lncRNA-HBBP1通过调控珠蛋白加重β地中海贫血的分子机制

The clinical manifestation and evaluation of efficacy on β thalassemia have been bewildered by the problem that phenotype of patients does not correspond to their genotype. We found that lncRNA-HBBP1 (in brief as HBBP1) increased significantly by comparing the expression level of lncRNA in the peripheral blood of major and minor thalassemia patients and normal controls. When the expression of HBBP1 was up-regulated in zebrafish, the amount of red blood cells and hemoglobin level were decreased significantly. It was found that HBBP1 could bind to β globin and some proteins of ubiquitin system in K562 erythroleukemic cells tested with CHIRP technology. The results of RIP experiments also confirmed the binding of HBBP1 to β globin. We propose the hypothesis that by combining with β globin and some proteins of ubiquitin system, HBBP1 could aggravate the imbalance of α and β globin as well as the anemic phenotype. This study includes：(1) To collect more samples and verify the degree of correlation between HBBP1 level and anemia in patients with β thalassemia; (2) To construct HBBP1 transgenic mice and HBBP1 overexpressed erythroleukemic cells, and identify the effects of HBBP1 on red blood cells, hemoglobin and globin levels; (3) To explore the molecular mechanism of HBBP1 effecting on β globin degradation. The conclusions of this project will help to understand new molecular mechanism of β thalassemia and provide new strategy of diagnosis and therapy.

β地中海贫血基因型与表型不对应的问题影响对患者病情判断和疗效评估。我们前期研究发现β地中海贫血患者外周血lncRNA-HBBP1(简称HBBP1)较对照组显著升高。在斑马鱼中上调HBBP1导致红细胞数量和血红蛋白水平均显著下降。应用CHIRP技术在K562红白血病细胞中发现HBBP1可以与β珠蛋白和泛素系统的某些蛋白结合，RIP实验结果也验证了HBBP1与β珠蛋白的结合。据此我们提出假说：HBBP1通过结合β珠蛋白及泛素系统相关蛋白，促进β珠蛋白的降解，加重α和β珠蛋白的不平衡，使得贫血表型加重。本研究拟进行：1.扩大样本验证HBBP1水平与β地中海贫血患者贫血程度的相关性；2.构建转基因小鼠及过表达的红白血病细胞株，明确HBBP1对红细胞数量、血红蛋白及珠蛋白水平的影响；3.从泛素化角度探索HBBP1促进β珠蛋白降解的分子机制。项目的完成将揭示β地中海贫血新的分子机制，提供新的诊治思路。

β-地中海贫血是世界范围内最常见的单基因疾病之一。其基因型与表型不对应的问题影响对患者病情的判断和疗效评估。调节珠蛋白含量一直是探索β-地中海贫血治疗的切入点。血红蛋白亚单位β-假基因1（lncRNA-HBBP1）被认为通过升高胎儿血红蛋白（HbF）水平而导致β-地中海贫血的较轻症状。但在我们的研究中发现，与正常对照组相比重型β-地中海贫血（Tm）患者的lncRNA-HBBP1水平显著升高。采用模式动物观察到，lncRNA-HBBP1过度表达导致斑马鱼红细胞数量和血红蛋白含量减少，lncRNA-HBBP1转基因小鼠出现类似的贫血表型。在体外，lncRNA-HBBP1在K562细胞中与β-珠蛋白和α-珠蛋白结合。其高表达招募了更多的泛素结合酶2K（UBE2K）和泛素C（UBC）分别与β-珠蛋白和α-珠蛋白相互作用，然后通过泛素-蛋白酶体途径（UPP）不同程度地促进它们的降解。因此，β-/α-珠蛋白比值失衡加剧，可能加重贫血。此外，缺氧诱导因子1亚单位α（HIF-1α）的激活通过结合其调节序列促进lncRNA-HBBP1转录。总之，通过上述实验所见，我们首次提出了lncRNA-HBBP1对β-地中海贫血调控的一种新功能，它可以通过不同程度地降解β-珠蛋白和α-珠蛋白而加重β-地中海贫血程度，这与以往的研究所有不同。因此，lncRNA-HBBP1可能是参与治疗β-地中海贫血的潜在分子靶点，以及评估β-地中海贫血预后的分子标志物。