Immunological Characterization of DNA-Dependent RNA Polymerase(s) of Spinach Chloroplasts

Abstract

Chloroplasts from spinach are easily isolated in an intact form, free of cytoplasmic and nuclear contamination. The chloroplast genomes of tobacco and Marchantia have been sequenced (Shinozaki et al. 1986, Ohyama et al. 1986). Sequences homologous to the genes coding for the β, β’ and α subunits of RNA polymerase of E. coli have been found. Two apparently independent transcriptional activities have been isolated. One, the soluble RNA poly-merase (sRNAP), is present within the chloroplast stroma. The second, the transcriptionally active DNA-protein complex (TAC), can be isolated from the thylakoid membrane fraction. The TAC contains chloroplast DNA, tightly bound chloroplast DNA-dependent RNA polymerase, and other proteins. According to Hallick and his co-workers the transcriptionally active chromosome of Euglena specifically transcribes the rRNA operon (Rushlow et al. 1980, Gruissem et al. 1983a, Greenberg et al. 1984, Narita et al. 1985), whereas the sRNAP is selective for transcription of tRNA genes from spinach (Gruissem et al. 1983b, Gruissem, 1984) and Euglena (Gruissem et al. 1983a, Greenberg et al. 1984). However, mustard chloroplast TAC is capable of transcribing plastid rRNA genes as well as protein coding genes (Reiss and Link, 1985).