iProXProteomicsLin GuoMus Musculushttp://www.iprox.org/page/project.html?id=IPX0001777000guol@whu.edu.cnWuhan UniversityiProXPseudorabies virus (PRV) has been widely used as a live trans-synaptic tracer for mapping neuronal circuits. Systematically identifying mature PRV virion proteomes and defining co-purified host proteins are necessary to fully understand the detailed mechanism underlying PRV transmission processes. Here, a PRV virion purification strategy based on sorting with flow cytometry is developed and the mature extracellular and intracellular PRV virion proteomes using LC coupled with MS/MS are characterized. In addition to viral proteins, a large number of host proteins are also identified, including proteins related to actin cytoskeletal dynamics and membrane protrusion. How many of these host proteins are true virion components are unknown and the majority of these may not be. Through functional analysis, it is found that IRSp53 and fascin are critical for the egress process and play a role in direct cell-cell transmission. Moreover, it is shown that CDC42 and Rac1 are also involved in the production of mature extracellular virions. The results suggest that the formation of the filopodia-like cytoskeleton and the rearrangement of the membrane, which are both associated with IRSp53 and fascin, may be important for the transmission of viruses used in neuronal tracing.Proteomics Analysis Identifies IRSp53 and Fascin as Critical for PRV Egress and Direct Cell-Cell Transmission.Yu Fei-Long FL, Miao Huan H, Xia Jinjin J, Jia Fan F, Wang Huadong H, Xu Fuqiang F, Guo Lin LfalseProteomics analysis identifies IRSp53 and Fascin as critical for PRV egress and direct cell-cell transmissionPseudorabies virus (PRV) has been widely used as a live transsynaptic tracer for mapping neuronal circuits. Systematically identifying mature PRV virion proteomes and defining copurified host proteins are necessary to fully understand the detailed mechanism underlying PRV transmission processes. Here, we developed a PRV virion purification strategy based on sorting with flow cytometry and characterized the mature extracellular and intracellular PRV virion proteomes using liquid chromatography coupled with tandem mass spectrometry. In addition to viral proteins, a large number of host proteins were also identified, including proteins related to actin cytoskeletal dynamics and membrane protrusion. How many of these host proteins are true virion components are unknown and the majority of these may not be. Through functional analysis, we found that IRSp53 and fascin were critical for the egress process and play a role in direct cell-cell transmission. Moreover, we showed that CDC42 and Rac1 were also involved in the production of mature extracellular virions. Our results suggest that the formation of the filopodia-like cytoskeleton and the rearrangement of the membrane, which are both associated with IRSp53 and fascin, may be important for the transmission of viruses used in neuronal tracing.Wed Sep 18 00:00:00 BST 2019PXD0154901009031531927