Bleb Formation in Human Fibrosarcoma HT1080 Cancer Cell Line Is Positively Regulated by the Lipid Signalling Phospholipase D2 (PLD2)

Abstract

Blebs are spherical plasma membrane protrusions formed when the membrane detaches from the underlying cortex as a result of actomyosin contractility-powered increase of hydrostatic pressure in the cytoplasm. Different tumour cells metastasize using blebbing as alternative mode of migration by squeezing through pre-existing pores in the extracellular matrix (ECM). This study investigated the role of the lipid signalling phospholipases D1 and D2 (PLD1/PLD2) in bleb formation in human fibrosarcoma HT1080 cell line in the extracellular matrix, and reports that pharmacological inhibition of PLD1 and PLD2 with a potent universal PLD inhibitor potently inhibited bleb formation in HT1080 cells embedded in three-dimensional (3D) matrigel matrix. Use of smartpool small interfering RNAs (siRNAs) that target PLD1 and PLD2 isoforms at four different sequences revealed that PLD2, but not PLD1 is involved in blebbing of HT1080 cells. Furthermore, we demonstrate that PLD2-mediated bleb formation is via the PA-LPAR-Rho-ROCK signalling pathway. Thus, PLD2 is a promising therapeutic target in combating metastasis of cancers of fibrous connective tissues.