Herbivory could unlock mutations sequestered in stratified shoot apices of genetic mosaics.

Many higher plants have shoot apical meristems that possess discrete cell layers, only one of which normally gives rise to gametes following the transition from vegetative meristem to floral meristem. Consequently, when mutations occur in the meristems of sexually reproducing plants, they may or may not have an evolutionary impact, depending on the apical layer in which they reside. In order to determine whether developmentally sequestered mutations could be released by herbivory (i.e., meristem destruction), a characterized genetic mosaic was subjected to simulated herbivory. Many plants develop two shoot meristems in the leaf axils of some nodes, here referred to as the primary and secondary axillary meristems. Destruction of the terminal and primary axillary meristems led to the outgrowth of secondary axillary meristems. Seed derived from secondary axillary meristems was not always descended from the second apical cell layer of the terminal shoot meristem as is expected for terminal and primary shoot meristems. Vegetative and reproductive analysis indicated that secondary meristems did not maintain the same order of cell layers present in the terminal shoot meristem. In secondary meristems reproductively sequestered cell layers possessing mutant cells can be repositioned into gamete-forming cell layers, thereby adding mutant genes into the gene pool. Herbivores feeding on shoot tips may influence plant evolution by causing the outgrowth of secondary axillary meristems.

LAM1 is required for dorsoventrality and lateral growth of the leaf blade in Nicotiana.

The role of LAM1 in dorsoventrality and lateral growth of the leaf blade was investigated in the 'bladeless' lam1 mutant of Nicotiana sylvestris and in periclinal chimeras with lam1 and wild-type (N. glauca) cell layers. Mutant lam1 primordia show normal dorsoventrality at emergence, but produce blade tissue that lacks dorsal cell types and fails to expand in the lateral plane. In leaves of a lam1-glauca-glauca (L1-L2-L3) chimera, we observed restoration of dorsal identity in the lam1 upper epidermis, suggesting non-cell-autonomous movement of a dorsalizing factor between cell layers of the blade. A lam1-lam1-glauca chimera generated a leaf blade with lam1 cells in the L1-derived epidermis and the L2-derived upper and lower mesophyll. An in situ lineage analysis revealed that N. glauca cells in the L3-derived middle mesophyll restore palisade differentiation in the adjoining lam1 upper mesophyll. Movement of dorsalizing information appears short-range, however, having no effect on the upper lam1 epidermis in lam1-lam1-glauca. Clusters of lam1 mesophyll in distal or proximal positions show a localized default to radial growth, indicating that the LAM1 function is required for dorsoventrality and lateral growth throughout blade expansion.

Cell-layer interactions influence the number and position of lateral shoot meristems in Nicotiana.

In higher plants the formation of lateral shoot meristems (i.e., axillary meristems) in the axils of leaves establishes potential growth centers along the principal axis of the stem. The position and number of lateral buds in Nicotiana glauca, two genotypes of Nicotiana tabacum, and a series of interspecific periclinal chimeras composed of these species was studied to establish the role of position (location of the node along the main axis), flowering, and cell-layer interactions on the pattern of lateral meristem initiation. In N. glauca, both the number of nodes generated and the transition to flowering influenced the number and position of lateral meristems. A short-day mutant of N. tabacum grown under long days remained vegetative and never produced multiple lateral buds per node, indicating that attaining a certain node number was not sufficient to cause the formation of multiple buds. Yet, flowering plants of both short-day and day-neutral N. tabacum possessed multiple buds in their upper nodes. An analysis of the number of buds per node and bud position along the main axis in periclinal chimeras indicated that the genotype of the third apical layer (LIII) of the meristem had the greatest influence on the pattern of lateral shoot meristems in both vegetative and flowering plants. The lineage of the three apical layers (LI, LII, and LIII) of the terminal shoot meristem was preserved in primary (1 degree) lateral meristems but minor deviations in lineage were observed in secondary (2 degrees) buds (i.e., those formed later but in the same node as the 1 degree bud). An analysis of the phenotype of 2 degrees shoots that displayed deviations from expected lineage indicated that in most cases the periclinal cell divisions that destabilized the lineage occurred at the flanks of the meristem and began before the most basal node, indicating that periclinal cell divisions most likely occurred prior to the inception of the 2 degrees lateral meristem. Based on our studies, we conclude that both 1 degree and 2 degrees lateral meristems in Nicotiana ultimately descend from derivatives of all three apical layers of the terminal shoot meristem.

Origin and development of adventitious shoot meristems initiated on plant chimeras.

Most studies concerning the initiation and development of the shoot apical meristem have been performed on meristems that developed during embryogenesis. We characterized the in situ formation of adventitious shoots originating from cells located near leaf axils in a series of six interspecific periclinal tobacco chimeras. Shoots were generated by decapitating the plants and removing all of the axillary buds. Eighty-four of the 413 shoots regenerated were chimeral. Many of the shoots were complex mericlinal chimeras, with several of their axillary buds possessing meristems arranged as periclinal chimeras. The adventitious shoots originated from derivatives of only the second and third cell layers in the meristem of the source plant since derivatives of the outermost meristem layer were removed with the removal of the axillary buds. With time, nearly all adventitious shoot apices that were initially chimeral became nonchimeral or stabilized as periclinal chimeras. A statistical analysis indicated that the arrangement of genetically distinct tissues in the cell layers of adventitious meristems was influenced by competition between cell types. Our method for generating these shoots can be used to create small genetically distinct sectors analogous to radiation-induced sectors as well as a complete series of periclinical chimeras, both of which have potential for use in determining tissue-to-tissue interactions. An analysis of sector length and position on chimeral shoots indicates that the first one to three leaves of adventitious shoots do not arise as derivatives of the shoot apical initials of a meristem proper.