Pseudomonas aeruginosa exotoxin A (PA), a potent protein synthesis inhibitor, was found to be a weak T-cell mitogen for murine splenocytes. Maximal stimulation of [3H]thymidine incorporation was obtained with 10 to 100 ng of toxin per ml following a 4-day induction. PA was also shown to be a polyclonal activator of cytolytic T lymphocytes (CTL), effective against concanavalin A-treated target cells. The effective PA dose for CTL induction was the same as that for mitogenic stimulation, only with a prolonged priming time (7 days). In contrast to other mitogens, PA could not reactivate memory CTL into secondary CTL. The stimulation of CTL by subcytotoxic doses of PA may be relevant to its modulatory effect on the immunocellular system.
EDTA dissociation of reticulocyte polyribosomes liberates globin mRNA as a nucleoprotein. An attempt to compare the proteins forming the mRNA * protein complexes with the initiation factors of protein synthesis is presented here.Two conditions are described under which initiation factors are removed from hemoglobinsynthesizing ribosomes. One is after treating the polyribosomes with a high salt concentration ; the other is by isolating ribosomes from reticulocytes at an advanced state of maturation which have lost their initiation factors. I n both cases it is shown that the mRNA remains associated with the ribosomes and with the proteins forming the mRNA * protein.Moreover both mRNA and mRNA -protein are translatable in a heterologous cell-free system and their translation is stimulated to the same extent by the addition of reticulocyte initiation factors.Initiation factors are therefore different physically and functionally from the proteins of the mRNA -protein. and little information has been provided regarding the biological significance of these mRNA * protein associations. Some progress towards understanding the functional role of mRNA-protein has been made in the rabbit reticulocyte system where the 15-S mRNA ' protein (in which form globin messenger RNA is released from the polyribosomes by EDTA treatment) can be isolated in a nearly homogeneous form. The proteins attached to globin mRNA are well defined and a t least one of them appears to be required for binding mRNA to native 40-5 ribosomal subunits washed with deoxycholate [4].It was therefore of interest to study the possible relationship between these mRNA * protein proteins and the factors isolated by concentrated-salt washing of ribosomes which have been shown to be required for initiation and to be specific for mRNA recogniAbbreviations. mRNA * protein, messenger ribonucleoprotein; Brij 35, polyoxyethylene cetylether.Definition. Asso unit, the quantity of material contained in 1 ml of a solution which has an absorbance of I at 260 nm, when measured in a 1-cm path-length cell.tion [6-81. However, a direct comparison has been impossible to perform since initiation factors have not yet been p d e d to homogeneity. I n order to compare these seemingly separate classes of proteins we have used two types of ribosomes which are both dependent on the addition of initiation factors in order to synthesize globin chains. These are ribosomes washed in 0.5 M KC1 [6,7] and ribosomes taken from a mature population of reticulocytes [Y].We have shown that in both cases the major part of the globin mRNA is still attached to the remaining population of monomers and subunits, and that their lack of activity is primarily due to the loss of their initiation factors. Moreover, the mRNA remains attached to the ribosomes in the form of a nucleoprotein complex apparently identical to the mRNA * protein isolated from normal polyribosomes. Removal of initiation factors does not therefore release the mRNA -protein complexes. Studies of the translation of mRNA and mRNA.prote...
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