Localization of Adenyl Cyclase in Meristems of Young Pea Hypocotyls

Abstract

Electron micrographs of Pisum sativum L. hypocotyl tips treated to localize adenyl cyclase revealed discrete deposits on the internal membranes of cytoplasmic vacuoles which correspond to previously localized enzymes described as acid phosphatases. It remains to be determined whether the specificity of the substrate, adenylyl‐imidodiphosphate, used in the present study is such as to exclude all phosphatase activity other than adenyl cyclase. The acid phosphatase localized in earlier studies by other investigators may be an adenyl cyclase. In the differentiated cells of the root cap, lead precipitate was localized in distinct areas bound to the smooth endoplasmic reticulum.

Cyclic Nucleotides and Nucleotide Cyclases in Plant Stress Responses

“…This led to disputes that were only resolved in the late 1970s when experimental procedures to analyze cAMP were adapted to suit the plant cellular and extracellular environments. Since then a number of research groups reported more reproducible data, for example, the activity of PDE to hydrolyze 3',5'-cAMP through enzyme inhibition reaction by methylxanthines (Brown et al, 1977) and the localization of AC activity in meristems of young pea (Pisum sativum) hypocotyl (Hilton & Nesius, 1978). Efforts to identify endogenous cAMP in plant cells coincided with the development of sequential chromatographic and electrophoretic methodologies for the extraction and isolation of cAMP (Brown & Newton, 1973) that led to improved cAMP recovery by cochromatography.…”

“…Endogenously applied cAMP can be used as an indicator of functional activity for adenylate cyclase signaling system (Lomovatskaya et al, 2011). The presence and localization of AC activity was first experimentally demonstrated in meristems of young pea hypocotyls (Hilton & Nesius, 1978) making use of adenylyl-imidodiphosphate as a substrate specific for the enzyme and lead nitrate as a precipitating product that can be visualized with electron microscopy and quantitated in situ. Lead precipitate was found to be localized in distinct areas bound to the smooth endoplasmic reticulum in differentiated cells of the pea root cap (Hilton & Nesius, 1978).…”

“…The presence and localization of AC activity was first experimentally demonstrated in meristems of young pea hypocotyls (Hilton & Nesius, 1978) making use of adenylyl-imidodiphosphate as a substrate specific for the enzyme and lead nitrate as a precipitating product that can be visualized with electron microscopy and quantitated in situ. Lead precipitate was found to be localized in distinct areas bound to the smooth endoplasmic reticulum in differentiated cells of the pea root cap (Hilton & Nesius, 1978). Later, the use of mass spectrometric techniques explicitly identified the reaction product of AC activity in pea for the first time, thus erasing doubts casted on the methodology (Pacini et al, 1993).…”

Des variations d'activités de la 5′-nucléotidase et de l’adénylate-cyclase sont-elles des composantes de la levée d'inhibition du bourgeon cotylédonaire du pois?

“…Comme I'indique le schCma ci-dessous, I'adknylate-cyclase et la 5'-nuclCotidase jouent un r6le majeur dans la rCgulation du taux endogkne en nuclCotides et, en consCquence, dans la synthkse de polynuclCotides (Polya 1974) Howell ct Whitficld (1972) contcnant en concentration finale 20 mM dc tampon Tris-rnalCatc pH 7,4 avcc saccharosc i 8%; 2 mM de thCophyllinc inhibiteur (putatit] dc la destruction dc I'adCnosine monophosphate cycliquc (AMP-c) avcc transformation en 5'-AMP par la phosphodicstCrase (Duffus et Duffus 1968;Hilton ct Nesius 1978); 2 mM de sulfatc dc magnCsium; 0,5 mM de 5'-adCnylyl-imidodiphosphate; 20 mM de fluorurc dc sodium (NaF), activateur de I'enzyme dcs cellulcs anirnalcs (Rcik el 01. 1970; Rodbcll el a/.…”

Tansley Review No. 106

“…In the late 1970s and early 1980s, the experimental set-up for cAMP analysis was adapted to a great extent to cope with plant matrices. Taking care to minimize the artefactual origin of cAMP, various research groups produced more reproducible and coherent data (Brown et al, 1977 ;Katsumata et al, 1978 ;Hilton & Nesius, 1978 ;Tu, 1979). In the 1980s, chromatographic techniques in plant cAMP analysis such as adsorption, ion-exchange, paper and thin-layer chromatographies were largely reduced to preparative steps and replaced by the far superior HPLC, which is able to separate the 3h,5h-cyclic nucleotides and their 2h,3h isomers (Van Onckelen et al, 1982 ;Brown, 1983).…”