The morphological and physiological basis of delayed pollination overcoming pre-fertilization cross-incompatibility in Nicotiana

Delayed pollination is widely used in overcoming the pre-fertilization incompatibility, but its regulatory mechanisms are unclear. When Nicotiana tabacum was cross pollinated with pollen of N. alata, the incompatibility occurring in the basal 1/4 region of the style (pollinated at anthesis: 0-d-old pistil) was overcome by delayed pollination (of 6-d-old pistil), and the morphological changes and the corresponding physiological basis were explored here. The structure and ultrastructure of the pistil were observed under the fluorescence microscopy and transmission electron microscopy. Differentially expressed proteins were screened with a monoclonal antibody chip of Nicotiana, and the protein expression and distribution were analyzed by immunofluorescence. The cellulase and pectinase activities were tested using the enzyme linked immunosorbent assay kits. The style of Nicotiana is solid in the basal region, and pollen tubes grow in the extracellular spaces (ECM) of the transmitting tissue (TTS) cells. Seven of the twenty-two identified proteins were cell wall associated proteins, and they were expressed at higher levels during the pistil senescence. Cellulase and pectinase activities were increased. The TTS cells in the basal 1/4 region of the 0-d old style were polygonal and tightly arranged with narrow ECM, but were oval, or partially dissolved in the 6-d-old pistil, leading to wider ECM and richer secretions. The increased expression of the cell wall proteins and the enhanced enzyme activity during the pistil senescence might partially be responsible for the cells becoming oval and the ECM enlarged, providing the morphological basis for delayed pollination overcoming the pre-fertilization incompatibility between N. tabacum and N. alata.

Keywords: Nicotiana; cell morphology; cell wall proteins; delayed pollination; incompatibility.