A novel adapter protein employs a phosphotyrosine binding domain and exceptionally basic N-terminal domains to capture and localize an atypical protein kinase C: characterization of Caenorhabditis elegans C kinase adapter 1, a protein that avidly binds protein kinase C3.
Atypical protein kinase C isoforms (aPKCs) transmit regulatory signals to effector proteins located in the cytoplasm, nucleus, cytoskeleton, and membranes. Mechanisms by which aPKCs encounter and control effector proteins in various microenvironments are poorly understood. By using a protein interaction screen, we discovered two novel proteins that adapt a Caenorhabditis elegans aPKC (PKC3) for specialized (localized) functions; protein kinase C adapter 1 (CKA1, 593 amino acids) and CKA1S (549 amino acids) are derived from a unique mRNA by alternative utilization of two translation initiation codons. CKA1S and CKA1 are routed to the cell periphery by exceptionally basic N-terminal regions that include classical phosphorylation site domains (PSDs). Tethering of PKC3 is mediated by a segment of CKA1 that constitutes a phosphotyrosine binding (PTB) domain. Two aromatic amino acids (Phe(175) and Phe(221)) are indispensable for creation of a PKC3-binding surface and/or stabilization of CKA1.aPKC complexes. Patterns of CKA1 gene promoter activity and CKA1/CKA1S protein localization in vivo overlap with patterns established for PKC3 expression and distribution. Transfection experiments demonstrated that CKA1/CKA1S sequesters PKC3 in intact cells. Structural information in CKA1/CKA1S enables delivery of adapters to the lateral plasma membrane surface (near tight junctions) in polarized epithelial cells. Thus, a PTB domain and PSDs collaborate in a novel fashion in CKA1/CKA1S to enable tethering and targeting of PKC3. Avid ligation of a PKC isoform is a previously unappreciated function for a PTB module.
Zhang
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Department of Molecular Pharmacology, Atran Laboratories, Albert Einstein College of Medicine, Bronx, New York 10461, USA.
Wu
S L
SL
Rubin
C S
CS
D048868:Adaptor Proteins, Signal Transducing; D000595:Amino Acid Sequence; D000596:Amino Acids / Q000737:chemistry; D000818:Animals; D030801:Animals, Genetically Modified; D001483:Base Sequence; D001665:Binding Sites; D015152:Blotting, Northern; D015153:Blotting, Western; D017173:Caenorhabditis elegans / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D029742:Caenorhabditis elegans Proteins; D002352:Carrier Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D003062:Codon; D018076:DNA, Complementary / Q000378:metabolism; D004285:Dogs; D004591:Electrophoresis, Polyacrylamide Gel; D017353:Gene Deletion; D015723:Gene Library; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010649:Phenylalanine / Q000737:chemistry; D010766:Phosphorylation; D019000:Phosphotyrosine / Q000378:metabolism*; D011233:Precipitin Tests; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D014176:Protein Biosynthesis; D020033:Protein Isoforms; D011493:Protein Kinase C / Q000737:chemistry* / Q000378:metabolism; D017434:Protein Structure, Tertiary; D012333:RNA, Messenger / Q000378:metabolism; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D013552:Swine; D014158:Transcription, Genetic; D014162:Transfection; D014407:Tumor Cells, Cultured; D020798:Two-Hybrid System Techniques
D048868:Adaptor Proteins, Signal Transducing; D000596:Amino Acids; D029742:Caenorhabditis elegans Proteins; D002352:Carrier Proteins; D003062:Codon; D018076:DNA, Complementary; C425314:PKC adapter protein 2, C elegans; D020033:Protein Isoforms; D012333:RNA, Messenger; C425313:num-1 protein, C elegans; D019000:Phosphotyrosine; D010649:Phenylalanine; C115015:PKC-3 protein; D011493:Protein Kinase C