The effect of oleate and spermine on the subcellular distribution of phosphatidate phosphohydrolase (PAH, EC 3.1.34).

Phospha t i d a t e p h o s p h o h y d r o l a s r (PAH) c a t a l y s e s t h e c o n v e r s i o n o f phospha t i d a t e t o d i a c y l g l y c r r o l and is a n i m p o r i a n t r e g u l a t o r y s t e p i n t r i a c y l g l y r e r o l synthesis. Long t e r m c o n t r o l o f enzyme a c t i v i t y is m e d i a t r d by hormones r i t h e r v i a i n c r r a s e d r n z y m c s y n t h r s i s or s t a b i l i t y (1.2). I n t h e s h o r t t c r m , t h e p h y s i o l o g i c a l e x p r e s s i o n of PAH a c t i v i t y I S t t lmigti t t o b e dc,pcndant on t h r t r a n s l o c a t i o n o f PAH from t h e r c l a t i v c l y i n a c t i v e c y t o s o l i c poo l 1.0 t h e mic rosomal membranes where i t s s u b s t r a t e is s y n t h e s i s e d and i t s a c t i v i t y may h e e x p r c s s c d ( 1 ,2). T h i s translocat i o n o f a c t i v i t y c a n be i n d u c e d by f a t t y a c i d s or t h e i r CoA e s t e r s and p o l y a m i n e s and c a n b e r e v e r s c d by c-111 o rp rom, rz i rie . t ' r r v i oils s t u d i e s r i se~d i n c a b a t ion of c u 1 t u r t . d t i c p a t o c y t c s ( 3 ) o r p o s t m i t o c h o n d r i a 1 s u p e r n a t a n t s ( 4 ) . T h i s st .udy w a s des ig r l cd t o i n v e s t i g a t e t h e cha1igt.s on t h e d i s t r i b u t i o n o f PAH a f t e r i n c u b a t i o n o f rat l i v e r t o t a l homogenate w i t h o l e a t e or s p e r m i n e . Ct-1 lu la r f r a c t . i o n a t i o n was u n d e r t a k e n u s i n g c o n t i n u o u s surrose g r a d i e n t s , c e n t r i f u g e d i n a v e r i i c n l p o c k e t r o t o r . PA11 a c t i v i t y ( 5 ) a n d o t h e r m a r k e r enzymes wc'rc assayed ( 6 ) and r e s u l t s c a l c u l a t e d as f r e q u e n c y / d e n s i t y . I n u n t r e a t e d homogena te PAH a c t i v i t y r e m a i n e d c y t o s o l i c ( F i g . ] 1 , t r e a t m e n t w l t h o l f a t e o r spc r rn ine s h i f t s a c t i v i t y on t o h i g h e r d e n s i t y of t h e g r a d i e n t . 'The e f f e c t o f t r e a t m e n t w i t h o l e a t c and spe rmi r i e is a d d i t i v e ( F i g . 2 ) . Thc. peak d i s t r i b u t i o n of I'AH