Evidence that genetic information for chloroplast coupling factor I is shared by nuclear and chloroplast DNA

Abstract

Using the method developed by Lien and Racker (Lien, S. and Racker, E. (1971) in Methods in Enzymology (San Pietro, A., ed.), Vol. 23, Part A, pp. 547–555, Academic Press, New York) for spinach leaves, coupling factor I was isolated from 2-kg quantities of Nicotiana glutinosa, N. tabacum, N. excelsior, N. langsdorffii leaves, and shown to consist of 5 subunits of comparable monomeric molecular weights to coupling factor I from spinach and also to be a trypsin-activated, Ca 2+-dependent ATPase. Limitation of hybrid plant material necessitated use of polyacrylamide electrophoresis to separate Fraction I protein ( M r = 550 000 with a monomeric large subunit of 56 000) from coupling factor I ( M r = 350 000; β-subunit of 56 000) so that approx. 50 μg of pure coupling factor I could be isolated from 50-g quantities of coupling factor I reciprocal hybrid plants and the coupling factor I subjected to electrofocussing in 8 M urea simultaneously with coupling factor I from the two parental species. Electrofocusing resolved 3 or 4 of the 5 subunits contained in the coupling factor I from leaves of the four Nicotiana species tested. All were like N. tabacum in regard to isoelectric points and staining intensity of 4 subunits except N. glutinosa coupling factor I which displayed 2 subunits which differed in staining intensity from 2 comparable subunits in coupling factor I of the other species. N. glutinosa coupling factor I also lacked one subunit resolved in the other species. The genetic information determining the difference in staining intensity of the 2 subunits of coupling factor I contained in N. tabacum × N. glutinosa reciprocal coupling factor I hybrids was inherited only by the maternal line and is therefore most likely contained in chloroplast DNA. Information determining the isoelectric point of a third subunit was transmitted by pollen to an coupling factor I hybrid and was therefore contained in nuclear DNA. No difference in staining intensity or isoelectric points could be detected in a fourth subunit so the source of its coding information remains unknown.