JPT2/HN1L functions as an NAADP-binding protein in a cell type-specific manner

Nicotinic acid adenine dinucleotide phosphate (NAADP) is a second messenger evoking Ca{superscript 2} release from intracellular Ca{superscript 2} stores by targeting several Ca2+ channels including two-pore channels (TPC1/2), transient receptor potential mucolipin-1 (TRPML1), or ryanodine receptor type 1 (RYR1). For activation of Ca2+ channels, NAADP requires binding proteins, such as JPT2/HN1L and LSM12. So far, their function has been analyzed in several cell lines and only a very limited number of primary cells; however, their physiological relevance in cell types known to utilize NAADP signaling remains unclear. Here, we generated JPT2/HN1L-deficient (Jpt2/Hn1l-/-) mice to evaluate the contribution of JPT2/HN1L proteins to platelet aggregation, and Ca2+ signaling in cardiomyocytes, mast cells, and CD4 T cells. NAADP is known to contribute to collagen-related peptide (CRP)-evoked platelet aggregation, but this was not altered by JPT2/HN1L deletion. Functional Ca{superscript 2} imaging revealed that JPT2/HN1L plays a strikingly cell-type specific role in NAADP-mediated Ca2+ release. In beating ventricular cardiomyocytes, {beta}-adrenergic stimulation is known to evoke arrhythmogenic spontaneous diastolic Ca2+ transients, which were not altered in their frequency in Jpt2/Hn1l-/- myocytes. Further, antigen-evoked Ca2+ transients in peritoneal mast cells (PMCs) are not changed in Jpt2/Hn1l-/- PMCs. However, CD4 T cells displayed a pronounced requirement for JPT2/HN1L. Following T cell receptor/CD3 stimulation, global Ca{superscript 2} elevations and early NAADP-driven Ca{superscript 2} microdomains, which occur within tens of milliseconds of TCR/CD3 engagement and serve as initiating signals for downstream immune activation, were significantly decreased in Jpt2/Hn1l-/- CD4+ cells. We conclude that JPT2/HN1L is indispensable for NAADP-mediated Ca{superscript 2} release in T cells, but dispensable in cardiomyocytes, platelets, and mast cells, at least for the agonists employed. Accordingly, LSM12 might compensate for the loss of JPT2/HN1L. Together, JPT2/HN1L is not universally required as an NAADP-binding protein but exhibits cell-type specificity, with an essential function in T cell Ca{superscript 2} signaling. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=76 SRC="FIGDIR/small/687795v2_ufig1.gif" ALT="Figure 1"> View larger version (31K): org.highwire.dtl.DTLVardef@393a62org.highwire.dtl.DTLVardef@128045aorg.highwire.dtl.DTLVardef@35dc63org.highwire.dtl.DTLVardef@180363_HPS_FORMAT_FIGEXP M_FIG C_FIG HighlightsO_LIJPT2/HN1L has been identified as an NAADP-binding protein mediating NAADP-evoked Ca{superscript 2} release via its target channels TPC1, TPC2and RyR1. C_LIO_LIWe investigated the role of JPT2/HN1L in NAADP-dependent Ca{superscript 2} signaling across four distinct cell types. C_LIO_LINAADP-dependent Ca{superscript 2} responses were unaltered in JPT2/HN1L-/- cardiomyocytes, platelets, and mast cells. C_LIO_LIJPT2/HN1L is essential for NAADP-mediated CD4 T-cell receptor activation. C_LI