Studies on the Control of Development

Abstract

Ribosomes from vegetative cells (vegetative ribosomes) of Bacillus subtilis synthesize guanosine 3′(2′)‐diphosphate 5′‐diphosphate (p2Gp2) and guanosine 3′(2′)‐diphosphate 5′‐triphosphate (p3Gp2). In contrast, ribosomes isolated from sporulating cells (sporulation ribosomes) synthesize adenosine 3′(2′)‐diphosphate 5′‐diphosphate (p2Ap2) and adenosine 3′(2′)‐diphosphate 5′‐triphosphate (p3Ap2). The products obtained with the aid of vegetative ribosomes were characterized by comparison with authentic substances in two chromatographic systems. The reaction mechanism is identical with that found in Escherichia coli, since [3H]GTP or GDP is pyrophosphorylated by ATP to form p2Gp2 and p3Gp2. Omission of ATP completely stops the formation of any reaction products. Furthermore, no reaction products are observed when GTP or GDP are omitted in a reaction mixture containing vegetative ribosomes. In contrast, incubation of sporulation ribosomes with ATP alone or ADP and ATP results in the formation of two products identified as p2Ap2 and p3Ap2 by their mechanism of synthesis as well as chemical and enzymatic degradation. Sporulation ribosomes (isolated at least an hour after the end of logarithmic growth) synthesize drastically less (by a factor of 100) p2Gp2 and p3Gp2. Vegetative ribosomes, on the other hand, synthesize only 1/100 the amount of p2Ap2 and p3Ap2 compared to sporulation ribosomes when isolated at least an hour before the end of logarithmic growth. By using washed ribosomes and ribosomal wash from vegetative and sporulation ribosomes, we were able to show that changes in the composition of ribosomes (probably in ribosomal proteins and/or ribosome‐associated factors) must occur during the transition from vegetative to sporulating cells. Washed ribosomes from vegetative cells cannot replace washed ribosomes from sporulating cells to synthesize p2Ap2 and p3Ap2 in the presence of ribosomal wash from sporulating cells and vice versa. During the transition from one type of ribosomes to the other, as far as the unusual nucleotide‐synthesizing activity is concerned, both types of nucleotides can be synthesized. While the ability to synthesize p2Gp2 and p3Gp2 decreases approximately at the end of logarithmic growth, the ability to synthesize p2Ap2 and p3Ap2 increases at the same time. The reason behind this obvious change in ribosomal structure and associated factors is discussed in the light of our finding that these changes are not observed in some early blocked asporogenous mutants.The observation that ribosomes from sporulating cells synthesize traces of p2Gp2 and p3Gp2 (by a factor of almost 100 less than vegetative ribosomes) is probably due to the presence of some non‐sporulating cells in any B. subtilis culture. Likewise, there are always some spores contained in a vegetatively growing culture, which are probably responsible for the observed small amounts of p2Ap2 and p3Ap2 synthesized.