O fator de crescimento epidermal (EGF) induz a sinalização de cálcio nuclear por meio da ação da fosfolipase C Delta-4 em SK-Hep-1

Abstract

Calcium signaling within the cell nucleus regulates specific cellular events such as gene transcription and cell proliferation. Nuclear and cytosolic calcium levels can be regulated independently and nuclear translocation of receptor tyrosine kinases (TRK) is one way to locally activate signaling cascades within the nucleus. Nuclear RTK, including epidermal growth factor receptor (EGFR), are important for processes such as transcriptional regulation, DNA-damage repair, and cancer therapy resistance. RTK can hydrolyze phosphatidylinositol-4,5-biphosphate (PIns-4,5-P2) within the nucleus, leading to calcium release from the nucleoplasmic reticulum by inositol-1,4,5-triphosphate receptors (InsP3). PIns-4,5-P2 hydrolysis is mediated by phospholipase C (PLC). However, it is unknown which nuclear PLC isoform is triggered by EGFR. Here, using subcellular fractionation, immunoblotting, fluorescence, siRNA and FRET-based biosensor reporter assay, we investigated the role of PLC delta 4 (PLCδ4) in epidermal growth factor (EGF)-induced nuclear calcium signaling and downstream events. We found that EGF-induced calcium signals are inhibited when nuclear translocation of EGFR is impaired. Nuclear calcium signaling also were reduced by selectively buffering inositol-1,4,5-triphosphate (InsP3) within the nucleus. EGF induced hydrolysis of nuclear Pins-4,5-P2 by the intranuclear PLCδ4, rather than PLC gamma 1 (PLCγ1). Moreover, protein kinase C (PKC), a downstream target of EGF, was active in the nucleus of stimulated cells. Furthermore, PLCδ4 and InsP3 modulated cell cycle progression by regulating the expression of cyclin A and B1. These results provide evidence that nuclear PLCδ4 mediates EGF-induced nuclear signaling and suggest new therapeutic targets to modulate proliferation effects of this growth factor.