Role of auxin in regulating Arabidopsis flower development.

To elucidate the role of auxin in flower morphogenesis, its distribution patterns were studied during flower development in Arabidopsis thaliana (L.) Heynh. Expression of DR5::GUS was regarded to reflect sites of free auxin, while immunolocalization with auxin polyclonal antibodies visualized conjugated auxin distribution. The youngest flower bud was loaded with conjugated auxin. During development, the apparent concentration of free auxin increased in gradual patterns starting at the floral-organ tip. Anthers are major sites of high concentrations of free auxin that retard the development of neighboring floral organs in both the acropetal and basipetal directions. The IAA-producing anthers synchronize flower development by retarding petal development and nectary gland activity almost up to anthesis. Tapetum cells of young anthers contain free IAA which accumulates in pollen grains, suggesting that auxin promotes pollen-tube growth towards the ovules. High amounts of free auxin in the stigma induce a wide xylem fan immediately beneath it. After fertilization, the developing embryos and seeds show elevated concentrations of auxin, which establish their axial polarity. This developmental pattern of auxin production during floral-bud development suggests that young organs which produce high concentrations of free IAA inhibit or retard organ-primordium initiation and development at the shoot tip.

Root-synthesized cytokinin in Arabidopsis is distributed in the shoot by the transpiration stream.

To clarify how root-synthesized cytokinins (CKs) are transported to young shoot organs, CK distribution patterns were analysed in free-CK-responsive ARR5::GUS transformants of Arabidopsis thaliana (L.) Heynh. together with free plus bound CKs using specific CK monoclonal antibodies. Plants were subjected to two different growth conditions, completely protected from any air movement, or exposed to gentle wind 3 h before harvesting. In wind-protected plants the strongest ARR5::GUS expression was found in the root cap statocytes, spreading upwards in the vascular cylinder. This pattern in roots was congruent with that found by CK immunolocalization. Shoots of wind-protected plants displayed either no or only low ARR5::GUS expression in the stem vascular bundles, nodal ramifications, and the bases of flower buds; shoot vascular bundles showed patterns of acropetally decreasing staining and the apical parts of buds and leaves were free from ARR5::GUS expression. In wind-exposed plants ARR5::GUS expression was considerably increased in shoots, also in basal-to-apical decreasing gradients. Immunolabelled shoots showed differential staining, with the strongest label in the vascular bundles of stems, leaves, and buds. The fact of the apparent absence of free CK in the buds of wind-protected plants and the typical upward decreasing gradients of free and conjugated CKs suggest that the bulk of the CK is synthesized in the root cap, exported through the xylem and accumulates at sites of highest transpiration where cuticles do not yet exist or do not protect against water loss.

Role of cytokinin in the regulation of root gravitropism.

The models explaining root gravitropism propose that the growth response of plants to gravity is regulated by asymmetric distribution of auxin (indole-3-acetic acid, IAA). Since cytokinin has a negative regulatory role in root growth, we suspected that it might function as an inhibitor of tropic root elongation during gravity response. Therefore, we examined the free-bioactive-cytokinin-dependent ARR5::GUS expression pattern in root tips of transformants of Arabidopsis thaliana (L.) Heynh., visualized high cytokinin concentrations in the root cap with specific monoclonal antibodies, and complemented the analyses by external application of cytokinin. Our findings show that mainly the statocytes of the cap produce cytokinin, which may contribute to the regulation of root gravitropism. The homogenous symmetric expression of the cytokinin-responsive promoter in vertical root caps rapidly changed within less than 30 min of gravistimulation into an asymmetrical activation pattern, visualized as a lateral, distinctly stained, concentrated spot on the new lower root side of the cap cells. This asymmetric cytokinin distribution obviously caused initiation of a downward curvature near the root apex during the early rapid phase of gravity response, by inhibiting elongation at the lower side and promoting growth at the upper side of the distal elongation zone closely behind the root cap. Exogenous cytokinin applied to vertical roots induced root bending towards the application site, confirming the suspected inhibitory effect of cytokinin in root gravitropism. Our results suggest that the early root graviresponse is controlled by cytokinin. We conclude that both cytokinin and auxin are key hormones that regulate root gravitropism.