Differentiation of cellular and biochemical features of the single-cell C4 syndrome during leaf development in Bienertia cycloptera (Chenopodiaceae).

The terrestrial plant Bienertia cycloptera has been shown to accomplish C(4) photosynthesis within individual chlorenchyma cells by spatially separating the phases of carbon assimilation into distinct peripheral and central compartments. In this study, anatomical, physiological, and biochemical techniques were used to determine how this unique compartmentation develops. Western blots show ribulose-1,5-bisphosphate carboxylase (Rubisco) (chloroplastic) is present in the youngest leaves and increases during development, while levels of C(4) enzymes-pyruvate,Pi dikinase (chloroplastic), phosphoenolpyruvate carboxylase (PEPC) (cytosol), and NAD-malic enzyme (mitochondrial)-increase later in development. Immunolocalization confirmed this for Rubisco and PEPC. The youngest chlorenchyma cells have a central nucleus surrounded by monomorphic granal chloroplasts containing Rubisco. Later stages show progressive development of a central cytoplasmic compartment enriched with chloroplasts and mitochondria and of a peripheral cytoplasm with chloroplasts. A complex reticulum of connections between the compartments also developed and was characterized. δ(13)C isotope analyses show mature leaves have distinct C(4)-type isotope composition, while the composition in younger leaves is "C(4)-like." Based on the results, this form of single-cell C(4) photosynthesis develops from a common pool of organelles through partitioning to separate compartments, and the development of biochemically and ultrastructurally dimorphic chloroplasts.