c Paramyxovirus membrane glycoproteins F (fusion protein) and HN, H, or G (attachment protein) are critical for virus entry, which occurs through fusion of viral and cellular envelopes. The F protein folds into a homotrimeric, metastable prefusion form that can be triggered by the attachment protein to undergo a series of structural rearrangements, ultimately folding into a stable postfusion form. In paramyxovirus-infected cells, the F protein is activated in the Golgi apparatus by cleavage adjacent to a hydrophobic fusion peptide that inserts into the target membrane, eventually bringing the membranes together by F refolding. However, it is not clear how the attachment protein, known as HN in parainfluenza virus 5 (PIV5), interacts with F and triggers F to initiate fusion. To understand the roles of various F protein domains in fusion triggering and metastability, single point mutations were introduced into the PIV5 F protein. By extensive study of F protein cleavage activation, surface expression, and energetics of fusion triggering, we found a role for an immunoglobulin-like (Ig-like) domain, where multiple hydrophobic residues on the PIV5 F protein may mediate F-HN interactions. Additionally, destabilizing mutations of PIV5 F that resulted in HN trigger-independent mutant F proteins were identified in a region along the border of F trimer subunits. The positions of the potential HN-interacting region and the region important for F stability in the lower part of the PIV5 F prefusion structure provide clues to the receptor-binding initiated, HN-mediated F trigger.

Oocytes of many species lack centrioles and therefore form acentriolar spindles. Despite the necessity of oocyte meiosis for successful reproduction, how these spindles mediate accurate chromosome segregation is poorly understood. We have gained insight into this process through studies of the kinesin-4 family member Kif4 in mouse oocytes. We found that Kif4 localizes to chromosomes through metaphase and then largely redistributes to the spindle midzone during anaphase, transitioning from stretches along microtubules to distinct ring-like structures; these structures then appear to fuse together by telophase. Kif4’s binding partner PRC1 and MgcRacGAP, a component of the centralspindlin complex, have a similar localization pattern, demonstrating dynamic spindle midzone organization in oocytes. Kif4 knockdown results in defective midzone formation and longer spindles, revealing new anaphase roles for Kif4 in mouse oocytes. Moreover, inhibition of Aurora B/C kinases results in Kif4 mislocalization and causes anaphase defects. Taken together, our work reveals essential roles for Kif4 during the meiotic divisions, furthering our understanding of mechanisms promoting accurate chromosome segregation in acentriolar oocytes.

In many enveloped virus families, including HIV and HSV, a crucial, yet unexploited, step in the viral life cycle is releasing particles from the infected cell membranes. This release process is mediated by host ESCRT complex proteins, which are recruited by viral structural proteins and provides the mechanical means for membrane scission and subsequent viral budding. The prazole drug, tenatoprazole, was previously shown to bind to ESCRT complex member Tsg101 and to quantitatively block the release of infectious HIV-1 from cells in culture. In this report we show that tenatoprazole and a related prazole drug, ilaprazole, effectively block infectious Herpes Simplex Virus (HSV)-1/2 release from Vero cells in culture. By electron microscopy, we found that both prazole drugs block the transit of HSV particles through the cell nuclear membrane resulting in their accumulation in the nucleus. Ilaprazole also quantitatively blocks the release of HIV-1 from 293T cells with an EC50 of 0.8-1.2 μM, which is much more potent than tenatoprazole. Our results indicate that prazole-based compounds may represent a class of drugs with potential to be broad-spectrum antiviral agents against multiple enveloped viruses, by interrupting cellular Tsg101 interaction with maturing virus, thus blocking the budding process that releases particles from the cell. Importance These results provide the basis for the development of drugs that target enveloped virus budding that can be used ultimately to control multiple virus infections in humans.

In many enveloped virus families, including HIV and HSV, a crucial, yet unexploited, step in the viral life cycle is releasing particles from the infected cell membranes. This release process is mediated by host ESCRT complex proteins, which is recruited by viral structural proteins and provides the mechanical means for membrane scission and subsequent viral budding. The prazole drug, tenatoprazole, was previously shown to bind to ESCRT complex member Tsg101 and quantitatively block the release of infectious HIV-1 from cells in culture. In this report we show that tenatoprazole and a related prazole drug, ilaprazole, effectively block infectious Herpes Simplex Virus (HSV)-1/2 release from Vero cells in culture. By electron microscopy, we found that both prazole drugs block the release of HSV particles from the cell nuclear membrane resulting in their accumulation in the nucleus. Ilaprazole also quantitatively blocks the release of HIV-1 from 293T cells with an EC50 of 0.8 µM, which is more potent than tenatoprazole. Finally, we synthesized and tested multiple novel prazole-based analogs that demonstrate both binding to Tsg101 and inhibition of viral egress in the nanomolar range of HIV-1 from 293T cells. Our results indicate that prazole-based compounds may represent a class of drugs with potential to be broad-spectrum antiviral agents against multiple enveloped viruses, by interrupting cellular Tsg101 interaction with maturing virus, thus blocking the budding process that releases particles from the cell. ImportanceThese results provide the basis for the development of drugs that target enveloped virus budding that can be used ultimately to control multiple virus infections in humans.