PIP2 promotes membrane curvature and is a signaling hub in human sperm acrosome exocytosis.

ALTAMIRANO, KARINA N; SUHAIMAN LAILA; LUCCHESI, ORNELLA; RUETE, CELESTE; BELMONTE SILVIA A

Paraná, Entre Ríos

Congreso; 54th Annual Meeting Argentine Society for Biochemistry and Molecular Biology. SAIB; 2018

Argentine Society for Biochemistry and Molecular Biology. SAIB

The human sperm has a secretory vesicle that undergoes exocytosis when challenged with different stimuliknown as acrosome reaction (AR),which requires the fusion of the outer acrosome membrane and plasma membrane. We reported that DAG stimulates AR by feeding into a PKC and PLD1-dependent positive loop that supplies PIP2. We hypothesize that PIP2 synthesis is required to produce DAG and IP3, and to induce a change in the acrosomal membrane curvature. Our Molecular Dynamics simulations and TEM experiments demonstrated that PIP2 increase induces the formation of deep acrosomal membrane invaginations; although it was not able to induce acrosome swelling, AR or membrane disruption. However, for the AR to proceed, PIP2 needs to be hydrolyzed. Previous work led us to propose the following signaling pathway: cAMP-Epac-Rap1-PLCepsilon. The hydrolysis of PIP2 generates IP3, which binds IP3-sensitive channels and releases Ca2+ from theacrosome. We demonstrate the presence of a nucleotide exchange factor activated by DAG (RasGRP1), described to activate Rap1 in secretory cells, by WB and IFI. Also, we proved RasGRP1 ability to trigger AR in a dose-dependent manner. Furthermore, as shown by Far-IFI, DAG was able to activate Rap1 indicating that it is involved in the pathway described previously. Our findings highlights the dual role of PIP2 in exocytosis and report a direct evidence of the presence and function of RasGRP1 in the signaling module cAMP-Epac-Rap1-PLCε in AR.