Odaira Koya, Kawashima Fumika, Tamura Shogo, Suzuki Nobuaki, Tokoro Mahiru, Hayakawa Yuri, Suzuki Atsuo, Kanematsu Takeshi, Okamoto Shuichi, Takagi Akira, Katsumi Akira, Matsushita Tadashi, Shima Midori, Nogami Keiji, Kojima Tetsuhito, Hayakawa Fumihiko
Division of Cellular and Genetic Sciences, Department of Integrated Health Sciences, Nagoya University Graduate School of medicine, Nagoya, Japan.
Division of Cellular and Genetic Sciences, Department of Integrated Health Sciences, Nagoya University Graduate School of medicine, Nagoya, Japan.
Thromb Res. 2022 May;213:91-96. doi: 10.1016/j.thromres.2022.03.010. Epub 2022 Mar 15.
Hemophilia B (HB) is a hereditary bleeding disorder caused by the genetic variation of the coagulation factor IX (FIX) gene (F9). Several F9 structural abnormalities, including large deletion and/or insertion, have been observed to cause HB development. However, there is limited information available on F9 deep intronic variations. In this study, we report about a novel large deletion/insertion observed in a deep region of F9 intron 1 that causes mRNA splicing abnormalities.
The patient was a Japanese male diagnosed with moderate HB (FIX:C = 3.0 IU/dL). The genomic DNA of the patient was isolated from peripheral blood leukocytes. DNA sequences of F9 exons and splice donor/acceptor sites were analyzed via polymerase chain reaction and Sanger sequencing. Variant-affected F9 mRNA aberration and FIX protein production, secretion, and coagulant activity were analyzed by cell-based exon trap and splicing-competent FIX expression vector systems.
A 28-bp deletion/476-bp insertion was identified in the F9 intron 1 of a patient with moderate HB. A DNA sequence identical to a part of the inverted HNRNPA1 exon 12 was inserted. Cell-based transcript analysis revealed that this large intronic deletion/insertion disrupted F9 mRNA splicing pattern, resulting in reduction of protein-coding F9 mRNA.
A novel deep intronic F9 rearrangement was identified in a Japanese patient with moderate HB. Abnormal F9 mRNA splicing pattern due to this deep intronic structural variation resulted in a reduction of protein-coding F9 mRNA, which probably caused the moderate HB phenotype.
乙型血友病（HB）是一种由凝血因子IX（FIX）基因（F9）的基因变异引起的遗传性出血性疾病。已观察到几种F9结构异常，包括大片段缺失和/或插入，可导致HB的发生。然而，关于F9内含子深部变异的信息有限。在本研究中，我们报告了在F9内含子1深部区域观察到的一种新型大片段缺失/插入，其导致mRNA剪接异常。
该患者为一名日本男性，被诊断为中度HB（FIX：C = 3.0 IU/dL）。从外周血白细胞中分离出患者的基因组DNA。通过聚合酶链反应和桑格测序分析F9外显子以及剪接供体/受体位点的DNA序列。通过基于细胞的外显子捕获和具有剪接能力的FIX表达载体系统，分析受变异影响的F9 mRNA异常以及FIX蛋白的产生、分泌和凝血活性。
在一名中度HB患者的F9内含子1中鉴定出一个28bp的缺失/476bp的插入。插入了一段与反向的HNRNPA1外显子12的一部分相同的DNA序列。基于细胞的转录分析表明，这种大片段内含子缺失/插入破坏了F9 mRNA的剪接模式，导致编码蛋白的F9 mRNA减少。
在一名中度HB的日本患者中鉴定出一种新型的F9内含子深部重排。这种内含子深部结构变异导致的F9 mRNA剪接模式异常，导致编码蛋白的F9 mRNA减少，这可能是导致中度HB表型的原因。
