IN-VIVO EVIDENCE FOR THE INVOLVEMENT OF PHOSPHOLIPASE-A AND PROTEIN-KINASE IN THE SIGNAL-TRANSDUCTION PATHWAY FOR AUXIN-INDUCED CORN COLEOPTILE ELONGATION

Auxin-induced elongation of corn coleoptiles is accompanied by cell wa ll acidification, which depends upon Hf-pump activity. We tested the h ypothesis that phospholipase A and a protein kinase are involved in th e pathway of auxin signal transduction leading to H+ secretion, and el ongation of corn coleoptiles. Initially, the pH of the bath solution a t 50-100 mu m from the surface of a coleoptile segment (pH(o)) ranged between 4.8 and 6.6 when measured with an H+-sensitive microelectrode. Twenty or 50 mu M lysophosphatidylcholine, 50 mu M linolenic acid or 50 mu M arachidonic acid induced a decline in pH(o) by 0.3 to 2.1 unit s. The effect was blocked by 1 mM vanadate, suggesting that lysophosph atidylcholine or linolenic acid induced acidification of the apoplast by activating the H+-pump. Lysophosphatidylcholine and linolenic acid also accelerated the elongation rate of the coleoptiles. While linolen ic acid and arachidonic acid, highly unsaturated fatty acids, promoted pH, decrease and coleoptile elongation, linoleic acid, oleic acid, an d stearic acid, fatty acids with a lesser extent of unsaturation, had no such effects. The effects of lysophosphatidylcholine, linolenic aci d, and arachidonic acid on H+ secretion were not additive to that of i ndoleacetic acid (IAA), suggesting that lysophospholipids, fatty acids and auxin use similar pathways for the activation of the H+-pump. The phospholipase A(2) inhibitors, aristolochic acid and manoalide, inhib ited the IAA-induced pH, decrease and coleoptile elongation. The gener al protein kinase inhibitors, H-7 or staurosporine, blocked the IAA- o r lysophosphatidylcholine-induced decrease in pH(o). H-7 also inhibite d the coleoptile elongation induced by IAA or lysophosphatidylcholine. These results support the hypothesis that phospholipase A is activate d by auxin, and that the products of the enzyme, lysophospholipids and fatty acids, induce acidification of the apoplast by activating the H +-pump through a mechanism involving a protein kinase, which in turn p romotes corn coleoptile elongation.