A DNA polymerase has been extracted from spinach chloroplasts and purified by chromatography on DEAE-cellulose and hydroxyapatite. A great similarity between the purified chloroplast polymerase and the mammalian mitochondrial DNA polymerase y was found by several criteria :preference for the synthetic primer-template (dT)lz -18 . poly(rA), optimal requirement for Mn2+ (0.1 -1.0 mM), KCl(lO0 mM) and pH (8 -9), high relative molecular mass (approximately 105 000), resistance to aphidicolin and inhibition by N-ethylmaleimide. Some peculiar features of the chloroplast DNA polymerase have, however, been noticed.The mammalian DNA polymerase y has been suggested to be responsible for the replication of mitochondrial DNA. Thus, both the presence of a y-like DNA polymerase in chloroplasts and the similarities between the chloroplast and the mitochondrial DNA (absence of a nucleosomal structure and presence of displacement loops) lead to the suggestion that chloroplast DNA is also replicated by a y-like DNA polymerase and that the y polymerases present in eukaryotes are, therefore, involved in a strand-displacement DNA synthesis.An a-like DNA polymerase activity, present and predominant in crude leaf extracts, was practically absent from purified chloroplast preparations.Developed chloroplasts contain multiple copies (up to 50) of possibly identical supertwisted circular DNA molecules, one circle thus constituting the entire chloroplast genome [l-31. ctDNA resembles mtDNA and bacterial DNA in several aspects, among them the absence of supromolecular organization in chromatin structures, and is thus substantially different from the chromosomal DNA present in nuclei [2,4,5].Recent data indicate that animal mitochondria contain a unique DNA polymerase (DNA polymerase y) [6 -81, which is responsible for the replication of mtDNA [9,10]. On the other hand, no information is available on the enzyme(s) involved in the replication of ctDNA [2,5]. Chloroplasts isolated from tobacco [ l l ] and spinach [12] are capable of incorporating deoxynucleotides into chloroplast DNA. However, this activity was not purified and characterized.In this work we show that spinach leaves contain at least two DNA polymerases and that only one of them is localized in the chloroplast. The properties of the partially purified chloroplast enzyme are similar in many aspects to those of DNA polymerase y, the replicative enzyme of animal mtDNA. Some peculiarities have, however, been noticed. MATERIALS AND METHODS MaterialsN-Ethylmaleimide was from Sigma Chemicals Corp. ; p-toluenesulfonyl fluoride (TosF) was from Aldrich; poly(rA), poly(dA), poly(rC) and poly(dC) from Miles Laboratories; (dT)1Z -18 and (dG)12 -18 were from PL-Biochemicals ; [3H]dTTP was from Amersham; Teepol 610 was from BDH Chemicals; D-mannitol was from Carlo Erba; sucrose was from Merck; DEAE-cellulose (DE-52) and phosphocellulose (Pll) were from Whatman; hydroxyapatite was prepared according to Bernardi [13].Ahbreviutions. ctDNA, chloroplast DNA; mtDNA, mitochondrial DNA; TosF, p-tolue...

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A DNA polymerase activity was isolated from cells of Oryza sativa L. grown in suspension culture. Molecular mass (∼ 180,000), optimal requirements for pH (neutral), Mg(2+) (5-10 mM), Mn(2+) (1 mM), template preference (activated DNA), lack of activity with native or denatured DNA, and sensitivity to N-ethylmaleimide and ionic strength are similar to those of the vertebrate α-polymerase. Like DNA polymerase α, the DNA polymerase described in this work is the most abundant in proliferating cells of Oryza sativa L., Parthenocissus tricuspidata (Siebold et Zucc.) Planchon, Acer pseudoplatanus L., and Medicago sativa L. and responds to changes in the rate of cell multiplication. We therefore postulate that this α-like DNA polymerase is the replicating enzyme of plant cells.