Morphogenesis of a Dissected Birch Leaf in vitro Culture

Abstract

The Betula pendula f. “dalecarlica” is a variety of silver birch (B. pendula) that has dissected leaves. The possible nature of the disturbance of leaf morphogenesis (the transition from a dissected shape to a normal whole leaf) in one of the eight clones of Betula “dalecarlica” during in vitro cultivation was investigated. Karyological analysis revealed a higher degree of chromatin compaction of metaphase chromosomes in the revertant clone versus the clone with dissected leaves. Reversion to the wild type is suggested to have an epigenetic nature. This is indicated by a pronounced reaction (appearance of leaves with dissection) of the explants of the revertant clone to the effect of the 5-azacytidine epimutagen (which has a demethylating effect on DNA) and a significant increase (by six times) in the proportion of cells with residual nucleoli in the metaphase and anaphase of mitosis. It is assumed that changes in the hormonal status of the cell (when explants were introduced into in vitro culture), the level of DNA methylation, and the degree of chromatin compaction could change the expression pattern of regulatory genes in the shoot apex (including the repression of transcription of key homeobox KNOX1 genes) and leaf reversion to the wild type. Moreover, the “epigenetic silence” induced by the in vitro conditions is quite stable and long-term: it has been observed for 19 years of clonal micropropagation using nutrient media without hormones, and it is also preserved after planting microplants in the greenhouse (ex vitro). Cultivated in vitro clones of Betula “dalecarlica,” contrasting in stability manifestations of the sign of dissected leaves, are a promising model for further studying the genetics of leaf morphogenesis and mechanisms of epigenetic variability.