Open AccessDuck‐Haemoglobin Synthesis in Frog Cells
Author(s) -
Lane CharlesDaniel,
Gregory Caroline M.,
Morel Carlos
Publication year - 1973
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1973.tb02749.x
Subject(s) - reticulocyte , microinjection , globin , messenger rna , biology , xenopus , protein biosynthesis , rna , polysome , biochemistry , translation (biology) , oocyte , microbiology and biotechnology , ribosome , hemoglobin , embryo , gene
9‐S RNA was prepared from duck reticulocytes by dodecylsulfate treatment of the ribonucleoprotein particle derived from EDTA‐treated reticulocyte polyribosomes. The messenger content of this RNA was assayed by microinjection into oocytes of the frog Xenopus laevis. The 9‐S RNA was found to direct the synthesis of a molecular species that eluted from Sephadex G‐100 with added duck reticulocyte haemoglobin. When analysed on a cation‐exchange resin this material was found to contain the three globin‐chain species characteristic of duck reticulocyte haemoglobin. Moreover, paper electrophoresis at pH 6.5 and pH 3.5, and paper chromatography showed that these globin‐like species made in the oocyte were virtually indistinguishable, in terms of their histidine‐containing tryptic peptides, from the corresponding globin chains derived from duck reticulocyte haemoglobin. A comparison between the products made following the injection into oocytes of reticulocyte 9‐S RNA from rabbits, mice or ducks proved that it is the exogenous messenger that gives rise to the information content of the proteins whose synthesis is elicited by the injected RNA. The results also showed that the oocyte contains translational systems that are neither cell‐type nor species specific, and that duck globin messenger RNAs require no reticulocyte‐specific factors for their translation in oocytes. The construction of a saturation curve showed that there is a linear relationship between the amount of messenger injected and the amount of haemoglobin synthesized, up to the point at which haemoglobin synthesis constitutes over 35% of the total protein being made. Consequently the frog oocyte provides a convenient assay system for small quantities of duck globin mRNAs, and the messenger activity of as little as 1 ng of 9‐S RNA can be detected very simply. Thus a combination of the duck reticulocyte and frog oocyte may provide a useful system for studying post‐transcriptional control, and the formation of mRNA, in a eukaryotic cell.