nuclear rigidity using our lab’s innovative micromanipulation single nucleus force-extension technique that has the unique ability to separate chromatin and lamin rigidity contributions. We will couple this with measuring nuclear shape/blebbing and rupture through tracking NLS-GFP via live cell microscopy. Our preliminary data show that heterochromatin histone modifications have distinct effects on nuclear shape while transcription inhibition suppresses nuclear blebbing and rupture independent of nuclear rigidity. Our preliminary data also show that miotic segregation errors cause both abnormal nuclear morphology and rupture. This work will provide a deeper understanding of the established interphase blebbing pathway by determining heterochromatin subtypes’ distinct physical contributions and a new contributor in transcription-driven deformation. This proposal will also identify a new mechanism for abnormal shape via mitotic segregation error. Identifying the role of heterochromatin subtypes, transcription activity, and mitotic segregation error to nuclear rigidity, shape, and rupture will enlighten the underlying causes of abnormal nuclear morphology and provide therapeutic targets for restoring nuclear shape and function in human disease.
