Characteristics of anion-stimulated Mg-ATPase from rat parotid gland secretory granules.

Magnesium-dependent adenosine triphosphatase (Mg-ATPase) was assayed in highly purified secretory granules. The enzyme was stimulated by sulphite and isethionate, unaffected by chloride and inhibited by fluoride and thiocyanate. Inhibition was not related to the permeant properties of the anion, but the relative inhibitory potency of the anions was similar to that in some other studies of secretory granule ATPases. Maximum contribution to the anion-stimulated ATPase by contaminating mitochondria was estimated at 9.3%. The enzyme was inhibited by the stilbene disulphonic acid inhibitor, 4-acetamido-4'-isothiocyano-2,2'-stilbene disulphonic acid (SITS). The IC50 was 0.16 mM in the absence of sulphite and increased in the presence of sulphite. The relation of the inhibition by SITS to sulphite was complex. Both Vmax and Km parameters were changed by SITS. Furthermore the data are consistent with the presence of two anion-stimulated ATPases. The ATPase was sensitive to tributyltin, dicyclohexylcarbodiimide (DCCD) and oligomycin, only moderately sensitive to azide, probenecid and N-ethylmaleimide (NEM) and rather insensitive to carbonylcyanide m-chlorophenylhydrazone (CCCP) and sulphisoxazole. ATPase activity was stimulated by calcium both in the presence and absence of magnesium. These findings suggest that the ATPase(s) present in parotid secretory granules is unique among secretory granule ATPases.

Calcium-dependent protein kinase reactions associated with parotid gland secretory granule membranes.

Rat parotid secretory granule membranes were examined for the presence of calcium-dependent protein kinase activities and kinase substrates. Protein kinase C (C-kinase), which is stimulated by certain phospholipids, was present in the membranes, as indicated by its ability to catalyze the phosphorylation of histone. Two substrates for protein kinase C were seen in the granule membranes. The cytosolic fraction from the cell contained kinase activity, which was stimulated by phosphatidylserine and which caused the phosphorylation of two granule membrane polypeptides. In addition, when both granule membranes and cytosol were incubated together, phosphorylation of the cytosolic substrates was inhibited, indicating that the granule membrane substrates were phosphorylated preferentially. The results indicate that the granule membranes may react with cytosolic protein kinase C activity in a way which would direct an intracellular calcium and diacylglycerol signal toward the granule membrane. Since these signals occur during stimulation by various agonists, the mechanism may contribute to secretion.

Graviresponsiveness and cap dimensions of primary and secondary roots of Ricinus communis (Euphorbiaceae).

After branching from the primary root, secondary roots of castor bean (Ricinus communis) grow laterally for 15-20 mm, after which they bend downward (i.e., become positively gravitropic). During the first 10 mm of growth, the lengths of caps of secondary roots increase from 120 +/- 26 to 220 +/- 28 micrometers. Although this increase is statistically significant (P < 0.1%), the resulting secondary roots are only minimally graviresponsive. A subsequent doubling of the lengths and widths of the root caps (i.e., to 420 +/- 34 and 450 +/- 41 micrometers, respectively) is positively correlated with the onset of gravicurvature. The graviresponsiveness and dimensions of caps of positively gravitropic secondary roots are not significantly different from those of positively gravitropic primary roots. These results indicate that (i) a statistically significant increase in the length and length : width ratio of a root cap does not necessarily result in the root becoming positively gravitropic, (ii) there may be a minimum cap length and (or) width necessary for graviresponsiveness, and (iii) the degree of graviresponsiveness exhibited by a particular root may be related to the size of its root cap.

Graviresponsiveness and the development of columella tissue in primary and lateral roots of Ricinus communis.

Half-tipped primary and lateral roots of Ricinus communis cv Hale bend toward the side of the root on which the intact half-tip remains. Therefore, the minimal graviresponsiveness of lateral roots is not due to the inability of their caps to produce growth effectors (presumably inhibitors). The columella tissues of primary (i.e. graviresponsive) roots are (a) 4.30 times longer, (b) 2.95 times wider, (c) 37.4 times more voluminous, and (d) composed of 17.2 times more cells than those of lateral roots. The onset of positive gravitropism by lateral roots is positively correlated with a (a) 2.99-fold increase in length, (b) 2.63-fold increase in width, (c) 20.7-fold increase in volume of their columella tissues. We propose that the minimal graviresponsiveness of lateral roots is due to the small size of their columella tissues, which results in their caps being unable to (a) establish a concentration gradient of the effector sufficient to induce gravicurvature and (b) produce as much as the effector as caps of graviresponsive roots.

Graviresponsiveness and columella cell structure in primary and secondary roots of Ricinus communis.

In order to determine what structural changes are associated with the onset of graviresponsiveness by plant roots, we have monitored the quantitative ultrastructures of columella (i.e., graviperceptive) cells in primary and secondary roots of Ricinus communis. The relative volumes of cellular components in lateral (i.e., minimally graviresponsive) roots were not significantly different from those of primary roots. The relative volumes of cellular components in secondary roots growing laterally were not significantly different from those of graviresponsive secondary roots. Therefore, the onset of graviresponsiveness by secondary roots of R. communis is not correlated with changes in organellar concentrations in columella cells. These results are discussed relative to a model for the differential graviresponsiveness of plant roots.

Graviresponsiveness and columella cell structure in primary and secondary roots of Ricinus communis.

In order to determine what structural changes are associated with the onset of graviresponsiveness by plant roots, we have monitored the quantitative ultrastructures of columella (i.e., graviperceptive) cells in primary and secondary roots of Ricinus communis. The relative volumes of cellular components in lateral (i.e., minimally graviresponsive) roots were not significantly different from those of primary roots. The relative volumes of cellular components in secondary roots growing laterally were not significantly different from those of graviresponsive secondary roots. Therefore, the onset of graviresponsiveness by secondary roots of R. communis is not correlated with changes in organellar concentrations in columella cells. These results are discussed relative to a model for the differential graviresponsiveness of plant roots.