PT - JOURNAL ARTICLE AU - Catherine B. Carbone AU - Nadja Kern AU - Ricardo A. Fernandes AU - Enfu Hui AU - Xiaolei Su AU - K. Christopher Garcia AU - Ronald D. Vale TI - In vitro reconstitution of T cell receptor–mediated segregation of the CD45 phosphatase AID - 10.1101/097600 DP - 2016 Jan 01 TA - bioRxiv PG - 097600 4099 - http://biorxiv.org/content/early/2016/12/31/097600.short 4100 - http://biorxiv.org/content/early/2016/12/31/097600.full AB - Binding of the T cell receptor (TCR) to peptide-major histocompatibility complex (pMHC) initiates signaling leading to T cell activation. Signaling is proposed to be triggered by the segregation of the transmembrane phosphatase CD45 from TCR-pMHC, caused by a difference in sizes of their extracellular domains. Here, we have reconstituted the segregation of CD45 from TCR-pMHC using purified proteins on model membranes. Similar to the sensitivity of T cell signaling, TCR-pMHC interactions with Kds of ≤15 μM were needed to exclude CD45; the larger of two developmentally-regulated isoforms of CD45 was excluded more rapidly and efficiently. We further show that two receptor-ligand pairs, TCR-pMHC and the inhibitory co-receptor PD-1 with its ligand PD-L1, segregate from one another and together create zones of CD45 exclusion. These results demonstrate that the binding energy of TCR-pMHC at membranemembrane interfaces is sufficient to create spatial partitioning of the TCR relative to CD45.