Abstract
We have previously demonstrated that CD45 physically associates with the endoplasmic reticulum processing enzyme glucosidase II (GII). GII consists of the catalytic alpha-chain and an associated beta-chain. To gain insight into the basis of the association between CD45 and GII, we examined the biochemical requirements for the interaction. We show that the alpha-subunit is essential for the interaction. Interestingly, only a higher molecular weight form of GIIalpha is capable of associating with CD45 in a competitive situation where multiple GIIalpha isoforms are expressed. Further, transfection studies demonstrate that only isoforms containing the alternatively spliced sequence Box A1 are capable of binding CD45, although all isoforms are catalytically active. The interaction between CD45 and GII is dependent on the active site of GII, is mediated through the carbohydrate on CD45, and can be inhibited with mannose. Taken together, these results suggest that GIIalpha acts as a lectin and binds to CD45 in an exon-dependent manner. This lectin activity of GII may be a novel mechanism for the regulation of CD45 biology and play a role in immune function, possibly by regulating CD45 glycosylation.
Highlights
CD45 is a highly abundant, transmembrane, protein-tyrosine phosphatase expressed on all cells of hematopoietic origin [1]
The CD45-glucosidase II (GII) Interaction Requires the GII␣ Subunit—Recently, it was demonstrated that the BW5147 mutant cell line BW/PHAr is deficient in expression of the GII␣ subunit [23]
To reconstitute the association between CD45 and GII, there is a dependence on Endo H-sensitive carbohydrate on CD45 suggesting that GII is binding via a lectin-like interaction [11]
Summary
CD45 is a highly abundant, transmembrane, protein-tyrosine phosphatase expressed on all cells of hematopoietic origin [1]. A sample of the BW/T200Ϫ lysate before and after the sequential reconstitution assays was taken and examined for the presence of GII␣ protein and GII enzymatic activity.
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