Tim-1, Tim-1 Y276F, and Tim-1 cytoplasmic tail truncation were generated as described previously.
Jurkat cells were transfected as described above with pCDEF3 (empty vector), WT Tim-1, Tim-1 QGQ, or Tim-1 lacking the cytoplasmic tail truncation (Tim-1 ∆Cyto).
20×10 Jurkat T cells were transfected with empty vector, Tim-1, or Tim-1 QGQ.
Furthermore, while WT Tim-1 can enhance cytokine production, Tim-1 QGQ cannot.
Given the dramatic effect on Tim-1 localization, we further characterized the Tim-1 QGQ mutant.
Specifically, we tested the effect of Tim-1 Y276F, a cytoplasmic tail truncation (Tim-1 del.cyto), and Tim-1 244–246 KRK-QGQ (Tim-1 QGQ) on Tim-1 localization.
To define patterns of Tim-1 localization on T cells, we transfected Tim-1 into the murine Th2 line D10, which does not expresses endogenous Tim-1.
We observed that the Tim-1 QGQ construct has lower surface expression than wild type Tim-1, even when higher concentrations of Tim-1 QGQ plasmid are transfected.
BCR signaling and TIM-1 are closely related.
Next, a Tim-1 cytoplasmic tail truncation was utilized.
