Campari2 genomic interrogation of homeostatic calcium activity identifies TIM1 as a negative regulator of T cell function

Calcium signals regulate crucial cellular functions yet many genes coding for Ca²⁺handling proteins remain unknown as their identification relies on low-throughput single-cell approaches. Here we describe a method to measure Ca²⁺ activity using CaMPARI2, flow cytometry and pooled genome interrogation. CAMPARI2 screen (CaMP-Screen) identified enhancers and inhibitors of homeostatic Ca²⁺ activity, highlighting a predominant role for store-operated Ca²⁺ entry (SOCE) and lipid signalling pathways. Genes reducing basal Ca²⁺ activity were linked to Prader Willy syndrome, T cell dysfunction, and deafness. Silencing of HAVCR1 gene, coding for T cell transmembrane immunoglobulin and mucin (TIM1), enhanced Ca²⁺ signals in T cells and promoted signaling under resting but not after TCR engagement. Our findings establish CaMP-Screen as an efficient detector of low-amplitude Ca²⁺ signals and identify new genes associated to pathologies that regulate Ca²⁺ homeostasis, reporting TIM1 as a negative regulator of Ca²⁺ signals driving T cell function.