Maize miniendosperm proliferation in vitro is characterized by tracheary element formation

Maize grain is of a great economic importance worldwide. Morphometric and cytological descriptions of developing maize endosperms are available in inbred lines growing under natural conditions. Despite tremendous potential in biotechnology, similar studies focused on endosperm development in vitro are rather rare. Here we provide a comparative study on growth and development of endosperms in planta versus in vitro, considering regulation by different cytokinins during endosperm cultivation. Histochemical analyses were performed on hand-cut sections of samples stained with Sudan III. Microtubules (MTs) were immunolocalized in whole mount samples and Steedman?s wax sections of endosperm and miniendosperm from in vitro conditions. Our results confirmed that in vitro cultured maize endosperms proliferate and establish aleurone and starchy cells like endosperms grown in planta, however later ones undergo a distinct developmental program triggered/controlled by cytokinins. Higher mitotic activity of surface layer cells resulted in the miniendosperm proliferation, while the centrally localized parenchymatic starchy-like cells differentiated into tracheary elements (TEs). Furthermore, we report, for the first time, on the formation of numerous lobes and trichomes on pericarp cells. MT rearrangements play an important regulatory role in the differentiation of TEs. Formation of functional vascular tissue and lipid accumulation appears to be fundamental for maize miniendosperm proliferation. In contrast, the presence of maternal tissue (pericarp and nucellus) prevents miniendosperm growth and development. We assume the formation of lobes and trichomes is an adaptive response of in vitro maize endosperm to high temperatures.