Cholestrol dependent cytolysins (CDCs) are key virulence factors of pathogenic bacteria. CDCs are shown to activate diverse cellular responses in host cells such as cytoskeletal rearrangement, membrane trafficking and cell damage. It has been suggested that these responses critically depend on the initial Ca²⁺ signaling induced by the CDCs. However, the mechanistic basis of this Ca²⁺ signaling is controversial and poorly understood. Here, using streptolysin O (SLO), a CDC from the lung pathogen Streptococcus pyogenes, we studied the mechanistic basis of this Ca²⁺ signaling. We show that in H441 lung epithelial cells non-lytic concentration of SLO induces a complex oscillatory Ca²⁺ signaling. SLO not only mobilizes intracellular Ca²⁺ stores but also causes Ca²⁺ influx. Surprisingly these mechanisms were found to be independent of pore formation by SLO as non-pore forming mutant SLO-N402 activated similar Ca²⁺ signaling. We found that SLO induced Ca²⁺ oscillations (SICO) were sensitive to the blockers of store operated Ca²⁺ (SOC) entry and gene silencing of STIM1 and Orai1 abolished SICO. Furthermore, Ca²⁺ add-back experiments showed that SLO activated SOC entry. In STIM1 and Orai1 over-expressing cells, SLO-induced co-localization of STIM1 and Orai1 at the plasma membrane. Similarly, in primary cultivated alveolar type II (ATII) cells, SLO activated SOC channels but SICO was short-lived. RT-PCR and functional over-expression showed that expression pattern of STIM1 and Orai1 affects SICO. Hence, our results demonstrate that SLO-induced Ca²⁺ signaling depends on Ca²⁺ release and STIM1/Orai1 dependent SOC entry.