VMP1 prevents Ca2+ overload in endoplasmic reticulum and maintains naive T cell survival

Abstract

Ca2+ in endoplasmic reticulum (ER) dictates T cell activation, proliferation, and function via store-operated Ca2+ entry. How naive T cells maintain an appropriate level of Ca2+ in ER remains poorly understood. Here, we show that the ER transmembrane protein VMP1 is essential for maintaining ER Ca2+ homeostasis in naive T cells. VMP1 promotes Ca2+ release from ER under steady state, and its deficiency leads to ER Ca2+ overload, ER stress, and secondary Ca2+ overload in mitochondria, resulting in massive apoptosis of naive T cells and defective T cell response. Aspartic acid 272 (D272) of VMP1 is critical for its ER Ca2+ releasing activity, and a knockin mouse strain with D272 mutated to asparagine (D272N) demonstrates all functions of VMP1 in T cells in vivo depend on its regulation of ER Ca2+. These data uncover an indispensable role of VMP1 in preventing ER Ca2+ overload and maintaining naive T cell survival.

TMEM41B is an endoplasmic reticulum Ca2+release channel maintaining T cell metabolic quiescence and responsiveness

“…The observed decrease in CD8 T cell number is likely attributed to heightened ER stress in these cells (Figures S3J and S3K). However, unlike VMP1 - deficient T cells, TMEM41B - deficient T cells did not exhibit mitochondrial Ca 2+ overload (Figures S3L and S3M), which caused massive apoptosis of VMP1 - deficient T cells 37 . Thus, TMEM41B plays a distinct role in T cells compared to VMP1.…”

“…To identify potential ER Ca 2+ release channel(s) in T cells, we performed a genome-wide CRISPR screening in Jurkat T cells, using the Ca 2+ -NFAT-PD-1 axis as a readout (Figures 1A and S1A), as recently published 37 . The rationale of this screening is that ER Ca 2+ levels determine SOCE, which, in turn, induces PD-1 expression in T cells through a NFAT-dependent manner 19,20,38 (Figure 1A).…”

“…(G) The interaction between RFP, TMEM41B or TMEM41B-K4A with COPI was examined by coimmunoprecipitation. The first lane was used in our previous study 37 .…”

“…Given that TMEM41B is an ER-resident membrane protein 27,28 , it is technically challenging to measure Ca 2+ transport across the ER membrane. We thus redirected TMEM41B to plasma membrane using a strategy that we put VMP1 on plasma membrane in our recent study 37 . TMEM41B possesses a lysine-rich Golgi-to-ER retrieve motif at its C-terminus (Figure 2F), which interacts with COPI complex facilitating the retro-transport of membrane proteins from the trans-Golgi back to ER 40 .…”

“…To investigate whether the Ca 2+ influx observed in HEK293T cells overexpressing TMEM41B or TMEM41B-K4A was SOCE, we repeated these experiments in STIM1- or ORAI1-deficient HEK293T cells incapable of inducing SCOE 37 . Ca 2+ influx induced by TMEM41B overexpression was abolished in STIM1- or ORAI1-deficient HEK293T cells, while TMEM41B-K4A-induced Ca 2+ influx remained unaffected (Figures S2A - S2C), demonstrating that Ca 2+ influx induced by TMEM41B overexpression is SOCE (due to its depletion of ER Ca 2+ ), while TMEM41B-K4A induced Ca 2+ influx is not SOCE.…”