Functional Domains of the HSV-1 UL37 Protein in Virus Infectivity and Retrograde Transport in Neurons

Author

Carolyn Clark, Louisiana State University and Agricultural and Mechanical CollegeFollow

Degree

Doctor of Philosophy (PhD)

Department

Pathobiological Sciences

Document Type

Dissertation

Abstract

Herpes simplex virus type 1 is large alphaherpesvirus with 20 envelope proteins and 23 tegument proteins. Virus entry is via fusion of the viral envelope with cellular membranes mediated by a complex of viral glycoproteins. Glycoprotein B (gB) is the only fusogenic glycoprotein, while other viral glycoproteins and membrane proteins including gD, gH, gL, gK and UL20 membrane protein modulate gB’s fusogenicity. Previously, Our laboratory developed the HSV-1 (VC2) vaccines strain with partial deletions in the amino termini of gK (ΔgK(31–68)) and the UL20 membrane protein (ΔUL20(4–22)). VC2 enters epithelial cells only via endocytosis followed by release of the capsid into the cytoplasm from endocytic vesicles, but is unable to enter into neuronal axons that occurs exclusively via membrane fusion. Tegument proteins serve important roles in virus infectivity immediately following capsids deposition into the cytoplasm of infected cells. The UL37 tegument protein is a deamidase, which deactivates pathogen recognition receptors cGAS and RIG-I. Therefore, we engineered a C819S substitution in UL37 to inactivate the predicted deamidase active site. This UL37 C819S mutation enhanced replication and spread of VC2 approaching replication levels torly wild-type virus. Additionally, the C819S mutation engineered into the wild-type genome independently increased secretion of GM-CSF, however, it did not further increase GM-CSF secretion produced by the VC2 virus. These results suggest that the observed increase of GM-CSF secretion by the the C819S mutation in the context of the wild-type virus affected the same intracellular functions affected by the VC2 mutations. These results suggest that the C819 deamidation site on UL37 may be important for its direct or indirect interactions with UL20 and gK affecting viral assembly, egress and spread.

It has been reported that the UL37 protein is involved in retrograde transport of capsids in neuronal axons. Furthermore, our laboratory has shown that the UL37 binds the Dynein intermediate chain. Specifically, a combination of 5 alanine substitutions of UL37 at Q403, E452, Q455, Q511, and R515 (called HSV-1 (R2)) in the HSV-1(F) genetic background prevented the virus from efficiently transporting through neuronal axons. Testing various amino and carboxyl deletions of UL37 for ability to inhibit virus replication and retrograde transport revealed that a 480 amino terminal deletion in the HSV-1(KOS) genetic background allowed the virus to replicate fairly efficiently and to be transported in a retrograde manner in neuronal axons. These results combined suggested that the Q511A and/or the R515A mutations alone or combined may be responsible for the R2 defect in retrograde transport in neurons. The Q511A and the R515A mutations were engineered in the HSV-1(McKrae) genetic background, however, these viruses could be recovered indicating that these mutations alone or in combination were lethal for virus replication. However, transfection-complementation of UL37 genes containing either or both mutations efficiently complemented a UL37-null virus.

The UL37 protein physically interacts with the UL37 tegument protein, and these interactions and intracellular transport of both proteins are independent of capsid formation. Examination of the amino acid sequences of the HSV-1(F), HSV-1(KOS) and HSV-1(McKrae) revealed that the McKrae UL37 protein contained amino acid domains that differed from those of KOS and F strains. Therefore, it is likely that the Q511A and R515A mutations are lethal in the context of the McKrae genetic background because the structure of the UL36/Ul37 protein complex differs from those encoded by the KOS and F viral strains. Therefore, it is predicted that the R2 mutations in the McKrae background will produce a lethal phenotype. Overall, these results underscore the importance of the UL36/UL37 protein complex in virus infectivity and retrograde transport in neurons.

Date

4-22-2023

Recommended Citation

Clark, Carolyn, "Functional Domains of the HSV-1 UL37 Protein in Virus Infectivity and Retrograde Transport in Neurons" (2023). LSU Doctoral Dissertations. 6121.
https://repository.lsu.edu/gradschool_dissertations/6121

Committee Chair

Kousoulas, Konstantin G.

DOI

10.31390/gradschool_dissertations.6121