講演情報
[BP-3]深部イントロンPOU1F1バリアントの多階層機能解析系の構築： c.143-83A＞Gは分枝部位破壊によるスプライス異常を起こす
○秋葉 和壽1,2, 長谷川 行洋2, 福井 由宇子1, 寺尾 美穂3, 高田 修治3, 深見 真紀1, 鳴海 覚志1 (1.国立成育医療研究センター研究所 分子内分泌研究部, 2.東京都立小児総合医療センター 内分泌・代謝科, 3.国立成育医療研究センター研究所 システム発生・再生医学研究部)
※12月15日（木）と16日（金）で同一演題の発表を行います。
Introduction: POU Class 1 Homeobox1 (POU1F1/Pou1f1) is a pituitary-specific transcription factor, and causes, when mutated, combined pituitary hormone deficiency. POU1F1/Pou1f1 has two isoforms, α and β isoforms. Recently, in human, pathogenic variants around the exon-intron boundary for the unique region of β isoform (β domain) were reported, although their functional consequences remain obscure.Materials and Methods: (1) We generated mice with the Pou1f1 c.143-83A＞G substitution that recapitulates the human intronic variant. (2) We created a vector for POU1F1 including all exonic and intronic sequences (16-kb) and artificial variants in or near β domain, which were candidate positions of the branch site of splicing. These vectors were transfected into HEK293 cells and transcripts were evaluated.Results: (1) Homozygous mice showed growth failure with body weight 35% of wildtype (WT) at 12 weeks, which was accompanied by anterior pituitary hypoplasia. The RNA-seq analysis of the pituitary gland showed the reduction of somatotrophs. The proportion of isoforms of Pou1f1 mRNA in the pituitary was evaluated, showing α isoform was the main isoform (99.0%) in WT, while homozygous mice produced exon 2 skip and β isoforms (78.5% and 17.1%, respectively). These results implied that c.143-83A＞G breaks the branch site of splicing. (2) In vitro assay showed that only c.143-83A＞G produced transcripts comparable to the mice model.Discussion: Our report is the first to show that the c.143-83A＞G variant leads splicing error and causes morphological and functional abnormalities.