Def-Capn3蛋白降解新途径调控肝脏发育和再生的分子机制研究

Def是我们首先研究并命名的一种新型核仁蛋白。在斑马鱼中def基因失活导致肝脏变小。我们最近发现Def与半胱氨酸蛋白酶Calpain3形成复合体介导p53的降解,从而解释了为什么def突变体中p53蛋白水平增加，且富集于核仁中。def 突变体中p53的激活诱导其异构体Δ113p53转录表达。Δ113p53能促进p53抑制细胞周期的功能，但拮抗p53促细胞凋亡功能，这解释了为什么def突变体肝脏变小是由于细胞增殖受阻而不是由于细胞凋亡异常导致的。最近我们观察到def杂合突变体的肝脏切除后不能正常再生。但目前对Def-Capn3蛋白降解途径如何行使其功能、如何调节肝脏发育和再生仍是未知数。本项目旨在利用斑马鱼和小鼠为模式深入研究以Def-Capn3为轴心的蛋白降解途径的作用机理以及Def蛋白本身受调控的机理，以期揭示核仁中是否存在一个新的途径来调节细胞的增殖和死亡并进而调控肝脏的发育和再生。

Def (Digestive organ expansion factor) was first identified by our lab as a pan-endoderm enriched factor that is essential for the growth of liver and other digestive organs in zebrafish. Def belongs to a novel protein family evolutionally conserved from yeast to human and is a component of the ribosomal small subunit (SSU) complex in the nucleolus. In zebrafish, loss-of-function of def (e.g. the defhi429 null mutant) confers a small liver phenotype. We recently reported that loss-of-function of def led to up-regulation of the level of the p53 protein, a well-known tumor suppressor, which was notablely specifically accumulated in the nucleolus in the mutant cells. Consequently, the p53 pathway was activated to selectively upregulate the transcriptional expression of Δ113p53, which encodes an N-terminal truncated p53 isoform. The transcriptional expression of Δ113p53 is initiated by an alternative promoter contained in the intron 4 of the p53 gene which is totally p53-dependent. We also reported that both zebrafish Def and its human counterpart hu-Def (c1orf107, 1q32.2) teamed up with Calpain3, a cystein proteinase, to mediate the degradation of the p53 protein. In additon, our unpublished data showed that defhi429 heterozygous mutant was defective in liver regeneration. This research proposal plans a detailed research program on this novel finding with the purpose to elucidate the molecular mechanism of the Def-Calpain 3 protein degradation pathway and to define the biological role of this pathway in the processes of liver development and liver regeneration. Our findings will not only help us to understand more about the liver biology but to define a novel function for the nucleolus in the context of cell biology as well.