Date of Presentation

4-17-2020

College

College of Science & Mathematics

Faculty Sponsor(s)

Claude Krummenacher

Poster Abstract

Nectin-1 is an adhesion protein that also serves as a receptor for herpes simplex virus (HSV) to enter cells. In epithelial tissues nectin-1 proteins naturally bind to other nectin-1 molecules on adjacent cells to organize intercellular junctions. Recently, our lab showed that human nectin-1 also binds to the natural killer (NK) cell receptor CD96 (Holmes et al, PloS One, 2019). CD96 modulates the immune response of NK cells, which are part of the innate immune system of mammals. NK cells play an important role in eliminating virally infected cells and cancer cells. When HSV infection occurs, nectin-1 located at the cell surface becomes internalized by endocytosis. We hypothesize that HSV down-regulates nectin-1 from the surface of infected cells to prevent their detection and destruction by NK cells. To study how nectin-1 and CD96 proteins bind to each other at cell contacts, cell lines expressing fluorescently tagged nectin-1 and CD96 are being generated. Previously, we used mouse melanoma B78H1 cells as a model which are normally deficient in both nectin-1 and CD96. B78H1 cells were transfected to stably express fluorescent human nectin-1 (hNectin-1GFP, green), however expression of fluorescent human CD96 (hCD96mCherry, red) appears to be unstable in these cells. Here, human cell lines 293 and K562 are being used. 293 cells are robust human embryonic kidney cells that are being used as a positive control for transfection. K562 cells are human myelogenous leukemia cells that have been shown previously in this lab to be deficient in nectin-1 and CD96. We are currently attempting selection of clonal cell lines for these constructs by fluorescent microscopy and flow cytometry. The cells that express fluorescent CD96 and nectin-1 will be mixed in co-cultures to observe co-localization of nectin-1 and CD96 at cell contacts. This approach will provide a unique setting to investigate and understand how HSV affects the NK cell innate immune response.

Student Keywords

binding, nectin-1, CD96, human cells, herpes simplex virus, innate immune system

Disciplines

Biology

DOI

10.31986/issn.2689-0690_rdw.buss.1006

Included in

Biology Commons

Share

COinS
 
Apr 17th, 12:00 AM

Assessing the binding of nectin-1 and CD96 in human cells to investigate how herpes simplex virus evades the innate immune system

Nectin-1 is an adhesion protein that also serves as a receptor for herpes simplex virus (HSV) to enter cells. In epithelial tissues nectin-1 proteins naturally bind to other nectin-1 molecules on adjacent cells to organize intercellular junctions. Recently, our lab showed that human nectin-1 also binds to the natural killer (NK) cell receptor CD96 (Holmes et al, PloS One, 2019). CD96 modulates the immune response of NK cells, which are part of the innate immune system of mammals. NK cells play an important role in eliminating virally infected cells and cancer cells. When HSV infection occurs, nectin-1 located at the cell surface becomes internalized by endocytosis. We hypothesize that HSV down-regulates nectin-1 from the surface of infected cells to prevent their detection and destruction by NK cells. To study how nectin-1 and CD96 proteins bind to each other at cell contacts, cell lines expressing fluorescently tagged nectin-1 and CD96 are being generated. Previously, we used mouse melanoma B78H1 cells as a model which are normally deficient in both nectin-1 and CD96. B78H1 cells were transfected to stably express fluorescent human nectin-1 (hNectin-1GFP, green), however expression of fluorescent human CD96 (hCD96mCherry, red) appears to be unstable in these cells. Here, human cell lines 293 and K562 are being used. 293 cells are robust human embryonic kidney cells that are being used as a positive control for transfection. K562 cells are human myelogenous leukemia cells that have been shown previously in this lab to be deficient in nectin-1 and CD96. We are currently attempting selection of clonal cell lines for these constructs by fluorescent microscopy and flow cytometry. The cells that express fluorescent CD96 and nectin-1 will be mixed in co-cultures to observe co-localization of nectin-1 and CD96 at cell contacts. This approach will provide a unique setting to investigate and understand how HSV affects the NK cell innate immune response.