Cytosolic phospholipase C activity: II. Relationship to concanavalin A-induced phosphatidylinositol-turnover in splenocytes
✍ Scribed by Thomas Akompong; Robert L. Spencer; Bruce S. McEwen
- Publisher
- John Wiley and Sons
- Year
- 1994
- Tongue
- English
- Weight
- 845 KB
- Volume
- 56
- Category
- Article
- ISSN
- 0730-2312
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✦ Synopsis
We have described in the first paper the coupling between cytosolic Gia and cytosolic PLC activity in a cell free preparation. In order to establish the functional significance of the cytosolic Cia coupled soluble PLC, we examined the effects of DEX, NaF, and trifluoperizine (TFP) on concanavalin A (Con A)-induced PI-turnover in intact splenocytes and, in parallel, on soluble PLC activity in cytosol preparations. Cytosolic PLC activity was measured with L3H1PI and [3HlPlP2 as substrates. 1) The Con A-induced increase (2-4 fold) in PI-turnover in intact splenocytes was paralleled by an 1.2-5-fold increase in soluble PLC activity in vitro. Con A administration also increased cytosolic Gia immunoreactivity 3-6-fold as expected if cytosolic Cia was coupled to soluble PLC activation. 2) DEX (10-7 M), administered 6 h prior to Con A administration, inhibited the Con A-induced increase in PI-turnover in intact splenocytes. This was paralleled by DEX inhibition of the Con A-induced increase in soluble PLC activity measured in vitro and cytosolic Cia immunoreactivity. 3) We have demonstrated in the first paper that NaF and TFP inhibited soluble PLC activity. Here we show that NaF and TFP inhibited the Con A-induced increase in PI-turnover extending the similarities between soluble PLC activity and Con A-stimulated PLC activity in intact splenocytes. 4) In order to examine whether or not the Con A-induced PLC was similar to PLCy, we measured PI-turnover induced by Con A or NaV0, in combination with DEX and PMA. Whereas the Con A-induced PI-turnover was significantly inhibited (40-60%) by DEX, the NaV03-induced PI-turnover was not affected by DEX. The Con A-induced PI-turnover was not affected by PMA (50 nM), but the NaV0,-induced PI-turnover was increased over 2-fold by PMA (50 nM), suggesting that the Con A-induced PLC in intact splenocytes is different from NaV03-induced PLC. Based on these results a model for the sequential activation of substrate-specific PLCs in splenocyte by mitogen is presented.