Dual Activation of Phospholipase C-ϵ by Rho and Ras GTPases

Department of Pharmacology, University of North Carolina School of Medicine, Chapel Hill, North Carolina 27599, USA.
Journal of Biological Chemistry (Impact Factor: 4.57). 10/2008; 283(44):29690-8. DOI: 10.1074/jbc.M805038200
Source: PubMed

ABSTRACT Phospholipase C-epsilon (PLC-epsilon) is a highly elaborated PLC required for a diverse set of signaling pathways. Here we use a combination of cellular assays and studies with purified proteins to show that activated RhoA and Ras isoforms directly engage distinct regions of PLC-epsilon to stimulate its phospholipase activity. Purified PLC-epsilon was activated in a guanine nucleotide- and concentration-dependent fashion by purified lipidated K-Ras reconstituted in PtdIns(4,5)P(2)-containing phospholipid vesicles. Whereas mutation of two critical lysine residues within the second Ras-association domain of PLC-epsilon prevented K-Ras-dependent activation of the purified enzyme, guanine nucleotide-dependent activation by RhoA was retained. Deletion of a loop unique to PLC-epsilon eliminated its activation by RhoA but not H-Ras. In contrast, removal of the autoinhibitory X/Y-linker region of the catalytic core of PLC-epsilon markedly activates the enzyme (Hicks, S. N., Jezyk, M. R., Gershburg, S., Seifert, J. P., Harden, T. K., and Sondek, J. (2008) Mol. Cell, 31, 383-394), but PLC-epsilon lacking this regulatory region retained activation by both Rho and Ras GTPases. Additive activation of PLC-epsilon by RhoA and K- or H-Ras was observed in intact cell studies, and this additivity was recapitulated in experiments in which activation of purified PLC-epsilon was quantified with PtdIns(4,5)P(2)-containing phospholipid vesicles reconstituted with purified, isoprenylated GTPases. A maximally effective concentration of activated RhoA also increased the sensitivity of purified PLC-epsilon to activation by K-Ras. These results indicate that PLC-epsilon can be directly and concomitantly activated by both RhoA and individual Ras GTPases resulting in diverse upstream control of signaling cascades downstream of PLC-epsilon.

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    • "In the present study, PI-PLC β4 and PI-PLC δ3 were expressed. PI-PLC ε known to be a downstream effector of Ras superfamily GTPases and an upstream activator of small GTPases, both Ras and Rap (Seifert et al. 2008), was weakly expressed. The expression of PI-PLC β1 is significantly higher. "
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    Journal of Cell Communication and Signaling 02/2013; 8(3). DOI:10.1007/s12079-013-0194-6
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    • "Although not yet verified in conditions satisfying the four criteria, data from the literature may suggest interesting possibilities. Indeed, RhoA and Rac1 activate PLC isoforms, including the PLCβ, PLCΓ, and PLCε (Thodeti et al., 2002; Illenberger et al., 2003; Wing et al., 2003; Piechulek et al., 2005; Seifert et al., 2008; Walliser et al., 2008; Guo et al., 2010) which, in turn, could determine the activation of Rap1A through CALDAG–GEF. Notably, the lipid kinase activity of PIP5K1C itself may contribute to Rap1A activation by increasing the local concentration of PtdIns(4,5)P2, which, in turn, is substrate of PLCs leading to activation of CALDAG–GEF and, ultimately, of Rap1A. "
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    Frontiers in Immunology 05/2012; 3:127. DOI:10.3389/fimmu.2012.00127
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    • "Activaton of Gα12/13 leads to RhoA and phospholipase C-ε activation [34], [47]. Phospholipase C-ε is known to activate Ras and PKC [48], [49]. Besides, some activation mechanisms of RhoA by Ras had been reported [50], [51]. "
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