ABSTRACT
Our previous behavioural, biochemical and immunohistochemical analyses conducted in selected regions (supra/sub oesophageal masses) of the Octopus vulgaris brain detected a cytoplasmic poly-ADP-ribose polymerase (more than 90% of total enzyme activity). The protein was identified as the vault-free form of vault-poly-ADP-ribose polymerase. The present research extends and integrates the biochemical characterization of poly-ADP-ribosylation system, namely, reaction product, i.e., poly-ADP-ribose, and acceptor proteins, in the O. vulgaris brain. Immunochemical analyses evidenced that the sole poly-ADP-ribose acceptor was the octopus cytoskeleton 50-kDa actin. It was present in both free, endogenously poly-ADP-ribosylated form (70kDa) and in complex with V-poly-ADP-ribose polymerase and poly-ADP-ribose (260kDa). The components of this complex, alkali and high salt sensitive, were purified and characterized. The kind and the length of poly-ADP-ribose corresponded to linear chains of 30-35 ADP-ribose units, in accordance with the features of the polymer synthesized by the known vault-poly-ADP-ribose polymerase. In vitro experiments showed that V-poly-ADP-ribose polymerase activity of brain cytoplasmic fraction containing endogenous actin increased upon the addition of commercial actin and was highly reduced by ATP. Anti-actin immunoblot of the mixture in the presence and absence of ATP showed that the poly-ADP-ribosylation of octopus actin is a dynamic process balanced by the ATP-dependent polymerization of the cytoskeleton protein, a fundamental mechanism for synaptic plasticity.
Subject(s)
Actins/metabolism , Brain/enzymology , Octopodiformes/enzymology , Poly(ADP-ribose) Polymerases/metabolism , Vault Ribonucleoprotein Particles/metabolism , Actin Cytoskeleton/metabolism , Actins/chemistry , Animals , Neuronal Plasticity/genetics , Poly Adenosine Diphosphate Ribose/metabolism , Poly(ADP-ribose) Polymerases/chemistry , Polymerization , Synapses/metabolism , Synapses/physiology , Vault Ribonucleoprotein Particles/chemistryABSTRACT
In plants, the decline of poly(ADP-ribosyl)ation activity is involved in energy homeostasis and stress tolerance. By reducing stress-induced poly(ADP-ribosyl)ation activity, NAD(+) breakdown is inhibited preventing high energy consumption. Under these conditions, plants preserve their energy homeostasis without an overactivation of mitochondrial respiration, thus avoiding the production of reactive oxygen species. Therefore, plants with lowered poly(ADP-ribosyl)ation activity appear tolerant to multiple stresses. In this study, the evergreen species Cistus incanus L. was used as a model because of its capacity to overcome successfully the environmental constraints of the Mediterranean climate. The aim of the present work was to characterize and assess the role of poly(ADP-ribosyl)ation in C. incanus plants kept under different temperature in greenhouse (GH), outdoor during winter (WO) and outdoor during spring (SO). Data showed that in C. incanus polyADPribose metabolism occurs. The enzyme responsible for poly(ADP-ribose) chains synthesis is a poly(ADP-ribose)polymerase of about 80 kDa, lacking "zinc finger" N-terminal domain and able to automodify. The lowest PARP activity, as well as the lowest quantum yield of PSII linear electron transport (Φ(PSII)) and photochemical quenching (q(P)), was found in WO plants. Instead, in SO plants the recovery of photochemical activity associated to a poly(ADP-ribose)polymerase activity increase of about 50%, as compared to GH plants, was observed. Taking into account both biochemical and eco-physiological responses, a possible explanation for the poly(ADP-ribosyl)ation deficiency in WO plants has been hypothesized.
