Peking University, Nov. 16, 2012: A research group from PKU the College of Engineering has discovered that clathrin and phosphatidylinositol-4,5-bisphosphate tightly controls lysosome homeostasis. Consisting of Professor Xi Jianzhong and his doctoral student Zhang Hanshuo, together with co-workers from Tsinghua University, China Agriculture University and National Institute of Biological Science, the team published their research in Nature Cell Biology (http://www.nature.com/ncb/journal/v14/n9/full/ncb2557.html).

Autophagy is a lysosome-based degradation pathway. During autophagy, lysosomes fuse with autophagosomes to form autolysosomes. Following starvation-induced autophagy, nascent lysosomes are generated from autolysosomal membranes through an evolutionarily conserved cellular process, autophagic lysosome reformation (ALR). The research reveals that over tens of new components get involved and clathrin and phosphatidylinositol-4,5-bisphosphate (PtdIns(4,5)P2) are the central components in ALR.

To identify proteins required for ALR in a systematic manner, they took a combined approach that included proteomic analysis, large-scale RNA-mediated interference (RNAi) screening and functional studies. They carried out the loss-of-function screen using a recently developed method, SAMCell, which allows the delivery of a large number of individual RNAi molecules to cell islands grown on single glass slides with high efficiency.

The SAMCell method was developed by Xi and his student Zhang and applied to study cancer metastasis (Zhang, H. et al, Nature Communication 2011). Through silencing genes one by one, SAMcell can identify functional genes regulating diverse cell behaviors as well as phenotypes of cell organelles in a large scale. This functional study demonstrates the central role of clathrin and its associated proteins in cargo sorting, phospholipid conversion, initiation of autolysosome tubulation, and proto-lysosome budding during ALR.

Identification of genes regulating ALR by proteomic analysis and SAMCell-RNAi screening

The researchers believe that this work not only uncovers a molecular pathway by which lysosome homeostasis is maintained through the ALR process, but also reveals unexpected functions of clathrin and PtdIns(4,5)P2 in lysosome homeostasis. The findings are considered to have provided a new insight for scientists to deeply understand cellular and molecular regulation mechanism of autophagy.

"We are very happy to have this kind of collaboration with Dr. Yu Li and others. In fact, the RNAi based large scale screening is a very powerful tool for the study of functional genes. I believe that more and more laboratories would like take use of this tool in future.” Xi said.

This project is supported by 973 Program, the National Science Foundation of China, etc.

Edited by: Zhang Jiang
Source: College of Engineering