Molecular basis of LMAN1 in coordinating LMAN1-MCFD2 cargo receptor formation and ER-to-Golgi transport of FV/FVIII.
The LMAN1-MCFD2 (lectin, mannose binding 1/multiple coagulation factor deficiency protein 2) cargo receptor complex transports coagulation factors V (FV) and VIII (FVIII) from the endoplasmic reticulum (ER) to the ER-Golgi intermediate compartment (ERGIC). LMAN1 (ERGIC-53) is a hexameric transmembrane protein with a carbohydrate recognition domain (CRD) on the ER luminal side. Here, we show that mutations in the first beta sheet of the CRD abolish MCFD2 binding without affecting the mannose binding, suggesting that LMAN1 interacts with MCFD2 through its N-terminal beta sheet, consistent with recently reported crystal structures of the CRD-MCFD2 complex. Mutations in the Ca(2+)- and sugar-binding sites of the CRD disrupt FV and FVIII interactions, without affecting MCFD2 binding. This interaction is independent of MCFD2, as LMAN1 mutants defective in MCFD2 binding can still interact with FVIII. Thus, the CRD of LMAN1 contains distinct, separable binding sites for both its partner protein (MCFD2) and the cargo proteins (FV/FVIII). Monomeric LMAN1 mutants are defective in ER exit and unable to interact with MCFD2, suggesting that the oligomerization of LMAN1 is necessary for its cargo receptor function. These results point to a central role of LMAN1 in regulating the binding in the ER and the subsequent release in the ERGIC of FV and FVIII.
Zheng
Chunlei
C
Lerner Research Institute, Cleveland Clinic Foundation, Cleveland, OH, USA.
Liu
Hui-Hui
HH
Yuan
Shuguang
S
Zhou
Jiahai
J
Zhang
Bin
B
D000595:Amino Acid Sequence; D001692:Biological Transport; D015153:Blotting, Western; D002118:Calcium / Q000378:metabolism; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D005165:Factor V / Q000378:metabolism*; D005169:Factor VIII / Q000378:metabolism*; D005455:Fluorescent Antibody Technique; D006056:Golgi Apparatus / Q000378:metabolism*; D020662:Guanine Nucleotide Exchange Factors / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D047468:Immunoprecipitation; D037241:Mannose-Binding Lectins / Q000235:genetics / Q000378:metabolism*; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D011485:Protein Binding; D012333:RNA, Messenger / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D064067:Rho Guanine Nucleotide Exchange Factors; D017386:Sequence Homology, Amino Acid
D020662:Guanine Nucleotide Exchange Factors; C084004:LMAN1 protein, human; C577400:MCF2L protein, human; D037241:Mannose-Binding Lectins; D008565:Membrane Proteins; D012333:RNA, Messenger; D064067:Rho Guanine Nucleotide Exchange Factors; D005165:Factor V; D005169:Factor VIII; D002118:Calcium
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