ACRIDINE-ORANGE AS AN INDICATOR OF THE CYTOPLASMIC RIBOSOME STATE

In hippocampal neurons of ground squirrels and neuroblastoma culture t he ribosome state was analyzed by staining with acridine orange (AO), labeling with radioactive amino acids, and electron microscopy, Electr on microscopy indicated that the extent to which ribosomes associated in polysomes varied from 25% in brain cells of torpid ground squirrels up to 93% in growing neuroblastoma cells, In control rat neurons, it was 75%. The fluorescence of the AO-rRNA complex in ribosomes changed with the polysome/monosome ratio, The red fluorescence of AO-single-st randed rRNA complex as well as ribosome association in polysomes decre ased greatly as a function of the shift from polysomes to monosomes, T he green fluorescence of AO intercalated in the double-stranded rRNA c hanged insignificantly As a result, the ratio of red to green fluoresc ence intensity, K-alpha, changed more than 3-fold with the changing po lysome/monosome ratio, Protein labeling also showed strong positive co rrelation with K-alpha. Thus, rRNA showed different accessibility for AO binding in active and inactive ribosomes, A possible mechanism of p artial rRNA shielding with proteins is proposed. AO fluorescence in th e cytoplasm, i.e., AO binding to rRNA in ribosomes, is presumed to ref lect adequately the profound changes in the state of cell protein synt hesizing system that can be regulated by both functional activity acid stress factors. (C) 1997 Wiley-Liss, Inc.