Increase in the translatable mRNA for acetylcholine receptor during embryonic development of Torpedo ocellata electric organ

Abstract

The availability of large quantities of the nicotinic acetylcholine receptor (AChR) in the highly specialized electroplates of Torpedo permitted detailed studies of its structure [l-3], as well as preparation of specific antibodies against the receptor in its native and in its denatured forms [4-61. Anti-AChR antibodies have also been used to immunoprecipitate all 4 nascent chains of the newly synthesized AChR, translated by Torpedo mRNA in vitro [7,8] and in ovo, in microinjected Xenopus oocytes [9]. The immunoprecipitation of newly synthesized AChR subunits with anti-AChR antibodies of various specificities provides a means to determine which of the receptor antigenic sites already exists in the nascent chains. Here, we used antibodies raised against the native and denatured AChR and have demonstrated that newly synthesized AChR subunits probably do not possess the final spatial. conformation of the ‘adult’ receptor, but rather appear to be closer in structure to the denatured receptor. Differentiation of electroplates within the embryonic electric organ is accompanied by a steep increase in cholinergic constituents such as AChR and acetylcholinesterase (AChE) [IO]. Moreover, messenger RNA from the embryonic electric organ of Torpedo ocellata induces the production of catalytically active AChE in microinjected Xenopus oocytes [ 111. Following their maturation, the electroplates lose most of their ribosomes [ 121 and their capacity for protein synthesis is hence greatly reduced. Several mechanisms can explain the differentiationinduced accumulation of AChR in the electric organ. The elevation in AChR level might be preceded and/ or accompanied by a selective increase in its specific mRNA species (AChR mRNA). Alternatively, AChR production could be a transient phenomenon, characteristic of electroplate maturation stage. In both cases, the regulation of AChR expression might involve variation in the rate of proteolytic degradation of the receptor itself. In order to distinguish between these possibilities, we determined the level of AChR mRNA and compared it to the concentration of the receptor protein at 3 stages during electroplate differentiation: the electroblast stage (I); the stage when the celk acquire a columnar shape (II); and when morphological appearance of electroplates is displayed (III). Our findings indicate that the maturation stage of Torpedo ocellata electroplates is accompanied by a burst in AChR synthesis, and that following this stage, the mRNA species directing the synthesis of AChR represents a constant fraction out of total poly(A)containing mRNA.