The association of chloroplast DNA with photosynthetic membrane vesicles from spinach chloroplasts.

To investigate the association between chloroplast DNA (cp DNA) and the photosynthetic membranes of spinach chloroplasts, previously suggested by electron-microscope autoradiography, use has been made of vesicles formed by isolating chloroplasts directly in 3.5 mM Mg2+. These chloroplast vesicles consist of photosynthetic membranes, separate from chloroplast envelope membranes. Light and electron microscopy confirm that the vesicles consist of swollen stroma lamellar membranes with some peripheral grana lamellae that are much less swollen. Vesicles labelled with [H]thymidine were obtained from [3H]thymidine-labelled chloroplasts from spinach disks in which chloroplast division and cp DNA synthesis and segregation were occurring. The chloroplast vesicle fraction retains about 45% of the cp DNA as determined by liquid scintillation counting. The cp DNA-membrane associations do not appear to be dependent on the presence of Mg2+. The chloroplast vesicles can be autoradiographed for light microscopy if they are fixed in formaldehyde and no centrifugation steps are used. Light-microscope autoradiography is consistent with a preferential labelling of grana as opposed to stroma membranes, and long lengths of membrane are labelled. It appears that in spinach chloroplasts cp DNA is associated with granal thylakoids at intervals along the length of a continuous photosynthetic membrane system. Such an organization would facilitate cp DNA segregation during chloroplast division.