PRP8 regulates chromatin organization to enforce transcriptional gene silencing independent of DNA methylation.

Abstract

Transcriptional gene silencing (TGS) is critical in maintaining genome integrity in plants, during which DNA methylation plays a fundamental role. The establishment and maintenance of DNA methylation are well characterized, while the downstream events mediating transcriptional repression remain poorly understood. In this study, we performed a forward genetic screen for suppressors of hdp1-induced report gene silencing to identify components involved in TGS downstream of DNA methylation and identified Pre-mRNA processing factor 8 (PRP8), a well-known component of the spliceosome complex. Through comprehensive genetic analysis coupled with RNA sequencing and whole-genome bisulfite sequencing, we demonstrated that PRP8-mediated TGS of the transgenes along with a subset of endogenous genes and transposable elements in a DNA methylation-independent manner. Through high-throughput chromosome conformation capture (Hi-C) analysis, we demonstrated that PRP8 is required for maintaining pericentromeric heterochromatin interactions and higher-order chromatin organization. Further genetic analysis revealed that PRP8 acts synergistically with Arabidopsis MICRORCHIDIA (MORC) protein MORC6, HISTONE DEACETYLASE 6 (HDA6), MORPHEUS MOLECULE 1 (MOM1), and NUCLEAR RNA POLYMERASE D1B (NRPE1), suggesting its involvement in a previously uncharacterized TGS pathway. In summary, our findings establish PRP8 as a dual-function factor involved in both splicing and DNA methylation-independent TGS.