Inhibition of Tetraspanin Functions Impairs Human Papillomavirus and Cytomegalovirus Infections.

Laura Fast,Peter Monk,Luise Florin,Christian Sinzger,Jonas Schwickert,Snježana Mikuličić,Anna Fritzen,Fatima Boukhallouk,Henar Suarez,María Yáñez-Mó,Diana Lieber,Daniel Hochdorfer

doi:10.3390/ijms19103007

Abstract

Tetraspanins are suggested to regulate the composition of cell membrane components and control intracellular transport, which leaves them vulnerable to utilization by pathogens such as human papillomaviruses (HPV) and cytomegaloviruses (HCMV) to facilitate host cell entry and subsequent infection. In this study, by means of cellular depletion, the cluster of differentiation (CD) tetraspanins CD9, CD63, and CD151 were found to reduce HPV16 infection in HeLa cells by 50 to 80%. Moreover, we tested recombinant proteins or peptides of specific tetraspanin domains on their effect on the most oncogenic HPV type, HPV16, and HCMV. We found that the C-terminal tails of CD63 and CD151 significantly inhibited infections of both HPV16 and HCMV. Although CD9 was newly identified as a key cellular factor for HPV16 infection, the recombinant CD9 C-terminal peptide had no effect on infection. Based on the determined half-maximal inhibitory concentration (IC50), we classified CD63 and CD151 C-terminal peptides as moderate to potent inhibitors of HPV16 infection in HeLa and HaCaT cells, and in EA.hy926, HFF (human foreskin fibroblast) cells, and HEC-LTT (human endothelial cell-large T antigen and telomerase) cells for HCMV, respectively. These results indicate that HPV16 and HCMV share similar cellular requirements for their entry into host cells and reveal the necessity of the cytoplasmic CD151 and CD63 C-termini in virus infections. Furthermore, this highlights the suitability of these peptides for functional investigation of tetraspanin domains and as inhibitors of pathogen infections.

Highlights

Tetraspanin proteins play important roles during the life cycle of numerous viruses [1,2,3,4,5]
Cluster of Differentiation (CD) Tetraspanins CD9, CD63, CD81 and CD151 Are Required for Human Papillomavirus Type 16 (HPV16) Infection
To test whether additional tetraspanins are involved in HPV type 16 (HPV16) infection, we applied a pseudovirus (PsV) infection assay, depleting either CD9 or CD81. siRNAs targeting CD151 and CD63 served as a control for impaired infection

Summary

Introduction

Tetraspanin proteins play important roles during the life cycle of numerous viruses [1,2,3,4,5]. Tetraspanins localize in the plasma membrane and the membranes of intracellular compartments They are structurally conserved harboring four transmembrane domains, three intracellular domains as well as a small and a large extracellular loop [7,8,9,10,11]. The large extracellular loop (LEL) of tetraspanins is a flexible functional domain with a variable sequence but different tetraspanins share a conserved structure [12,13,14]. The short cytoplasmic C-terminal tail of tetraspanins serves as a connector to cytoplasmic interaction partners [15,16,17,18,19] Despite their relative shortness of about a dozen amino acids, some tetraspanin C-termini contain functional units, such as the C-termini of tetraspanins CD63 and CD151 that contain a tyrosine-based sorting motif essential for their subcellular localization [16,20]. The ability to form TEMs and co-integrate interaction partners such as growth factor receptors, adhesion molecules, and proteases into these assemblies enables local concentration of factors required for virus entry [5]

Methods

Results

Conclusion