Phosphorylation of mitochondrial phospholipid scramblase 3 by protein kinase C-δ induces its activation and facilitates mitochondrial targeting of tBid
✍ Scribed by Yongwen He; Jihua Liu; Douglas Grossman; David Durrant; Trevor Sweatman; Leonard Lothstein; Raquel F. Epand; Richard M. Epand; Ray M. Lee
- Publisher
- John Wiley and Sons
- Year
- 2007
- Tongue
- English
- Weight
- 315 KB
- Volume
- 101
- Category
- Article
- ISSN
- 0730-2312
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✦ Synopsis
Abstract
Phospholipid scramblase 3 (PLS3) is a member of the phospholipid scramblase family present in mitochondria. PLS3 plays an important role in regulation of mitochondrial morphology, respiratory function, and apoptotic responses. PLS3 is phosphorylated by PKC‐δ at Thr21 and is the mitochondrial target of PKC‐δ‐induced apoptosis. Cells with overexpression of PLS3, but not the phosphoinhibitory mutant PLS3(T21A), are more susceptible to apoptosis induced by AD198, an extranuclear targeted anthracycline that activates PKC‐δ. Here we report that the phosphomimetic mutant of PLS3(T21D) by itself can induce apoptosis in HeLa cells. Using proteoliposomes with addition of pyrene‐labeled phosphatidylcholine (PC) at the outer leaflet, we measured the lipid flip‐flop activity of PLS3 and its phosphorylation mutant. PLS3(T21D) is more potent than wild‐type PLS3 or PLS3(T21A) to transfer pyrene‐PC from the outer leaflet to the inner leaflet of liposomes. Based on our previous finding that PLS3 enhances tBid‐induced mitochondrial damages, we tested the hypothesis that PLS3 enhances cardiolipin translocation to mitochondrial surface and facilitates tBid targeting. Fluorescein‐labeled tBid(G94E) was used as a probe to quantify cardiolipin on the surface of mitochondria. Mitochondria from cells treated with AD198 or cells expressing PLS3(T21D) had a higher level of tBid‐binding capacity than control cells or cells expressing wild‐type PLS3. These findings indicate that phosphorylation of PLS3 by PKC‐δ induces PLS3 activation to facilitate mitochondrial targeting of tBid and apoptosis. J. Cell. Biochem. 101:1210–1221, 2007. © 2007 Wiley‐Liss, Inc.