Phosphoinositide Kinases in Chick Brain and Sciatic Nerve, a Developmental Study

Abstract

Abstract— Phosphatidylinositol kinase and diphosphoinositide kinase activities were measured in homogenates of brain and sciatic nerve of developing chick embryos and chicks. Characteristics of the chick nervous system enzymes were similar to those reported for rat brain. Diphosphoinositide kinase was inhibited by high concentrations of ATP and by low concentrations of triphosphoinositide. Both activities were greatly enhanced by the non‐ionic detergent, Cutscum, and the ratio of detergent to protein in the reaction mixture was important. Optimum phosphatidylinositol kinase activity required a ratio of 7 : 1 for both tissues. The optimum ratio for diphosphoinositide kinase was 3:1 for nerve homogenates and 0.6:1 for brain. Cutscum increased the concentration of diphosphoinositide that is required for maximum diphosphoinositide kinase activity. Developmental changes were the same for both kinase activities, which were low in unmyelinated brain and sciatic nerve. The activities correlated with the concentration of polyphosphoinositides in chick brain where they increased 4‐5 fold during the period of active myelination and remained high in the mature brain. The kinase activities correlated with the rate of triphosphoinositide deposition in sciatic nerve. Following a 2‐3 fold increase during the initial phase of myelination the activities declined to values as low as those of embryonic nerve.

Catalytic properties of a purified phosphatidylinositol-4-phosphate kinase from rat brain

“…To our knowledge, PIP kinase has not yet been obtained in highly purified form (Cooper & Hawthorne, 1976;Shaikh & Palmer, 1977;Desmukh et al, 1984), although work with partially purified preparations has resulted in the proposal that the enzyme may be the target of a regulation in the intact cell (Jolles et al, 1980;Van Dongen et al, 1985). Gispen and co-workers (Jolles et al, 1980;Van Dongen et al, 1985) have described a brain PIP kinase preparation which co-purified as a complex with a 50 kDa phosphoprotein (B-50 protein), and a series of reports from his group resulted in the proposal that B-50 could be a modulator ofPIP kinase through a shuttle between a phosphorylated (inhibitory) and a dephosphorylated form (Van Dongen et al, 1985).…”

Phosphatidylinositol 4,5-bisphosphate hydrolysis in human sperm stimulated with follicular fluid or progesterone is dependent upon Ca2+ influx

“…The sperm were finally suspended to a concentration of 6 x 106/ml in Medium A (Med. A), containing 117.5 mM-NaCl, 25 containing 125 mM-NaCl, 10 mM-KCl, 2.5 mM-CaCI2, 0.25 mM-MgCl2, 19 mM-sodium lactate, 2.5 mM-sodium pyruvate, 20 mM-Hepes/NaOH and 3 mg of BSA/ml, which was supplemented with 10 mM-Ins. The cells were sedimented at 300 g for 10 min and the pellet was resuspended in 50 ml of Med.…”

“…The lipids were then separated on silica gel H plates impregnated with 1 % potassium oxalate using a solvent system ofchloroform/acetone/methanol/acetic acid/water (40:15:13:12:8, by vol.) [25]. Chromatograms were stained with iodine and the appropriate spots scraped from the plates.…”

Separation and Characterization of Inositol Phospholipids from the Pulvini of Samanea saman

“…The limits of detection of the assay are 0.5 to 1 nmol. The total lipid phosphate detected has averaged 1489 nmol in extracts of approximately 24 pulvini (n = 1 1). Thus there can be no more than 1 nmol phosphate in PIP2 per 1489 nmol of total lipid phosphate.…”