No abstract

In the present study we have determinated the acetylcholinesterase molecular forms present in rat liver hepatocytes; we have also studied the association of acetylcholinesterase with the cell surface of the hepatocytes. Subcellular fractionation indicated that rough endoplasmic reticulum and plasma-membrane-enriched fractions contains G4 and G1 acetylcholinesterase forms bound to membranes. Hepatocytes incubated with phosphatidylinositol-specific phospholipase C released about 70% of the surface acetylcholinesterase. Sedimentation analysis showed that all the solubilized acetylcholinesterase activity comes exclusively from a G2 dimer. The G4 hydrophobic form of acetylcholinesterase accounts for the additional cell-surface activity. The existence of these two forms of acetylcholinesterase on the surface of hepatocytes was further established by analyzing the phosphatidylinositol-specific phospholipase C sensitivity of the acetylcholinesterase molecular forms present in isolated rat liver plasma membranes.Acetylcholinesterase is the enzyme responsible of the hydrolysis of the neurotransmitter acetylcholine [l, 21. The enzyme exists in multiple molecular forms which can be separated by velocity sedimentation. The acetylcholinesterase molecular forms can be classified as globular (monomers GI, dimers G1 and tetramers G4) and asymmetric forms (containing one, two or three tetramers plus a collagen-like tail: A4, As and AI2) [I -41. Asymmetric forms of acetylcholinesterase are associated with the extracellular matrix by heparan sulfate proteoglycans [5,6]. Globular acetylcholinesterases have been identified as both soluble and membrane-bound forms and some of them appear to have hydrophobic domains that attach the enzyme to the plasma membrane [3, 71. The G2 forms of acetylcholinesterase are apparently anchored to the membrane through a glycolipid moiety which contains phosphatidylinositol (PtdIns) covalently linked to the C-terminal [8 -1 I]; the G4 acetylcholinesterase form contains a non-catalytic hydrophobic 20-kDa polypeptide subunit [12]. Futerman et al. [I31 have communicated that a G2 dimer of acetylcholinesterase was present in rat liver homogenates. Here we show that isolated hepatocytes and plasma membrane fractions contain both GZ and G4 forms of acetylcholinesterase. In particular, the dimer was mainly localized in the cell surface from where it can be entirely solubilized by treatment with PtdIns-specific phospholipase C.Correspondence to E. Brandan, Molecular Neurobiology Unit,

No abstract

No abstract

With the aim of understanding the organization of the nervous system in the Prosobranchia gastropod Concholepas concholepas, we studied the properties, specificity, sedimentation coefficient, and solubility of the cholinergic enzyme, acetylcholinesterase (AChE). It was found that 95% of the esterase was inhibited by BW284c51 dibromide but not by iso-OMPA, which is consistent with the specificity of AChE. The calculated K, 0.22 mM is eight to ten times higher than are the K,s for AChE of other invertebrates and similar to the values reported for fishes and vertebrates.The AChE shows a maximal activity around 22"C, has a glycoproteic character, and presents sedimentation coefficients of 6.5 S and 10.5 S. Most of this AChE activity is soluble under low ionic strength conditions; however, the enzyme aggregates in the absence of detergents.In conclusion, our evidence indicates the presence of a well-recognized molecular marker that could be useful for the study of the development of Concholepas concholepas.Acetylcholinesterase (AChE, EC 3.1.1.7) catalyzes the hydrolysis of the neurotransmitter acetylcholine at cholinergic synapses, thereby participating in the regulation of nerve transmission in the peripheral and central nervous system (Kuffler et al., '84). AChE exists in various molecular forms that can be distinguished by their solubility and hydrodynamic properties (Massoulie and Bon, '82). Asymmetric forms contain a collagen-like tail that anchors the enzyme to the synaptic extracellular matrix by interactions with heparan sulfate proteoglycans (Inestrosa et al., '82; Brandan et al., '85). Globular forms correspond to monomers (GJ, dimers (Gz), and tetramers (G4) of catalytic subunits (Inestrosa and Perelman, '89). The globular forms can be hydrophobically anchored to membranes or can occur as "soluble" forms that are released from them by low ionic strength solution in the absence of detergent (Silman and Futerman, '87). Hydrophobic dimers have been studied in detail and appear to have a glycophospholipid as a plasma membrane anchoring domain (Silman and Futerman, '87; Inestrosa et al., '88; Perelman and Brandan, '89). In contrast, tetrameric globular forms, Although in the animal kingdom the distribution of AChE determines important phylogenetic relationships, in invertebrates the enzyme has been poorly characterized, especially in marine organisms. The gastropod muricid Concholepas concholepas is the only species of the genus and is a very important marine resource in the Southeast Pacific (Castilla, '83; Castilla and Jerez, '86). However, despite decades of research, very little is known about the cellular and molecular biology of this organism.As a first step in the study of the nervous system of the C . concholepas, we describe here the This is the first paper of a series entitled "Molecular and Cellular Biology of Concholepas concholepas."