Evidence for mRNA in Higher Plants

Central to the concepts of transcription and translation is the existence of mRNA. For this reason, we shall examine some of the few pieces of evidence for mRNA in higher plants and, in so doing, the knowledge which we have just acquired about antimetabolites will come in handy. 

Isolated sections from the hypocotyl of the soya bean continue to lengthen after they have been excised. If such hypocotyl sections are treated with 5-fluorouracil, the lengthening proceeds uninterrupted. If, however, actinomycin Ci is added to the hypocotyls, then the lengthening is inhibited. More detailed analyses of their nucleic acids showed what had happened in the hypocotyls under the influence of the antimetabolites. 

First a word concerning the procedure used. Columns are filled with methylalbumin-kieselgur (MAK) and a mixture of nucleic acids is applied to such a column. This mixture contains all of the nucleic acids to be found in the plant, namely, DNA, mRNA, rRNA, and tRNA. After the nucleic acid mixture has seeped into the column, the column is eluted with an NaCl solution of increasing molarity. The ease with which the nucleic acids, adsorbed to the methylalbumin, are eluted by NaCl differs. Some are eluted at quite low molarities of NaCl and so are rapidly displaced from the column. Others are eluted only at higher molarities of NaCl and, accordingly, are displaced later from the column. The nucleic acids are collected from the column in fractions. The amount of nucleic acids in each of the fractions is determined by measurements of the extinction at 260 nm (the extinction maximum of purines and pyrimidines) and, where necessary, of the radioactivity. 

If the nucleic acids from the hypocotyl of the soya bean are separated in this way, the following sequence of fractions is obtained (Fig. 17) tRNA, DNA, rRNA, and a further RNA fraction which is presumed to contain mRNA. If the nucleic acids of our sections are analyzed after treatment with 5-fluorouracil, it becomes apparent that the synthesis of tRNA and rRNA is strongly inhibited. In contrast, the synthesis of the supposed mRNA remains unimpaired. This is not the case after treatment with actinomycin Ci. In this instance,-as already mentioned, growth is inhibited, and the synthesis of the supposed mRNA is also strongly im- 

We will mention one more piece of evidence for the existence of mRNA, which will enable us, at the same time, to become acquainted with a new technique. This is the evidence for mRNA in the cotyledons of the ground nut (Arachis hypogaea). During germination of such cotyledons a considerable increase in the activity of a whole series of enzymes which are necessary for the mobilization of reserve materials, occurs. It seemed likely that this increase in activity was due, at least in a large part, to the de novo synthQSis of enzymes. However, such de novo synthesis is impossible without mRNA. The technique with which we are concerned here is the hybridization of nucleic adds. If a DNA double helix is slowly heated, a sharp increase in the extinction of 260 m t over a par- 

---