Mechanistic and Epigenetic Partitioning of Lamina-Associated Chromatin Revealed by a Genome-Wide Imaging Screen

The nuclear periphery has long been recognized as a site of heterochromatin organization, where lamina-associated domains (LADs) play a crucial role in transcriptional repression and genome stability. However, the molecular mechanisms that tether LADs in human cells remain poorly defined. To address this gap, I conducted the first genome-wide, imaging-based siRNA screen to identify regulators of LAD positioning using HiDRO (high-throughput DNA or RNA labeling with optimized Oligopaints). Completion of the screen led to the identification of 101 genes required for proper perinuclear localization. Interestingly, our hits included a striking enrichment of RNA-binding and processing proteins, suggesting an unappreciated role in maintaining the spatial organization of heterochromatin. Among these candidates, the RNA-binding protein hnRNPK emerged as a central factor, required for positioning nearly two-thirds of LADs across the human genome. Knockdown of hnRNPK triggered widespread detachment of LADs from the nuclear periphery without altering the underlying heterochromatin state of these regions. Despite this, the observed detachment induced transcriptional dysregulation within these domains. Further analysis revealed that hnRNPK-sensitive LADs are uniquely enriched for H3K27me3, distinguishing them from hnRNPK-insensitive LADs, which are predominantly marked by H3K9me3. Collectively, these findings uncover a pivotal role for hnRNPK in spatial genome organization, highlight the broader diversity of LAD localization mechanisms, and establish a framework for systematically mapping genetic control of nuclear architecture.

Author dc:creator

• Walsh, Patrick, John

Advisor dc:contributor.advisor

• Joyce, Eric, F

• Genetics and Genomics

Language dc:language.iso

en

Repository record dc:identifier.uri

https://repository.upenn.edu/handle/20.500.14332/62704

OAI identifier oai:identifier

oai:repository.upenn.edu:20.500.14332/62704