Human FZD6 phosphorylation LC-MS/MS

Frizzleds (FZD) function as receptors for secreted lipoglycoproteins of the Wingless/Int-1 (WNT) family, initiating an important signal transduction network in multicellular organisms. FZDs are classified as G protein-coupled receptors (GPCRs), which control a wide variety of physiological functions. GPCRs are well known to be regulated by phosphorylation, which can lead to specification of downstream signaling or receptor desensitization. The role and underlying mechanisms of phosphorylation of FZDs remains largely unexplored. Here, we investigated the phosphorylation of human FZD6. In order to assess FZD6 phosphorylation experimentally, we employed mass spectrometry. HEK293 cells expressing FZD6-GFP were treated with either control, WNT-3A or WNT-5A containing conditioned medium for 30 min and processed for LC-MS/MS analysis. Six phosphorylated serine residues were detected in the C-terminus of FZD6 − S592, S620, S629, S648, S653 and S656.