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

通过对牙周炎潜在致病基因的遗传学研究，我们发现了第一个与牙周炎和冠心病(CHD)相关的共同遗传风险基因，称为ANRIL(CDKN2BAS)。目前，ANRIL的生物学功能尚不清楚。在以前的工作中，我们发现在特定的遗传背景下，病原菌刺激牙龈成纤维细胞时，选择性剪接的ANRIL转录本的调节不同。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上的相关影响。我们将解决以下问题：(1)详细阐明在牙周成纤维细胞(HGF)和动脉内皮细胞中ANRIL的不同调控剪接变体。(2)FISH鉴定两种主要的ANRIL转录本的核定位。(3)主要组织特异性ANRIL转录本的蛋白结合伙伴的鉴定。(4)ANRIL在纯合子背景的牙周成纤维细胞中诱导过表达和下调的全基因组表达谱以及常见的等位基因，以在全基因组水平上阐明ANRIL的转录调控作用。(5)对两个主要相关单倍型区块纯合子的患者进行ANRIL基因组区域的定向重测序，以确定功能性遗传变异。我们随后将测试这些变异在冠心病病因学上的相关性。

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.