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. 2012 Feb 21;7(2):e31689. doi: 10.1371/journal.pone.0031689

Figure 1. LCRMP-1 is a substrate of and phosphorylated by GSK3β at Thr-628.

Figure 1

(A) Protein sequence analysis showed the potential consensus site of LCRMP-1 for the phosphorylation by GSK3β. Protein sequences are aligned among CRMP2, CRMP-1, and LCRMP-1. The numbers represent amino acid sites and underline shows a potential phosphorylation site for Cdk5. (B) GSK3β phosphorylates LCRMP-1 (WT) in vivo. HEK293T cells were co-transfected with Flag-tagged LCRMP-1 and distinct activity of Flag-tagged GSK3β (WT, CA and KD form). Equal amount of plasmids were transfected into every condition by using empty vectors. Cells were lysed 30 hr post-transfection and protein extracts were analyzed by immunoblotting with anti-Flag antibodies. (C) GSK3β phosphorylates LCRMP-1 (WT) at Thr-628 in vivo. CL1-0 cells were co-transfected either Flag-tagged LCRMP-1(WT) or LCRMP-1 (T628A, T628D) mutants in the presence or absence of distinct activity of Flag-tagged GSK3β (CA and KD form). Empty vectors were used for supplement to equal amount of plasmids in transfection assay. Cell lysates were harvested 48 hr post-transfection and analyzed by immunoblotting with anti-Flag and anti-β-actin antibodies.