PARTICLE - a triplex-forming long ncRNA regulates locus specific methylation in response to low dose irradiation

Exposure to low dose irradiation causes transiently elevated expression of a long ncRNA PARTICLE (Gene PARTICL- 'Promoter of MAT2A-Antisense RadiaTion Induced Circulating LncRNA). PARTICLE affords both a cytosolic scaffold for the tumor suppressor methionine adenosyltransferase (MAT2A) and a nuclear genetic platform for transcriptional repression. In situ hybridisation discloses that PARTICLE-MAT2A associate together following irradiation. Bromouridine tracing and presence in exosomes indicate intercellular transport, and this is supported by ex-vivo data from radiotherapy-treated patients. Surface plasmon resonance indicates that PARTICLE forms a DNA-lncRNA triplex upstream of a MAT2A promoter CpG island. We show that PARTICLE represses MAT2A via methylation and demonstrate that the radiation-induced PARTICLE lncRNA interacts with the transcription repressive complex proteins G9a and SUZ12 (subunit of PRC2). The interplay of PARTICLE with MAT2A, implicates this lncRNA in intercellular communication as well as a recruitment platform for gene silencing machineries through triplex formation in response to irradiation. Total RNA was isolated from T47D (ductal breast epithelial tumor), HUVEC (umbilical vein endothelial) and U2OS (osteosarcoma) human cell lines (using Ambion RNA Life Technologies, cat. # AM1560) 4 hr after 2.5 Gy γ-irradiation. RNA was amplified, transcribed into Cy3 labelled cDNA and hybridized onto the human lncRNA array V2 (representing lncRNA probes (n) = 33,045, ArrayStar).