Functional characterization of the long antisense noncoding RNA CDKN2BAS (ANRIL) and elucidation of the specific role in the pathophysiology of periodontitis

对牙周炎潜在关键疾病基因的遗传探索使我们鉴定出第一个牙周炎和冠心病（CHD）的共同遗传风险基因，称为ANRIL （CDKN2BAS）。ANRIL的生物学功能目前在很大程度上是未知的。在之前的工作中，我们发现了在致病菌刺激下牙龈成纤维细胞中选择性剪接ANRIL转录物的不同调控与特定遗传背景的关系。这些转录本可能具有不同的生理和组织特异性功能，剪接模式的变化可能具有病理相关的影响。我们将解决以下问题：(1)在牙龈成纤维细胞（hGF）和动脉内皮细胞中不同调节的ANRIL剪接变异体的详细阐明。(2) FISH鉴定了两个主要ANRIL转录本的核定位。(3)鉴定主要组织特异性ANRIL转录本的推定蛋白结合伙伴。(4)通过对疾病相关纯合背景的牙龈成纤维细胞及常见等位基因诱导ANRIL过表达和低表达的全基因组表达谱，阐明ANRIL在全基因组水平上的转录调控作用。(5)对两个主要相关单倍型块纯合的ANRIL患者的基因组区域进行靶向重测序，鉴定功能性遗传变异。我们随后将测试这些变异在冠心病中的病因相关性。

Genetic exploration of potential key disease genes of periodontitis allowed us the identification of the first shared genetic risk gene of periodontitis and coronary heart disease (CHD), termed ANRIL (CDKN2BAS). The biological function of ANRIL is currently largely unknown. In previous works we identified different regulation of alternatively spliced ANRIL transcripts in gingival fibroblasts upon stimulation with pathogenic bacteria in relation to the specific genetic background. These transcripts may have distinct physiological and tissue specific functions, and changes in the splicing patterns potentially have pathological relevant effects. We will address the following questions: (1) Minute elucidation of differently regulated splicing variants of ANRIL within gingival fibroblasts (hGF) and arterial endothelial cells. (2) Identification of the nuclear localisation of the two main ANRIL transcripts by FISH. (3) Identification of putative protein binding partners of the major tissue specific ANRIL transcripts. (4) Genome-wide expression profiling upon inducible over-expression and knockdown of ANRIL in gingival fibroblastic cells of homozygous background for the disease associated as well as the common alleles to elucidate the transcriptional regulatory effects of ANRIL on a genome-wide level. (5) Targeted re-sequencing of the genomic region of ANRIL in patients who are homozygous for the two main associated haplotype blocks to identify the functional genetic variants. We will subsequently test these variants for their etiological relevance in CHD.