1. pUL21 regulation of pUs3 kinase activity influences the nature of nuclear envelope deformation by the HSV-2 nuclear egress complex
- Author
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Bruce W. Banfield, Michael A. Gulak, Jamil H. Muradov, Thomas J. M. Hay, and Renée L. Finnen
- Subjects
Physiology ,viruses ,Herpesvirus 2, Human ,Cultured tumor cells ,medicine.disease_cause ,Pathology and Laboratory Medicine ,Biochemistry ,Chlorocebus aethiops ,Medicine and Health Sciences ,Biology (General) ,Post-Translational Modification ,Phosphorylation ,Virus Release ,0303 health sciences ,Chemistry ,Kinase ,030302 biochemistry & molecular biology ,Viral tegument ,Transfection ,3. Good health ,Cell biology ,Body Fluids ,medicine.anatomical_structure ,Blood ,Herpes Simplex Virus-2 ,Medical Microbiology ,Viral Pathogens ,Viruses ,Cell lines ,Cellular Structures and Organelles ,Anatomy ,Pathogens ,Biological cultures ,Research Article ,Herpesviruses ,QH301-705.5 ,Viral protein ,Imaging Techniques ,Nuclear Envelope ,Immunology ,Phosphatase ,DNA construction ,Protein Serine-Threonine Kinases ,Research and Analysis Methods ,Microbiology ,03 medical and health sciences ,Viral Proteins ,Capsid ,Nuclear Membrane ,Virology ,Fluorescence Imaging ,Genetics ,medicine ,Inner membrane ,Animals ,Humans ,HeLa cells ,Nuclear membrane ,Kinase activity ,Molecular Biology Techniques ,Molecular Biology ,Microbial Pathogens ,Vero Cells ,030304 developmental biology ,Cell Nucleus ,Virus Assembly ,Organisms ,Biology and Life Sciences ,Proteins ,Herpes Simplex ,Cell Biology ,RC581-607 ,Blood Serum ,Cell cultures ,digestive system diseases ,Herpes Simplex Virus ,Herpes simplex virus ,Cytoplasm ,Plasmid Construction ,Parasitology ,Immunologic diseases. Allergy ,DNA viruses ,Immune Serum - Abstract
It is well established that the herpesvirus nuclear egress complex (NEC) has an intrinsic ability to deform membranes. During viral infection, the membrane-deformation activity of the NEC must be precisely regulated to ensure efficient nuclear egress of capsids. One viral protein known to regulate herpes simplex virus type 2 (HSV-2) NEC activity is the tegument protein pUL21. Cells infected with an HSV-2 mutant lacking pUL21 (ΔUL21) produced a slower migrating species of the viral serine/threonine kinase pUs3 that was shown to be a hyperphosphorylated form of the enzyme. Investigation of the pUs3 substrate profile in ΔUL21-infected cells revealed a prominent band with a molecular weight consistent with that of the NEC components pUL31 and pUL34. Phosphatase sensitivity and retarded mobility in phos-tag SDS-PAGE confirmed that both pUL31 and pUL34 were hyperphosphorylated by pUs3 in the absence of pUL21. To gain insight into the consequences of increased phosphorylation of NEC components, the architecture of the nuclear envelope in cells producing the HSV-2 NEC in the presence or absence of pUs3 was examined. In cells with robust NEC production, invaginations of the inner nuclear membrane were observed that contained budded vesicles of uniform size. By contrast, nuclear envelope deformations protruding outwards from the nucleus, were observed when pUs3 was included in transfections with the HSV-2 NEC. Finally, when pUL21 was included in transfections with the HSV-2 NEC and pUs3, decreased phosphorylation of NEC components was observed in comparison to transfections lacking pUL21. These results demonstrate that pUL21 influences the phosphorylation status of pUs3 and the HSV-2 NEC and that this has consequences for the architecture of the nuclear envelope., Author summary During all herpesvirus infections, the nuclear envelope undergoes deformation in order to enable viral capsids assembled within the nucleus of the infected cell to gain access to the cytoplasm for further maturation and spread to neighbouring cells. These nuclear envelope deformations are orchestrated by the viral nuclear egress complex (NEC), which, in HSV, is composed of two viral proteins, pUL31 and pUL34. How the membrane-deformation activity of the NEC is controlled during infection is incompletely understood. The studies in this communication reveal that the phosphorylation status of pUL31 and pUL34 can determine the nature of nuclear envelope deformations and that the viral protein pUL21 can modulate the phosphorylation status of both NEC components. These findings provide an explanation for why HSV-2 strains lacking pUL21 are defective in nuclear egress. A thorough understanding of how NEC activity is controlled during infection may yield strategies to disrupt this fundamental step in the herpesvirus lifecycle, providing the basis for novel antiviral strategies.
- Published
- 2021