Blebs regulate phosphoinositide distribution and promote cell survival through the Septin-SH3KBP1-PI3K axis

Cells rely on substrate adhesion to activate diverse signaling pathways essential for proliferation and survival. In the absence of proper adhesion to the extracellular matrix, cells undergo anoikis, a form of programmed cell death. Poorly attached cells often exhibit rounded morphology and form dynamic protrusions called blebs. Although the role of blebs in amoeboid migration is well-documented, recent studies have highlighted their role in anoikis resistance. However, little is known about whether the most abundant membrane components-phospholipids-function in blebs-facilitated anoikis resistance. Here, we report an enrichment of phosphatidylinositol (3,4,5)-trisphosphate [PI(3,4,5)P₃] and a depletion of phosphatidylinositol (4,5)-bisphosphate [PI(4,5)P₂] at bleb membrane, compared with nonbleb regions of the plasma membrane. Our results have shown that both PI(3,4,5)P₃ and PI(4,5)P₂ have restricted diffusion patterns between the bleb and nonbleb membrane regions. Subsequently, we reveal that phosphoinositide 3-kinase (PI3K) is recruited by SH3KBP1 via liquid-liquid phase separation and interacted with septin at the bleb necks. This Septin-SH3KBP1-PI3K axis then contributes to differential phosphoinositide (PI) distribution and anoikis resistance. These novel insights into PI dynamics and the associated molecular scaffolding not only elucidate the mechanisms of bleb formation and anoikis resistance but also highlight potential targets for therapeutic interventions in anchorage-independent cancers.<b>NEW & NOTEWORTHY</b> We reveal that bleb membranes are enriched in PI(3,4,5)P₃ but depleted in PI(4,5)P₂ compared with non-bleb regions. We find that the Septin-SH3KBP1-PI3K axis establishes differential phosphoinositide distribution, regulates phosphoinositide diffusion, and confers resistance to anoikis in detached tumor cells, providing novel targets to counteract anoikis resistance in cancer.