Dysregulation of DNA methylation and mRNA alternative cleavage and polyadenylation (APA) are both prevalent in cancer and have been studied as independent processes. We discovered a DNA methylation-regulated APA mechanism when we compared genome-wide DNA methylation and polyadenylation site usage between DNA methylation-competent and DNA methylation-deficient cells. Here, we show that removal of DNA methylation enables CTCF binding and recruitment of the cohesin complex, which, in turn, form chromatin loops that promote proximal polyadenylation site usage. In this DNA demethylated context, either deletion of the CTCF binding site or depletion of RAD21 cohesin complex protein can recover distal polyadenylation site usage. Using data from The Cancer Genome Atlas, we authenticated the relationship between DNA methylation and mRNA polyadenylation isoform expression in vivo. This DNA methylation-regulated APA mechanism demonstrates how aberrant DNA methylation impacts transcriptome diversity and highlights the potential sequelae of global DNA methylation inhibition as a cancer treatment.

OCRL1 and its paralog INPP5B encode phosphatidylinositol 5-phosphatases that localize to the primary cilium and have roles in ciliogenesis. Mutations in OCRL1 cause the X-linked Dent disease type 2 (DD2; OMIM# 300555), characterized by low-molecular weight proteinuria, hypercalciuria, and the variable presence of cataracts, glaucoma and intellectual disability without structural brain anomalies. Disease-causing mutations in INPP5B have not been described in humans. Here, we report the case of an 11-year-old boy with short stature and an above-average IQ; severe proteinuria, hypercalciuria and osteopenia resulting in a vertebral compression fracture; and Chiari I malformation with cervico-thoracic syringohydromyelia requiring suboccipital decompression. Sequencing revealed a novel, de novo DD2-causing 462 bp deletion disrupting exon 3 of OCRL1 and a maternally inherited, extremely rare (ExAC allele frequency 8.4×10−6) damaging missense mutation in INPP5B (p.A51V). This mutation substitutes an evolutionarily conserved amino acid in the protein’s critical PH domain. In silico analyses of mutation impact predicted by SIFT, PolyPhen2, MetaSVM and CADD algorithms were all highly deleterious. Together, our findings report a novel association of DD2 with Chiari I malformation and syringohydromyelia, and document the effects of digenic mutation of human OCRL paralogs. These findings lend genetic support to the hypothesis that impaired ciliogenesis may contribute to the development of Chiari I malformation, and implicates OCRL-dependent PIP3 metabolism in this mechanism.

We report that when expressed at similar levels from an isogenic locus, the Airn lncRNA induces Polycomb deposition with a potency that rivals Xist. However, when subject to the same degree of promoter activation, Xist is more abundant and more potent than Airn. Our data definitively demonstrate that the Airn lncRNA is functional and suggest that Xist achieved extreme potency in part by evolving mechanisms to promote its own abundance.

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Dysregulation of DNA methylation and mRNA alternative cleavage and polyadenylation (APA) are both prevalent in cancer, but they have been studied as independent processes. We discovered a DNA methylation regulated APA mechanism when we compared genome-wide DNA methylation and polyadenylation site usage between DNA methylation-competent and DNA methylation-deficient cells. Here we show that removal of DNA methylation enables CTCF binding and recruitment of the cohesin complex, which in turn, promotes proximal polyadenylation site usage. In this DNA de-methylated context, depletion of the RAD21 cohesin complex component can recover distal polyadenylation site usage. We also confirmed, using chromosomal conformation capture (3C), that CTCF and the cohesin complex mediate chromatin loops in the absence of DNA methylation to modulate polyadenylation site usage. Leveraging data from The Cancer Genome Atlas, we authenticated the relationship between DNA methylation and mRNA polyadenylation isoform expression in vivo. This DNA methylation regulated APA mechanism demonstrates how aberrant DNA methylation impacts transcriptome diversity and highlights the potential sequelae of global DNA methylation inhibition as a cancer treatment. Citation Format: Angela H. Ting, Vishal Nanavaty, Elizabeth Abrash, Byron Lee, Emily Fink, Tae Hyun Hwang, Changjin Hong. DNA methylation at the intersect of chromatin structure and transcriptome diversity [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 1081.