Mechanisms of nuclear egress in herpesviruses

PROJECT SUMMARY/ABSTRACT Herpesviruses use an unusual pathway to export their capsids from the nucleus where they are formed into the cytoplasm where they mature into infectious virions. Instead of using the canonical route through the nuclear pore, capsids undergo budding at the inner nuclear membrane to form perinuclear enveloped virions that subsequently fuse with the outer nuclear membrane, releasing the capsids into the cytoplasm. The nuclear egress complex (NEC), composed of two virally encoded proteins, is the key player in the membrane budding stage of this process. Previously, we discovered that the NEC from the prototypical a-herpesvirus, herpes simplex virus 1, is a complete virally encoded membrane-budding machine, the only one currently known to mediate budding of the nuclear, as opposed to cytoplasmic, membranes. We also showed that the NEC deforms and buds membranes by using a dual mechanism, in which membrane curvature is generated both locally (by peripheral membrane insertion) and globally (by the formation of a curved oligomeric scaffold). In addition, we have determined the high-resolution structures of several NEC homologs that are now used to guide functional studies. These results have transformed our understanding of how herpesviruses manipulate host membranes. The current application builds logically on these and other recent achievements and will leverage the combined power of structural and functional approaches to fill in the critical gaps in our understanding of both budding and fusion stages of nuclear egress. Aim 1 will expand the mechanistic and structural studies of the NEC to homologs from the distantly related g-herpesviruses. Aim 2 will visualize the nuclear egress process in infected cells in situ at a high resolution using cryoelectron tomography. Aim 3 will characterize a new host factor promoting fusion during nuclear egress. Our prior studies made important contributions to the current knowledge of how herpesvirus capsids bud at the nuclear envelope. We anticipate that upon successful completion, the proposed work will uncover new mechanisms and generate new models, thereby transforming our understanding of nuclear egress.