total src  (Cell Signaling Technology Inc)

Journal: Frontiers in cellular and infection microbiology

Article Title: Heparan sulfate proteoglycans remodel SARS-CoV-2 spike conformation to allow integrin interaction and infection of endothelial cells.

doi: 10.3389/fcimb.2025.1552116

Figure Lengend Snippet: FIGURE 6 SARS-CoV-2 Omicron BA.1 induces FAK/Src and ERK signaling after avb3 interaction. HL-mECs were not infected (NI) or infected with Omicron SARS-CoV-2 belonging to BA.1 and only XBB.1.5 sublineages (Omicron BA.1 and Omicron XBB.1.5, respectively) at a MOI of 1 for 1 h at 37°C. After 1 h p.i., cell lysates were analyzed by western blotting with anti-pFAK, anti-pSRC and anti-pERK antibodies (A-C, respectively). Blots are representative of three independent experiments with similar results. Quantification was carried out by densitometric analysis and plotting of the pFAK, pSrc and pERK normalized on GAPDH, tSrc or tERK as indicated in the graph. Values reported are the means ± the SD of three independent experiments. Statistical analysis was performed by one-way ANOVA, and the Bonferroni post-test was used to compare data (*p = 0.05, **p = 0.022, ***p = 0.0008). When indicated HL-mECs were pretreated with a mAb against avb3 integrin (mAb anti- avb3) or HSPGs inhibitors and analyzed for FAK/Src and ERK phosphorylation status (D-F).

Article Snippet: The proteins were purified from HEK293 cells with a purity ≥ 90% and run at a higher molecular weight by SDS-PAGE due to glycosylation; anti-phospho-extracellular signal-regulated kinase 1/ 2 (pERK) and anti-total ERK1/2 (tERK) antibodies were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA); antiphospho-focal adhesion kinase (pFAK), anti phospho-Src (pSrc), anti-Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and anti-total Src (tSrc) were purchased from Cell Signaling Technology (Danvers, MA, USA); recombinant human integrin alphaV beta3 (avb3), sodium chlorate, Heparinase III and heparin sodium salt from porcine intestinal mucosa were purchased from Merck (Darmstadt, Germany).

Techniques: Infection, Western Blot, Phospho-proteomics