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. 2023 Feb 9;21(2):e3001967. doi: 10.1371/journal.pbio.3001967

Fig 3. Confirmation that LRRC15 binds to SARS-CoV-2 spike protein.

Fig 3

(A) LRRC15 contains 15 leucine-rich repeats, a short cytoplasmic C-terminus, and 2 glycosylation sites. (B) Predicted protein structure of LRRC15 (from alpha fold). (C) LRRC15 is part of the LRR-Tollkin family. (D) Flow cytometry analysis of Alexa Fluor-647 (Spike647) binding in WT HEK293T cells, (E) HEK293T-ACE2, and (F) HEK293T cells with stable expression of ACE2 cDNA and TMPRSS2 cDNA (HEK293T-ACE2-TMPRSS2). Each cell line was transfected with plasmids encoding cDNA for GFP-tagged LRRC15 (transcript 1 or 2) or with empty GFP vector as negative control plasmid. (G). Histogram summary shows MFI of (D–F). (H) Representative images of interaction between LRRC15-GFP and Alexa Fluor 647-conjugated SARS-CoV-2 HexaPro spike protein in HEK293T cells (N = 2). Images were taken at 40× magnification. Green = LRRC15-GFP, red = Spike647, blue = Hoechst-stained nuclei. Scale bar = 25 μm. (I) Immunoprecipitation of LRRC15 with spike protein. Lysates of HEK293T cells transfected with GFP-tagged LRRC15 (transcript 1 or 2, LRRC15_1 and LRRC15_2, respectively) incubated with SARS-CoV-2 HexaPro spike protein were immunoprecipitated using anti-LRRC15 primary antibody. Immunoblots were performed for LRRC15 and for SARS-CoV-2 HexaPro spike. I = input, FT = flow-through, E = elute. The data underlying all panels in this figure can be found in DOI: 10.5281/zenodo.7416876. ACE2, angiotensin-converting enzyme 2; GP5, glycoprotein V platelet; LRRC15, leucine-rich repeat-containing protein 15; MFI, mean fluorescence intensity; SARS‑CoV‑2, Severe Acute Respiratory Syndrome Coronavirus 2; TLR, toll-like receptor; TMPRRS2, Transmembrane serine protease 2.