Investigation into the role of autophagy protease ATG4A in normal and impaired erythropoiesis

项目总结/摘要 自噬是一种在红细胞生成中具有组织特异性功能的自降解途径。红系期间 在分化过程中，自噬促进了大分子的降解和细胞的程序性清除。 线粒体这项提案的长期目标是确定自噬如何有助于正常和 红细胞生成受损骨髓增生异常综合征（MDS）患者通常有红细胞生成受损， 和线粒体功能障碍。MDS的一个子集，具有环形铁粒幼细胞（RS）的MDS，含有异常的铁粒幼细胞。 在成红细胞中，铁负载的线粒体的积累表明自噬可能在成红细胞中失调。 造血障碍和红细胞分化受损。虽然已知必需的自噬基因 虽然自噬在红细胞生成中起作用，但对自噬在红细胞生成过程中的组织特异性调节知之甚少。 人红系分化我们已经鉴定了自噬蛋白酶ATG 4A的选择性表达， 在红细胞谱系中。我们已经评估了ATG 4A在人红系细胞的离体模型中的贡献， 分化在分化早期，ATG 4A促进红系细胞的扩增和集落形成 祖先在分化后期，ATG 4A促进终末成熟、去核和线粒体膜电位。 间隙由于ATG 4蛋白酶受活性氧调节，因此ATG 4A在细胞内的功能可能与ATG 4蛋白酶的活性有关。 红细胞生成可能对氧化应激敏感。根据我们的初步结果，我们假设， ATG 4A在早期和晚期红细胞生成期间介导自噬，并且通过氧化应激失调。在aim中 1，我们将确定ATG 4A依赖性自噬在早期红细胞生成中的作用。在目标2中，我们将确定 ATG 4A依赖的线粒体清除机制。在目标3中，我们将研究 MDS-RS人类模型中的线粒体清除率。本提案中概述的实验将 在开发和使用人类iPSC的专家Sergei Doulatov博士的指导下进行 研究红细胞疾病的模型，以及Janis Abkowitz博士的共同指导， 华盛顿大学血液学博士，MDS和红细胞生物学国际专家。这个职业 发展奖将支持我的培训，成为一个独立的调查研究的贡献， 正常和功能失调的红细胞生成。

PROJECT SUMMARY/ ABSTRACT Autophagy is a self-degradation pathway that has tissue-specific functions in erythropoiesis. During erythroid differentiation, autophagy facilitates the degradation of macromolecules and the programmed clearance of mitochondria. The long-term objective of this proposal is to determine how autophagy contributes to normal and impaired erythropoiesis. Patients with Myelodysplastic syndromes (MDS) often have impaired erythropoiesis and mitochondrial dysfunction. A subset of MDS, MDS with ring sideroblasts (RS), contain the aberrant accumulation of iron-laden mitochondria in erythroblasts suggesting that autophagy may be dysregulated in hematopoietic disorders and impair erythroid differentiation. While essential autophagy genes are known to contribute to autophagy in erythropoiesis, little is known about the tissue-specific regulation of autophagy during human erythroid differentiation. We have identified the selective expression of the autophagy protease ATG4A in the erythroid lineage. We have evaluated the contribution of ATG4A in an ex vivo model of human erythroid differentiation. Early in differentiation, ATG4A promotes the expansion and colony formation of erythroid progenitors. Late in differentiation, ATG4A promotes terminal maturation, enucleation, and mitochondrial clearance. Since ATG4 proteases are regulated by reactive oxygen species, the function of ATG4A in erythropoiesis may be sensitive to oxidative stress. Based on our preliminary results, we hypothesize that ATG4A mediates autophagy during early and late erythropoiesis and is dysregulated by oxidative stress. In aim 1, we will identify the role of ATG4A-dependent autophagy in early erythropoiesis. In aim 2, we will determine the mechanism of ATG4A-dependent mitochondrial clearance. In aim 3, we will examine the regulation of mitochondrial clearance in human models of MDS-RS. The experiments outlined in this proposal will be conducted under the mentorship of Dr. Sergei Doulatov, an expert in the development and use of human iPSC models to study erythroid disorders, and co-mentorship of Dr. Janis Abkowitz, Chair of the Division of Hematology at the University of Washington and international expert in MDS and erythroid biology. This career development award will support my training to become an independent investigator studying the contribution of autophagy to normal and dysfunctional erythropoiesis.