Cloning and characterization of cDNAs associated with the embryogenic dedifferentiation of tobacco immature pollen grains

We conducted differential screening to obtain cDNAs showing that gene expression is highly associated with the transformation from immature pollen to embryogenic cell, so-called embryogenic dedifferentiation of pollen, in a Nicotiana tabacum pollen culture system and analyzed their expression and sequences. Seventy-seven cDNA clones were independently isolated and distinguished into 16 groups based on their sequences. The groups were further categorized into two classes, Class I and II, based on the gene expression pattern of the representative clone of each group under various pollen culture conditions arranged for examining the coincidence with the dedifferentiation. The 13 groups in Class I showed prominent expression under the conditions allowing or facilitating pollen dedifferentiation and the expression level increased earlier than A-type cyclin genes, but they were not markedly expressed in the cell populations rich in S-phase cells, i.e. young anthers with pollen mother cells, BY-2 cells at the growth phase and early phase embryos derived from immature pollen. The other three groups in Class II encoded homologs to H1 histone, H2A histone and minichromosome maintenance (MCM) protein, respectively. The level of their transcripts increased during dedifferentiation, but it was also high in anthers containing pollen mother cells and in the proliferating BY-2 cells indicating that their expression is coincident with the S phase but not with dedifferentiation. These findings suggest that pollen dedifferentiation is a complex process accompanied with the reentrance of cell cycle and unknown events probably caused by specific expression of many genes, at least, listed in Class I. These genes should be used as reliable markers and important clues for further studies on the molecular mechanism of dedifferentiation.