Identification of the motifs and amino acids in aggrecan G1 and G2 domains involved in product secretion.
Members of the large aggregating chondroitin sulfate proteoglycans are characterized by an N-terminal fragment known as G1 domain, which is composed of an immunoglobulin (IgG)-like motif and two tandem repeats (TR). Previous studies have indicated that the expressed product of aggrecan G1 domain was not secreted. Here we demonstrated that the inability of G1 secretion was associated with the tandem repeats but not the IgG-like motif, and specifically with TR1 of aggrecan. We also demonstrated that the G2 domain, a domain unique to aggrecan, had a similar effect on product secretion. The sequence of TR1 of G1 is highly conserved across species, which suggested similar functions played by these motifs. In a yeast two-hybrid assay, TR1 interacted with the calcium homeostasis endoplasmic reticulum protein. Deletion/mutation experiments indicated that the N-terminal fragment of TR1, in particular, the amino acids H(2)R(4) of this motif were key to its effect on product secretion. However, the N-terminal 55 amino acids were required to exert this function. Taken together, our study suggests a possible molecular mechanism for the function of the tandem repeats in product processing.
Kiani
Chris
C
Sunnybrook & Women's College Health Sciences Centre and Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, Canada.
Chen
Liwen
L
Lee
Vivian
V
Zheng
Peng-Sheng
PS
Wu
Yaojiong
Y
Wen
Jianping
J
Cao
Liu
L
Adams
Mark E
ME
Sheng
Wang
W
Yang
Burton B
BB
D053674:Aggrecans; D020816:Amino Acid Motifs; D000818:Animals; D001483:Base Sequence; D019556:COS Cells; D002417:Cattle; D002645:Chickens; D011508:Chondroitin Sulfate Proteoglycans / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D004721:Endoplasmic Reticulum / Q000737:chemistry; D016326:Extracellular Matrix Proteins; D006801:Humans; D037181:Lectins, C-Type; D008565:Membrane Proteins / Q000378:metabolism; D051379:Mice; D016296:Mutagenesis / Q000235:genetics; D017434:Protein Structure, Tertiary; D011509:Proteoglycans / Q000235:genetics / Q000378:metabolism*; D051381:Rats; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D020449:Repetitive Sequences, Amino Acid / Q000502:physiology; D015183:Restriction Mapping; D017386:Sequence Homology, Amino Acid; D014162:Transfection; D020798:Two-Hybrid System Techniques; D015003:Yeasts / Q000235:genetics
C508320:Acan protein, mouse; C508321:Acan protein, rat; D053674:Aggrecans; D011508:Chondroitin Sulfate Proteoglycans; D016326:Extracellular Matrix Proteins; D037181:Lectins, C-Type; D008565:Membrane Proteins; D011509:Proteoglycans; D011994:Recombinant Proteins