Tyrosine and serine phosphorylation regulate the conformation and subsequent threonine phosphorylation of the L1 cytoplasmic domain.
Previously we identified threonine-1172 (T1172) in the cytoplasmic domain of the cell adhesion molecule L1 as phosphorylated in pancreatic cancer cells. Although both CKII- and PKC-blockade suppressed this modification, only CKII was capable of phosphorylating T1172 of a recombinant L1 cytoplasmic domain, suggesting the requirement for additional events to facilitate availability of T1172 to PKC. In this study, we demonstrate that the region around T1172 exists in distinct conformations based on both T1172 phosphorylation and the integrity of surrounding residues. We further demonstrate the role of membrane-proximal and membrane-distal residues in regulating cytoplasmic domain conformation, and that modification of 3 of the 4 tyrosines in the L1 cytoplasmic domain promote conformational changes that facilitate other events. In particular, phenylalanine-substitution of tyrosine-1151 or tyrosine-1229 promote opening up of the cytoplasmic domain in a manner that facilitates phosphorylation of the other 3 tyrosines, as well as phosphorylation of T1172 by PKCalpha. Importantly, we show that phosphorylation of serine-1181 is required for T1172 phosphorylation by CKII. These data define a specific role for secondary structure in regulating the availability of T1172 that facilitates phosphorylation by PKC.
Chen
Maxine M
MM
Moores Cancer Center, University of California, San Diego, La Jolla, CA 92093-0803, USA.
Leland
Hyuma A
HA
Lee
Chia-Yao
CY
Silletti
Steve
S
D000595:Amino Acid Sequence; D045163:Antibodies, Phospho-Specific / Q000276:immunology; D047390:Casein Kinase II / Q000378:metabolism; D003593:Cytoplasm / Q000378:metabolism; D000939:Epitopes / Q000276:immunology; D006801:Humans; D008969:Molecular Sequence Data; D009154:Mutation; D039842:Neural Cell Adhesion Molecule L1 / Q000737:chemistry* / Q000235:genetics; D010766:Phosphorylation; D017510:Protein Folding; D051571:Protein Kinase C-alpha / Q000378:metabolism; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D012694:Serine / Q000737:chemistry* / Q000235:genetics; D013912:Threonine / Q000737:chemistry*; D014443:Tyrosine / Q000737:chemistry* / Q000235:genetics
D045163:Antibodies, Phospho-Specific; D000939:Epitopes; D039842:Neural Cell Adhesion Molecule L1; D013912:Threonine; D014443:Tyrosine; D012694:Serine; D047390:Casein Kinase II; D051571:Protein Kinase C-alpha