The Chromatin-Associated Phf12 Protein Maintains Nucleolar Integrity and Prevents Premature Cellular Senescence

Abstract

Pf1, also known as Phf12 (plant homeodomain [PHD] zinc finger protein 12), is a member of the PHD zinc finger family of proteins. Pf1 associates with a chromatin-interacting protein complex comprised of MRG15, Sin3B, and histone deacetylase 1 (HDAC1) that functions as a transcriptional modulator. The biological function of Pf1 remains largely elusive. We undertook the generation of Pf1 knockout mice to elucidate its physiological role. We demonstrate that Pf1 is required for mid-to late gestation viability. Pf1 inactivation impairs the proliferative potential of mouse embryonic fibroblasts (MEFs) and is associated with a significant decrease in bromodeoxyuridine incorporation; an increase in senescence-associated ␤-galactosidase (SA-␤-Gal) activity, a marker of cellular senescence; and elevated levels of phosphorylated H2AX (␥-H2A.X), a marker associated with DNA double-strand breaks. Analysis of transcripts differentially expressed in wild-type and Pf1-deficient cells revealed the impact of Pf1 in multiple regulatory arms of the ribosome biogenesis pathways. Strikingly, assessment of the morphology of the nucleoli exposed an abnormal nucleolar structure in Pf1-deficient cells. Finally, proteomic analysis of the Pf1-interacting complexes highlighted proteins involved in ribosome biogenesis. Taken together, our data reveal an unsuspected function for the Pf1-associated chromatin complex in the ribosomal biogenesis and senescence pathways.KEYWORDS transcription, ribosome, Pf1, senescence, nucleolus P f1, also known as Phf12 (plant homeodomain [PHD] zinc finger protein 12), is a member of the PHD zinc finger family of proteins. PHD domains are small, 50-to 80-amino-acid-long domains often found in clusters of two or three and/or in the proximity of other chromatin-interacting domains, such as bromo-or chromodomains. Consistently, many of the PHD-containing proteins are nuclear proteins that interact with chromatin. Increasing evidence suggests that PHD domains are capable of recognizing modified and unmodified histone tails and that PHD domain-containing proteins act as epigenetic readers (1).The Pf1 gene is conserved throughout evolution, and the Pf1 protein, like its Saccharomyces cerevisiae yeast homolog, Rco1, contains two PHD domains in its N terminus. Mammalian Pf1 was first identified in a yeast two-hybrid screen for proteins interacting with the paired amphipathic helix 2 (PAH2) domain of Sin3A and shown to function as a transcriptional repressor (2). A later report identified Pf1 to be one of the components of the human MRG15 complex, together with Sin3B but not together with its close homolog, Sin3A (3). The PHD domains of Pf1 are important for the interaction with the MRG domain of MRG15, which relies mainly on hydrophobic interactions (3-5). The interaction of Pf1 with a complex containing Sin3B but not Sin3A was also confirmed in experiments identifying associations between Sin3B, histone deacetylase

The HDAC-Associated Sin3B Protein Represses DREAM Complex Targets and Cooperates with APC/C to Promote Quiescence

“…Consistent with previous reports, we identified several members of the canonical Sin3-HDAC complex, which include the following: Sin3A, HDAC1, HDAC2, RBBP7, SAP130, ARID4B, ING1, ING2, SUDS3, SAP30, SAP30L, BRMS1L, and BRMS1 ( Bansal et al, 2016 ) ( Figure 1A ). Additionally, several members of a recently characterized smaller Sin3B complex were identified as well ( Graveline et al, 2017 ; Jelinic et al, 2011 ). These proteins include the following: PHF12, KDM5A, EMSY, and MORF4L1.…”

Family-based exome sequencing identifies rare coding variants in age-related macular degeneration

“…RXFP2 encodes a receptor for glycoprotein hormones that contains a unique low-density lipoprotein type A (LDLa) module ( 57 ); though this gene is not expressed in the adult human retina ( 54 ), its homolog in zebrafish is transcribed in the developing brain and retina ( 58 ). PHF12 encodes a member of the PHD zinc finger family of proteins involved in regulation of ribosomal biogenesis and senescence ( 59 ) and could contribute to AMD pathology through activation of senescence. Although present in only two families, the findings of rare variants in SPEF2 and BCAR1 are also noteworthy as both of these candidates were highlighted as most likely candidate at the AMD-GWAS loci based on a gene priority score ( 16 ).…”

Differential Complex Formation via Paralogs in the Human Sin3 Protein Interaction Network

“…SUDS3/BRMS1/BRMS1L, SAP30/SAP30L, and ING1/ING2 have homology to Rpd3L-specific components Sds3, Sap30, and Pho23, respectively ( 12 , 13 , 14 ). Components specifically found within Rpd3S, Rco1, and Eaf3, share homology with human PHF12 and MORF4L1, respectively ( 15 , 16 ). Although SIN3A can clearly interact with Rpd3L component homologs, data supports the existence of SIN3B complexes that contain Rpd3S component homologs ( 17 , 18 ).…”