Molecular association of herpes simplex virus type 1 glycoprotein E with membrane protein Us9

Abstract

Herpes simplex virus type 1 (HSV-1) glycoprotein E (gE), glycoprotein I (gI), and Us9 promote efficient anterograde axonal transport of virus from the neuron cytoplasm to the axon terminus. HSV-1 and PRV gE and gI form a heterodimer that is required for anterograde transport, but an association that includes Us9 has not been demonstrated. NS-gE380 is an HSV-1 mutant that has five amino acids inserted after gE residue 380, rendering it defective in anterograde axonal transport. We demonstrated that gE, gI and Us9 form a trimolecular complex in Vero cells infected with NS-gE380 virus in which gE binds to both Us9 and gI. We detected the complex using immunoprecipitation with anti-gE or anti-gI monoclonal antibodies in the presence of ionic detergents. Under these conditions, Us9 did not associate with gE in cells infected with wild-type HSV-1; however, using a nonionic detergent, TritonX-100, an association between Us9 and gE was detected in immunoprecipitates of both wild-type and NS-gE380-infected cells. The results suggest that the interaction between Us9 and gE is weak and disrupted by ionic detergents in wild-type infected cells. We postulate that the tight interaction between Us9 and gE leads to the antero-grade spread defect in the NS-gE380 virus.

Invasion of the Nervous System

“…Although the mechanism of axonal sorting is not fully understood, gI, gE and US9 could potentially change the balance of microtubule-dependent motor recruitment to increase the yield of viral sorting into axons. Mutant viruses lacking these proteins are unable to sort viral cargo into axons, and infection cannot spread to post-synaptically connected neurons (Awasthi and Friedman, 2016;Brideau et al, 2000;DuRaine et al, 2017;Howard et al, 2013;Husak et al, 2000;Kratchmarov et al, 2013;Polcicova et al, 2005;Snyder et al, 2008). In PRV, US9-null mutants have a strong defect in axonal sorting of virion components (Brideau et al, 2000;Lyman et al, 2007), but these mutants have no defects either in retrograde axonal transport or during infection of non-polarized cells.…”

A putative WAVE regulatory complex (WRC) interacting receptor sequence (WIRS) in the cytoplasmic tail of HSV-1 gE does not function in WRC recruitment or neuronal transport

“…The Input and Bound % represent the proportion of the total sample loaded per gel. HSV-1 pU S 9 was used as a positive control for gE interaction [ 89 ]. Highlighted in the red box is the presence of WRC subunit Sra1 (145 kDa) in the bound fraction of both 17–37 gE-GFP and 17–37 gE(AA)-GFP infected lysates.…”

“…Confluent monolayers of HeLa or HaCaT cells in T75 flasks were infected at an MOI of 3 and incubated for 24 h. Cells were lysed and the soluble fraction used as input for GFP-Trap co-immunoprecipitation.The Input and Bound % represent the proportion of the total sample loaded per gel. HSV-1 pU S 9 was used as a positive control for gE interaction[89]. Highlighted in the red box is the presence of WRC subunit Sra1 (145 kDa) in the bound fraction of both 17-37 gE-GFP and 17-37 gE(AA)-GFP infected lysates.…”

Discovery and Characterization of an Aberrant Small Form of Glycoprotein I of Herpes Simplex Virus Type I in Cell Culture

“…In retrospect, the mature form of HSV-1 gI was reported to be a product of 66–68 kDa in size that can be recognized by monoclonal antibody Fd69 and polyclonal antibodies ( 15 , 32 – 37 ). In the absence of N-linked carbohydrates, the product has a molecular weight of about 58 kDa ( 33 ).…”