Cell apoptosis is an important process that occurs during development or in response to stress stimuli such as oxidative stress. The serine-threonine kinase Akt enhances survival and suppress apoptosis. SHIP2 is known as a negative regulator of Akt. In addition to its lipid 5'-phosphatase activity, SHIP2 interacts and signals as a scaffolding complex with several proteins. Several findings have pointed out a possible role of SHIP2 in apoptosis regulation. However, the molecular mechanisms behind remain unknown. Using embryonic fibroblast lacking the lipid 5'-phosphatase domain as a genetic model system and human liver cancer cells treated with SHIP2 inhibitor (AS1949490), as a pharmacological model system. We provide the first evidence that SHIP2 regulates apoptosis independently of its 5'-phosphates activity. Indeed, absence of the 5'-phosphatase domain of SHIP2 did not prevent H2O2-induced apoptosis in fibroblasts. Whereas chemical inactivation or RNAi knockdown of SHIP2 blocked H2O2-induced apoptosis in HepG2 cells. We found that suppression of apoptosis upon SHIP2 inhibition is PI3K/Akt independent but rather MAP kinase dependent. In addition, we found that AS1949490 altered both 5'-phosphatase and scaffolding function of SHIP2. Indeed, AS1949490 mediated SHIP2 inhibition promotes protein complex formation of SHIP2 together with non-receptor tyrosine kinase SRC and ABL which in turn enhances PI3K/Akt and MAP kinase pathways activation. Dual inhibition of SRC/ABL blocked activation of both pathways upon SHIP2 inhibition and H2O2 treatment. Altogether, these findings indicate that SHIP2 protein play a determinant role in H2O2-induced apoptosis.