Two alanines juxtaposed to aggrecan's G1 domain alter its intracellular localization.
Nascent proteins translated and processed in the endoplasmic reticulum (ER) sometimes contain intrinsic signals for ER retention or ER retrieval. These signals are usually a few amino acids in length, and if alanine modifications are made within these sequences, normal transit patterns of the nascent protein frequently change. The purpose of this study was to determine whether two alanines juxtaposed to the first globular domain of aggrecan's core protein affect its transit in Chinese hamster ovary (CHO) cells. Results show that two alanines juxtaposed to the first globular domain (G1AA) minimized secretion of the protein. However, transgenic proteins with juxtaposed glutamate-phenylalanine (G1EF) or no additional amino acids (G1) were still secreted. GFP-tagged G1AA localized in the lumen of the ER but not in the Golgi. In contrast, a portion of GFP-tagged G1EF and G1 did appear in the Golgi compartment. More importantly, unique and striking accumulations of G1EF and G1 transgenic proteins were seen in large dilated regions of the ER cisternae, reminiscent of accumulations seen in alpha1-antitrypsin deficiency disease. G1AA transgenic proteins did not form these vesicles but were diffusely distributed throughout the ER lumen. These results indicate that just two juxtaposed alanines can profoundly affect a large globular protein's intracellular localization.
Oliver
Bonnie L
BL
Department of BioStructure and Function, School of Dental Medicine, University of Connecticut Health Center, Farmington, Connecticut 06030, USA. boliver@up.uchc.edu
Cronin
Chunxia G
CG
Bue
Catherine
C
Hand
Arthur R
AR
Tanzer
Marvin L
ML
D053674:Aggrecans; D000409:Alanine / Q000378:metabolism*; D000818:Animals; D001483:Base Sequence; D022181:COP-Coated Vesicles / Q000378:metabolism; D002451:Cell Compartmentation / Q000502:physiology; D002462:Cell Membrane / Q000378:metabolism; D020755:Coat Protein Complex I / Q000378:metabolism; D006224:Cricetinae; D003412:Cricetulus; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D016326:Extracellular Matrix Proteins; D006056:Golgi Apparatus / Q000378:metabolism*; D006801:Humans; D037181:Lectins, C-Type; D008856:Microscopy, Fluorescence; D016253:Microscopy, Immunoelectron; D008969:Molecular Sequence Data; D009154:Mutation; D017434:Protein Structure, Tertiary; D021381:Protein Transport / Q000502:physiology; D011509:Proteoglycans / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism
D053674:Aggrecans; D020755:Coat Protein Complex I; D016326:Extracellular Matrix Proteins; D037181:Lectins, C-Type; D011509:Proteoglycans; D011993:Recombinant Fusion Proteins; D000409:Alanine