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Morgan Basta, Heather Paulson, Alex V Levin, Svetlana Petruk, Alexander Mazo, Janice L Walker; De-condensed structure of post-replicative chromatin is essential for cell reprogramming in fibrotic disease. Invest. Ophthalmol. Vis. Sci. 2020;61(7):3982.
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© ARVO (1962-2015); The Authors (2016-present)
The epigenetic mechanisms that underlie reprogramming to a myofibroblast cell type that induces the lens fibrotic disease, Posterior Capsule Opacification (PCO) are not understood. Here, we examined if the structure of nascent chromatin is important during progenitor cell reprogramming to a myofibroblast cell fate, and if it is possible to manipulate the state of condensation of nascent chromatin in order to prevent reprogramming to a fibrotic phenotype.
An ex-vivo mock cataract surgery PCO model was used to study cell reprogramming to an αSMA+ myofibroblast fate, which is observed by 72hr post-injury. The fibrotic phenotype was examined by immunofluorescence, western blot and/or RT-PCR analysis. The chromatin assembly assay (CAA) was used to study how chromatin structure is re-established shortly after replication during the time of cell reprogramming. To manipulate nascent chromatin state to block the fibrotic outcome, the function of the ubiquitously transcribed tetratricopeptide repeat X chromosome (UTX) H3K27me3 demethylase was inhibited in both the ex-vivo surgery and pediatric human lens explant models.
CAA revealed a delay in the accumulation of H3K27me3 on nascent DNA in 28hr post-injury, while accumulation of this modified histone was rapid in 48hr after injury. Since H3K27me3 marks the most condensed structure of chromatin in the genome, this signifies a de-condensed state of nascent chromatin during progenitor cell reprogramming into myofibroblasts. This de-condensed state of nascent chromatin may be attributed to an accumulation of UTX on nascent DNA. Thus, UTX may be essential to create a ‘window of opportunity’ for binding of Myocardin-Related Transcription Factor-A (MRTF-A) to nascent DNA, a TF known to induce αSMA expression. Indeed, binding of MRTF-A occurred prior to the induction of αSMA mRNA expression at 48hr post-injury. Pharmacological inhibition of UTX activity induces accumulation of H3K27me3 to nascent DNA, leading to condensation of nascent chromatin, which prevents MRTF-A recruitment to DNA. Blocking UTX function prevents the emergence of αSMA+ myofibroblasts in both the ex-vivo mock cataract surgery model and human pediatric lens explants.
Pharmacological inhibition of UTX activity may provide an effective strategy to manipulate nascent chromatin structure leading to a block in fibrotic transcriptional reprogramming.
This is a 2020 ARVO Annual Meeting abstract.
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