ENAapplication/xmlftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR298/060/SRR29841760/SRR29841760_1.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR298/064/SRR29841764/SRR29841764_1.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR298/062/SRR29841762/SRR29841762_2.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR298/070/SRR29841770/SRR29841770_2.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR298/069/SRR29841769/SRR29841769_1.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR298/061/SRR29841761/SRR29841761_2.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR298/058/SRR29841758/SRR29841758_2.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR298/061/SRR29841761/SRR29841761_1.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR298/063/SRR29841763/SRR29841763_1.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR298/067/SRR29841767/SRR29841767_2.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR298/059/SRR29841759/SRR29841759_2.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR298/067/SRR29841767/SRR29841767_1.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR298/065/SRR29841765/SRR29841765_2.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR298/070/SRR29841770/SRR29841770_1.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR298/066/SRR29841766/SRR29841766_1.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR298/058/SRR29841758/SRR29841758_1.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR298/068/SRR29841768/SRR29841768_2.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR298/071/SRR29841771/SRR29841771_2.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR298/064/SRR29841764/SRR29841764_2.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR298/060/SRR29841760/SRR29841760_2.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR298/066/SRR29841766/SRR29841766_2.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR298/065/SRR29841765/SRR29841765_1.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR298/059/SRR29841759/SRR29841759_1.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR298/071/SRR29841771/SRR29841771_1.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR298/069/SRR29841769/SRR29841769_2.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR298/063/SRR29841763/SRR29841763_2.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR298/062/SRR29841762/SRR29841762_1.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR298/068/SRR29841768/SRR29841768_1.fastq.gzprimaryOK200GenomicsCITIID, University of Cambridgehttps://www.ebi.ac.uk/ena/browser/view/PRJNA1136420Homo sapiensThe human silencing hub (HUSH) complex is a transcription-dependent, epigenetic repressor complex that provides a genome-wide immunosurveillance system for the recognition and silencing of newly-integrated retroelements. The core HUSH complex of TASOR, MPP8 and Periphilin, represses these retroelements through SETDB1-mediated H3K9me3 deposition and MORC2-dependent chromatin compaction. HUSH-dependent silencing is RNA-mediated, yet no HUSH components encode any RNA-binding domain. Here we used an unbiased approach to identify which HUSH component was able to bind RNA and determine whether RNA-binding was essential for HUSH function. We identify Periphilin as the major RNA-binding component of the HUSH complex and show that Periphilin’s N-terminal domain is essential for both RNA binding and HUSH function. Periphilin binding to RNA was independent of its interaction with TASOR or MPP8, as its N-terminal domain was sufficient for RNA targeting. The artificial tethering of Periphilin, to a HUSH-insensitive, nascent transcript, enabled the HUSH-dependent silencing of the transcript. This tethering of Periphilin allowed the RNA-binding region of Periphilin to be removed such that only its C-terminal domain was required, for oligomerisation and interaction with TASOR. We therefore show that Periphilin is the predominant RNA-binding protein of the HUSH complex and this RNA-binding is essential for HUSH activity. Overall design: To characterize the endogenous targets of PPHLN1 binding we initially used UV-crosslinked RNA immunoprecipitation sequencing (UV-RIPseq) in 293T PPHLN1 and SETDB1 knockout cells, expressing a panel of HA tagged PPHLN1 constructsxref:PubMed:39658355ENAfalseRNA binding by Periphilin plays an essential role in initiating silencing by the HUSH complexRNA binding by Periphilin plays an essential role in initiating silencing by the HUSH complex2025-09-242024-11-13PRJNA1136420GSE272315960639658355