Rubinstein-Taybi syndrome (RSTS) is a multisystem developmental disorder characterized by facial dysmorphisms, broad thumbs and halluces, growth retardation, and intellectual disability. In about 8% of RSTS cases, mutations are found in EP300. Previously, the EP300 mutation has been shown to cause the highly variable RSTS phenotype. Using exome sequencing, we identified a de novo EP300 frameshift mutation in a proband with coloboma, facial asymmetry and imperforate anus with minimal RSTS features. Previous molecular studies have demonstrated the importance of EP300 in oculogenesis, supporting the possibility that EP300 mutation may cause ocular coloboma. Since a wide phenotypic spectrum is well known in EP300-associated RSTS cases, the atypical phenotype identified in our proband may be an example of rare manifestations of RSTS.

Sister chromatid cohesion is mediated by cohesin and is essential for accurate chromosome segregation. The cohesin subunits SMC1, SMC3, and Rad21 form a tripartite ring within which sister chromatids are thought to be entrapped. This event requires the acetylation of SMC3 and the association of sororin with cohesin by the acetyltransferases Esco1 and Esco2 in humans, but the functional mechanisms of these acetyltransferases remain elusive. Here, we showed that Esco1 requires Pds5, a cohesin regulatory subunit bound to Rad21, to form cohesion via SMC3 acetylation and the stabilization of the chromatin association of sororin, whereas Esco2 function was not affected by Pds5 depletion. Consistent with the functional link between Esco1 and Pds5, Pds5 interacted exclusively with Esco1, and this interaction was dependent on a unique and conserved Esco1 domain. Crucially, this interaction was essential for SMC3 acetylation and sister chromatid cohesion. Esco1 localized to cohesin localization sites on chromosomes throughout interphase in a manner that required the Esco1-Pds5 interaction, and it could acetylate SMC3 before and after DNA replication. These results indicate that Esco1 acetylates SMC3 via a mechanism different from that of Esco2. We propose that, by interacting with a unique domain of Esco1, Pds5 recruits Esco1 to chromatin-bound cohesin complexes to form cohesion. Furthermore, Esco1 acetylates SMC3 independently of DNA replication.

The cyclin B-dependent protein kinase Cdk1 is a master regulator of mitosis and phosphorylates numerous proteins on the minimal consensus motif Ser/Thr-Pro (S/T-P). At least in several proteins, however, not well-defined motifs lacking a Pro in the + 1 position, referred herein to as non-S/T-P motifs, have been shown to be phosphorylated by Cdk1. Here we show that non-S/T-P motifs in fact form consensus sequences for Cdk1 and probably play roles in mitotic regulation of physiologically important proteins. First, we show, by in vitro kinase assays, that previously identified non-S/T-P motifs all harbour one or more C-terminal Arg/Lys residues essential for their phosphorylation by Cdk1. Second, using Arg/Lys-scanning oriented peptide libraries, we demonstrate that Cdk1 phosphorylates a minimal sequence S/T-X-X-R/K and more favorable sequences (P)-X-S/T-X-[ R/K](2-5) as its non-S/T-P consensus motifs. Third, on the basis of these results, we find that highly conserved linkers (typically, T-G-E-K-P) of C2H2 zinc finger proteins and a nuclear localization signal-containing sequence (matching P-X-S-X-[R/K](5)) of the cytokinesis regulator Ect2 are inhibitorily phosphorylated by Cdk1, well accounting for the known mitotic regulation and function of the respective proteins. We suggest that non-S/T-P Cdk1 consensus motifs identified here may function to regulate many other proteins during mitosis.

本研究では、高等学校の『生物』の教科書で扱われているPCR法（ポリメラーゼ連鎖反応法）に関する新たな教材開発を行った。方法としては、高校生にとって身近なペット（犬）に着目し、そのSNP（一塩基多型）の有無の検出を行うことで遺伝疾患の罹患率を調べるものである。その結果、犬の高尿酸尿症と進行性網膜萎縮症の発症に係る遺伝子の変異を検出可能な実験系の構築を行うことが可能となった。本研究を継続した結果、現在も指導した高校生が課題研究を進めており、一定の教育的効果が見受けられた。今後も更なる教材化を図りたいと考えている。

高等学校の『生物』バイオテクノロジー分野で扱うPCR法（ポリメラーゼ連鎖反応法）の新たな教材開発を行った。方法としては、高校生にとって身近なペット（犬）に着目し、そのSNP（一塩基多型）の有無の検出を行った。その結果、犬の進行性網膜萎縮症の発症に係る遺伝子の変異をPCR法のみで検出可能な実験系の構築を行うことが可能となった。昨年度より、本研究を継続した結果、指導した高校生が課題研究を進めている。今後も更なる教材化を図りたいと考えている。

本研究では、高等学校の『生物』で扱うPCR法（ポリメラーゼ連鎖反応法）を利用して、高校生が遺伝子変異の有無を自らの手で検証可能な教材の開発を行った。検出用サンプルに関して、高校生にとって比較的身近なペット（犬）に着目し、教材化を図った。その結果、犬の高尿酸血症の発症に係る遺伝子の変異をPCR法のみで検出可能な実験系の構築を行うことが可能となった。また、現在、本研究を利用して、高校生が課題研究を進められるようになっている。今後も更なる教材化を図り、高校生を対象としたバイオテクノロジーの普及に努めたいと考えている。