A research team led by Prof. Guo Yusong at The Hong Kong University of Science and Technology (HKUST) has made a remarkable advancement in understanding cellular transport mechanisms. Their study, published in the Proceedings of the National Academy of Sciences (PNAS), reveals critical insights into how cells manage protein transport, a process vital for cellular function and associated with various genetic diseases.
The team employed an innovative vesicle proteomics platform to systematically identify new cargo proteins and essential accessory factors linked to two significant cellular transport complexes, AP-1 and AP-4. These findings provide a comprehensive map of previously unidentified cargo proteins and regulatory factors, enhancing our understanding of the secretory pathway, which functions as a cell’s postal service. This pathway is crucial for ensuring that proteins reach their designated locations within the cell, as errors can lead to severe physiological defects.
“For years, the field has struggled to comprehensively map the cargo repertoire of adaptor complexes like AP-1 and AP-4, whose malfunctions are directly linked to serious human conditions such as MEDNIK syndrome, X-linked intellectual disability, and AP-4 deficiency syndrome, ” said Prof. Guo. “However, the full list of proteins they transport has remained elusive.”
Through their innovative approach, the researchers reconstituted the transport process in vitro and applied quantitative proteomics. This method allowed the identification of proteins that require AP-1 or AP-4 for packaging into vesicles, transitioning from a fragmented understanding to a more holistic view of their cargo landscape. The study revealed new clients and unexpected cellular machinery that AP-4 relies on.
Breakthrough in Understanding Transport Mechanisms
The research team integrated in vitro vesicle reconstitution techniques with quantitative mass spectrometry. This powerful combination enabled them to create transport vesicles in a controlled environment, followed by a thorough analysis of their protein composition. Collaborating with Prof. Yao Zhong-Ping from The Hong Kong Polytechnic University (PolyU), the researchers identified specific cargo proteins that depend on AP-1 and AP-4 for their transport from the trans-Golgi network, a central sorting hub within the cell.
Among the significant discoveries was that the protein CAB45 is an AP-1-dependent cargo, while ATRAP emerged as a novel cargo for AP-4. A key finding addressed a longstanding question regarding the formation of transport vesicles by AP-4 without the well-known protein clathrin. The team found that two cytosolic factors, WDR44 and PRRC1, are essential for AP-4-mediated trafficking. When these factors were depleted, crucial AP-4 cargoes, such as ATG9A and ATRAP, failed to exit their organelles, resulting in defects in vital cellular processes like autophagy.
“Our findings not only reveal new cargo clients and essential co-factors for AP-1 and AP-4 but also provide a powerful toolkit for the scientific community to dissect the mechanisms of vesicular trafficking,” Prof. Guo noted. “This opens new avenues for researching the pathological mechanisms of related diseases and potentially identifying new therapeutic targets.”
The study significantly enhances our understanding of cellular transport processes, with implications for developing treatments for various genetic disorders. The co-corresponding authors of the study are Prof. Guo Yusong of HKUST and Prof. Yao Zhong-Ping of PolyU. Dr. Peng Ziqing, a postdoctoral researcher at HKUST, serves as the first author of this pivotal research.
