The Epsilon Isoform of Protein Kinase C Is Involved in Regulation of the LTD4-Induced Calcium Signal in Human Intestinal Epithelial Cells

Abstract

We investigated the potential roles of specific isoforms of protein kinase C (PKC) in the regulation of leukotriene D4-induced Ca2+ signaling in the intestinal epithelial cell line Int 407. RT-PCR and Western blot analysis revealed that these cells express the PKC isoforms α, βII, δ, ϵ, ζ, and μ, but not βI, γ, η, or θ. The inflammatory mediator leukotriene D4 (LTD4) caused the TPA-sensitive PKC isoforms α, δ, and ϵ, but not βII, to rapidly translocate to a membrane-enriched fraction. The PKC inhibitor GF109203X at 30 μM but not 2 μM significantly impaired the LTD4-induced Ca2+ signal, indicating that the response involves a novel PKC isoform, such as δ or ϵ, but not α. LTD4-induced Ca2+ signaling was significantly suppressed in cells pretreated with TPA for 15 min and was abolished when the pretreatment was prolonged to 2 h. Immunoblot analysis revealed that the reduction in the LTD4-induced calcium signal coincided with a reduction in the cellular content of PKCϵ and, to a limited extent, PKCδ. LTD4-induced Ca2+ signaling was also markedly suppressed by microinjection of antibodies against PKCϵ but not PKCδ. These data suggest that PKCϵ plays a unique role in regulation of the LTD4-dependent Ca2+ signal in intestinal epithelial cells.