Curr Genet (1995) 27:195-200 9 Springer-Verlag 1995 L. N. Mironova 9 M. G. Samsonova 9 G. A. Zhouravleva V. N. Kulikov. M. J. Soom Reversions to respiratory competence of omnipotent sup45 suppressor mutants may be caused by secondary sup45 mutations Received: t5 June 1994 Abstract The molecular nature of the sup45 respiratory deficient omnipotent suppressor, and of three reversions to respiratory competence which removed the suppressor ef- fect of the initial mutation, was examined. All reversions were caused by secondary sup45 mutations which indicates a direct connection between sup45 "respiratory" and "translational" functions. Computer analysis showed the local changes of Sup45 protein characteristics in the sup- pressor strain and revertants in comparison to the wild-type protein. The distribution of mutant sites in relation to ev- olutionary conserved, and tentatively functional, regions in the Sup45 protein is discussed. Key words Omnipotent suppression - Respiratory de- ficiency 9 Sup45 protein structure Introduction The Sup45 gene in the yeast Saccharomyces is one of the omnipotent (codon nonspecific) suppressor genes (see Inge-Vechtomov et al. 1994, for a review). It participates in the control of translational accuracy, but the exact func- tion of its product is unknown. In addition to translation the Sup45 protein is probably involved in other cellular processes (Pocklington et al. 1990). Proteins similar to Sup45 were found recently in human, Xenopus laevis and Arabidopsis thaliana cells (Grenett et al. 1992; Tassan et al. 1993; Quigley 1994). It was shown earlier (Ter-Avanesyan et al. 1982), that sup45 mutations inhibited mitochondrial translation. To study the relation of SUP45 functions in cytoplasmic and mitochondrial protein synthesis we have analysed muta- L. N. Mironova (~) 9 M. G. Samsonova. G. A. Zhouravleva V. N. Kulikov 9 M. J. Soom Department of Genetics, St. Petersburg St. University, 199034 St. Petersburg, Russia Communicated by L. Frontali tions which restore the respiratory capacity of sup45 mu- tants. This approach revealed the complicated system of interacting nuclear and mitochondrial genes which influ- ence cytoplasmic and mitochondrial protein synthesis (Mironova et al. 1986, 1988). For further investigation of SUP45 structure and func- tion we have now studied the molecular nature of an om- nipotent suppressor mutation, which also leads to respira- tory deficiency, as well as that of secondary SUP45 muta- tions selected as reversions to respiratory competence. We have analyzed the possible influence of suppressor muta- tions and reversions on the structure of the Sup45 protein and compared their intragenic distribution to the localiza- tion of mutations sequenced by other authors and to the position of tentative functionally important sites and evo- lutionary conserved regions in the Sup45 protein revealed by means of computer programs. Materials and methods Yeast strains and genetic methods. The list of strains employed is given in Table 1. Basic genetic procedures and media were as de- scribed previously (Sherman et al. 1988). Spontaneous or UV-in- duced (300 J/m2) revertants to respiratory competence were select- ed after incubation of respiratory deficient mutants for 7 days on glycerol media (YPGly). Molecular genetic procedures. Standard methods of recombinant DNA technics (Sambrook et al. 1989) were used. For amplification Table 1 Strains used in this study Strain Genotype 42B-P3990 MATaadel-14 his7-1 lys2-A12 3-42B-P3990 MATa adel-14 his7-1 lys2-A12 sup45-3 2G-P4364 MATaadel-14 his7-1 lys2-A12 ura3 leu2-1 sup45-3 5A-P4364 MATa adel-14 his7-1 ura3 Ieu2-1 thr4 sup45-3 6G-P4364 MATa adel-14 his7-1 thr4 sup45-3 1A-P4350 MATa his7-1 metl3-A1 thr4 arg4 1 B-P4350 MATa his7-1 ura3 arg4 thr4