A Study of Allostery Within Liganded RARα:RXR Heterodimers at the Genome Scale [RNA-seq]

We provide evidence of coordination between heterodimeric nuclear hormone receptor ligands at the genome scale to reveal three unique transcriptional outcomes. First, when stimulated in pancreatic acinar cells, distinct doubly-liganded complexes of the retinoic acid receptor-α (RARα) and retinoid-X-receptor (RXR) bind divergent DNA hormone response elements (HRE). Furthermore, the link between ligand pairs and HRE specificity is correlated with RNA transcript formation. Second, RXR ligands can function as either silent partners or as cooperative effectors on distinct HREs, also with correlated transcriptional outcomes. Third, the absence of unique binding sites on DNA by unliganded RARα:RXR suggests that productive HRE recognition and binding is an exclusive feature of the agonist-bound heterodimer. To achieve the above, DNA-bound heterodimers of RARα:RXR were isolated separately with an RARα-specific antibody, RARα-specific and RXR-specific agonists and the pan-ligand, 9-cis retinoic acid, using the CUT&RUN technique. At each liganded state, transcriptional activity was detected by mRNA sequencing. RNA-seq Profiling in mouse acinar pancreatic tissue (266-6) with differing agonistic treatment of the retinoic acid receptor alpha

本研究提供了全基因组范围内异二聚体核激素受体配体间协同作用的证据，揭示了三种独特的转录调控结局。其一，在胰腺腺泡细胞中受到刺激时，视黄酸受体α（retinoic acid receptor-α, RARα）与类维生素A X受体（retinoid-X-receptor, RXR）形成的不同双配体复合物，可结合序列相异的DNA激素反应元件（DNA hormone response elements, HRE）。此外，配体对与HRE特异性之间的关联，与RNA转录本的生成呈显著相关。其二，RXR配体可在不同HRE上分别作为沉默伴侣或协同效应因子发挥功能，同样伴随对应的转录调控结局。其三，未结合配体的RARα:RXR异二聚体在DNA上不存在特异性结合位点，这表明具有功能活性的HRE识别与结合，仅为激动剂结合的异二聚体所独有。为达成上述研究目标，本研究采用CUT&RUN技术，分别使用RARα特异性抗体、RARα特异性激动剂与RXR特异性激动剂，以及全配体9-顺式视黄酸，分离得到结合于DNA的RARα:RXR异二聚体。在每种配体结合状态下，通过mRNA测序检测转录活性。本研究对经不同视黄酸受体α激动剂处理的小鼠胰腺腺泡组织（266-6）开展了RNA测序分析。