Zhang Yang, David Natalie L, Pesaresi Tristan, Andrews Rosemary E, Kumar G V Naveen, Chen Hongyin, Qiao Wanning, Yang Jinzhao, Patel Kareena, Amorim Tania, Sharma Ankit X, Liu Silvia, Steinhauser Matthew L
School of Public Health (Shenzhen), Sun Yat-sen University, Shenzhen, Guangdong, China.
Aging Institute, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania, USA.
JCI Insight. 2024 Dec 10;10(2):e184140. doi: 10.1172/jci.insight.184140.
Mechanisms underpinning signals from genome-wide association studies remain poorly understood, particularly for noncoding variation and for complex diseases such as type 2 diabetes mellitus (T2D) where pathogenic mechanisms in multiple different tissues may be disease driving. One approach is to study relevant endophenotypes, a strategy we applied to the UBE2E2 locus where noncoding single nucleotide variants (SNVs) are associated with both T2D and visceral adiposity (a pathologic endophenotype). We integrated CRISPR targeting of SNV-containing regions and unbiased CRISPR interference (CRISPRi) screening to establish candidate cis-regulatory regions, complemented by genetic loss of function in murine diet-induced obesity or ex vivo adipogenesis assays. Nomination of a single causal gene was complicated, however, because targeting of multiple genes near UBE2E2 attenuated adipogenesis in vitro; CRISPR excision of SNV-containing noncoding regions and a CRISPRi regulatory screen across the locus suggested concomitant regulation of UBE2E2, the more distant UBE2E1, and other neighborhood genes; and compound heterozygous loss of function of both Ube2e2 and Ube2e1 better replicated pathological adiposity and metabolic phenotypes compared with homozygous loss of either gene in isolation. This study advances a model whereby regulatory effects of noncoding variation not only extend beyond the nearest gene but may also drive complex diseases through polygenic regulatory effects.
全基因组关联研究信号背后的机制仍知之甚少，尤其是对于非编码变异以及2型糖尿病（T2D）等复杂疾病而言，在这些疾病中，多种不同组织中的致病机制可能是疾病的驱动因素。一种方法是研究相关的内表型，我们将此策略应用于UBE2E2基因座，其中非编码单核苷酸变异（SNV）与T2D和内脏肥胖（一种病理性内表型）均相关。我们整合了对含SNV区域的CRISPR靶向和无偏倚的CRISPR干扰（CRISPRi）筛选，以建立候选顺式调控区域，并辅以小鼠饮食诱导肥胖或体外脂肪生成试验中的基因功能丧失研究。然而，确定单个因果基因很复杂，因为靶向UBE2E2附近的多个基因会减弱体外脂肪生成；对含SNV的非编码区域进行CRISPR切除以及对该基因座进行CRISPRi调控筛选表明，UBE2E2、距离更远的UBE2E1以及其他邻近基因受到协同调控；与单独敲除任一基因的纯合子相比，Ube2e2和Ube2e1的复合杂合子功能丧失能更好地重现病理性肥胖和代谢表型。这项研究提出了一个模型，即非编码变异的调控作用不仅延伸到最近的基因之外，还可能通过多基因调控作用驱动复杂疾病。
