Mutation analysis of the short cytoplasmic domain of the cell-cell adhesion molecule CEACAM1 identifies residues that orchestrate actin binding and lumen formation.
CEACAM1-4S (carcinoembryonic antigen cell adhesion molecule 1, with 4 ectodomains and a short, 12-14 amino acid cytoplasmic domain) mediates lumen formation via an apoptotic and cytoskeletal reorganization mechanism when mammary epithelial cells are grown in a three-dimensional model of mammary morphogenesis. We show by quantitative yeast two-hybrid, BIAcore, NMR HSQC and STD, and confocal analyses that amino acids phenylalanine (Phe(454)) and lysine (Lys(456)) are key residues that interact with actin orchestrating the cytoskeletal reorganization. A CEACAM1 membrane model based on vitamin D-binding protein that predicts an interaction of Phe(454) at subdomain 3 of actin was supported by inhibition of binding of actin to vitamin D-binding protein by the cytoplasmic domain peptide. We also show that residues Thr(457) and/or Ser(459) are phosphorylated in CEACAM1-transfected cells grown in three-dimensional culture and that mutation analysis of these residues (T457A/S459A) or F454A blocks lumen formation. These studies demonstrate that a short cytoplasmic domain membrane receptor can directly mediate substantial intracellular signaling.
Chen
Charng-Jui
CJ
Divisions of Immunology, Beckman Research Institute of the City of Hope, Duarte, California 91010, USA.
Kirshner
Julia
J
Sherman
Mark A
MA
Hu
Weidong
W
Nguyen
Tung
T
Shively
John E
JE
D000199:Actins / Q000235:genetics / Q000378:metabolism*; D019943:Amino Acid Substitution; D015703:Antigens, CD / Q000235:genetics / Q000378:metabolism*; D017209:Apoptosis / Q000502:physiology; D015815:Cell Adhesion Molecules / Q000235:genetics / Q000378:metabolism*; D018929:Cell Culture Techniques; D003599:Cytoskeleton / Q000235:genetics / Q000378:metabolism; D004847:Epithelial Cells / Q000166:cytology / Q000378:metabolism*; D005260:Female; D006367:HeLa Cells; D006801:Humans; D019169:Jurkat Cells; D042361:Mammary Glands, Human / Q000166:cytology / Q000378:metabolism*; D008958:Models, Molecular; D038081:Organogenesis / Q000502:physiology; D011485:Protein Binding / Q000502:physiology; D017434:Protein Structure, Tertiary / Q000502:physiology; D015398:Signal Transduction / Q000502:physiology*; D014809:Vitamin D-Binding Protein / Q000235:genetics / Q000378:metabolism
D000199:Actins; D015703:Antigens, CD; C069847:CD66 antigens; D015815:Cell Adhesion Molecules; D014809:Vitamin D-Binding Protein