The Subcellular Localization of DNA Components from Cyanophora paradoxa, a Flagellate Containing Endosymbiotic Cyanelles

Abstract

C. paradoxa, a unicellular flagellate, contains cyanelles which are supposed to be cyanobacterial in origin. DNA was isolated from subcellular fractions and separated according to density components in CsCl density gradients. The main DNA component, comprising > 90% of the total DNA, has a buoyant density of 1.724 g .cntdot. cm-3. Several subfractions in the range from 1.718-1.735 g .cntdot. cm-3 are contained in this component. This DNA of high complexity was considered to be host nuclear DNA. The DNA from the endosymbiotic cyanelles, which were isolated, treated with DNase and purified by sucrose density gradient centrifugation, exhibited a buoyant density of 1.692 g .cntdot. cm-3 in 1 strain and 1.695 g .cntdot. cm-3 in a 2nd strain. Both cyanelle DNA (cyDNA) have a complexity of .apprx. 126 .times. 103 base pairs and comprise .apprx. 5% of the total cellular DNA content. Two additional DNA components of low complexity were isolated from crude cyanelle pellets obtained without DNase treatment. The larger of these, .apprx. 48 .times. 103 base pairs in size, had a density of .apprx. 1.688 g .cntdot. cm-3. The 2nd component, .apprx. 15 .times. 103 base pairs in size, banded in the density range between 1.710 g .cntdot. cm-3 and 1.720 g .cntdot. cm-3. The latter is associated with nuclear DNA. The 48 .times. 103-base-pair component was located in the cytosol and could be obtained after CsCl/ethidium bromide density gradient centrifugation at the position of covalently closed circular DNA. Both these components amounted to .apprx. 0.5-1% of total DNA. A further DNA component with a complexity of > 150 .times. 103 base pairs, enriched in fractions where mitochondria are expected, was not characterized further. The density was intermediate between cyDNA and nuclear DNA (1.710-1.720 g .cntdot. cm-3) and it amounted to 1-2% of the total DNA. Evidently, the DNA from cyanelles, believed to be endosymbiotic cyanobacteria, is not more complex than higher plant chloroplast DNA.