A role for repressive complexes and H3K9 di-methylation in PRDM5-associated brittle cornea syndrome

Abstract

Type 2 brittle cornea syndrome (BCS2) is an inherited connective tissue disease with a devastating ocular phenotype caused by mutations in the transcription factor PR domain containing 5 (PRDM5) hypothesized to exert epigenetic effects through histone and DNA methylation. Here we investigate clinical samples, including skin fibroblasts and retinal tissue from BCS2 patients, to elucidate the epigenetic role of PRDM5 and mechanisms of its dysregulation in disease. First we report abnormal retinal vascular morphology in the eyes of two cousins with BCS2 (PRDM5 Δ exons 9-14) using immunohistochemistry, and mine data from skin fibroblast expression microarrays from patients with PRDM5 mutations p.Arg590* and Δ exons 9-14, as well as from a PRDM5 ChIP-sequencing experiment. Gene ontology analysis of dysregulated PRDM5-target genes reveals enrichment for extracellular matrix (ECM) genes supporting vascular integrity and development. Q-PCR and ChIP-qPCR confirm upregulation of critical mediators of ECM stability in vascular structures (COL13A1, COL15A1, NTN1, CDH5) in patient fibroblasts. We identify H3K9 di-methylation (H3K9me2) at these PRDM5-target genes in fibroblasts, and demonstrate that the BCS2 mutation p.Arg83Cys diminishes interaction of PRDM5 with repressive complexes, including NuRD complex protein CHD4, and the repressive chromatin interactor HP1BP3, by co-immunoprecipitation combined with mass spectrometry. We observe reduced heterochromatin protein 1 binding protein 3 (HP1BP3) staining in the retinas of two cousins lacking exons 9-14 by immunohistochemistry, and dysregulated H3K9me2 in skin fibroblasts of three patients (p.Arg590*, p.Glu134* and Δ exons 9-14) by western blotting. These findings suggest that defective interaction of PRDM5 with repressive complexes, and dysregulation of H3K9me2, play a role in PRDM5-associated disease.

The Ehlers–Danlos syndromes, rare types

“…To provide a comprehensive overview of the clinical phenotype of BCS, we reviewed the data on 51 patients ( ZNF469 : n = 32; PRDM5: n = 19) (Table S1) [Al‐Hussain et al, ; Abu et al, ; Christensen et al, ; Khan et al, ; Burkitt Wright et al, ; Al‐Owain et al, ; Aldahmesh et al, ; Rohrbach et al, ; Ramappa et al, ; Avgitidou et al, ; Porter et al, ]. Only those patients with individual clinical and molecular data were included in the review.…”

Systematic review of differential methylation in rare ophthalmic diseases

“…Following a comprehensive systematic review of the literature, this study found that the role of methylation in ophthalmic disease has been evaluated in histone methylation marks,41–43 gene body and promoter regions,44–53 and miRNAs,54 in the following rare ophthalmic conditions:…”

“…Brittle cornea syndrome, an autosomal recessive inherited disorder characterised by corneal fragility 42…”

“…Aberrant methylation was established as a pathogenic biomarker of ophthalmic disease in eight studies. This included: the identification of methylation as playing a role in the high mutability of exon 21 within KIF21A in CFEOM1,46 loss of H3K9me2-mediated repression in brittle cornea syndrome,42 differential methylation of autoinflammatory genes in retinopathy of prematurity (including NGPT1 , BDNF , CRP , MPO , SAA1 , SAA2 , TNFRSF1A and TNFRSF1B ),47 differential methylation of several genes corresponding to RNA sequencing gene expression analysis in keratoconus,50 hypomethylation in genes related to ion channel roles which are essential for corneal endothelium function in patients with FECD,44 differential miRNA expression in FECD cases compared with controls including the hypermethylation of miR-199b-5p which negatively regulates the corneal transcription factors Snail and ZEB1 ,54 altered histone methylation status associated with changes in TGFBIp expression levels in GCD243 and, finally, skewed inactivation of the X chromosome associated with a non-random DNA methylation pattern in choroideraemia 48…”

Zinc fingers 1, 2, 5 and 6 of transcriptional regulator, PRDM4, are required for its nuclear localisation

“…PRDM3 and 16 maintain mammalian heterochromatin integrity [14], while PRDM9 controls meiotic recombination [15][16][17][18][19][20]. Deregulated expression of many PRDM proteins is implicated in different types of cancers [1] and mutations in some are linked to congenital disorders [21][22][23] highlighting the importance of these proteins in normal development and disease.…”