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In vitro regeneration of Arabidopsis from roots is generally achieved via indirect organogenesis. First, transdifferentiation of lateral root primordia to calli is achieved by a balanced auxin-to-cytokinin ratio that is followed by the induction of shoot meristem formation using a high cytokinin level. Here we demonstrate that if the root explants were transferred onto a hormone-free medium after a transient (4-days) cytokinin treatment, embryogenic marker genes (LEC1, LEC2, FUS3) started to be expressed. App. 50% of the regeneration foci developed into plantlets with trichome-less cotyledon-like leaves. Moreover, the somatic embryogenesis defective lec1 mutant could regenerate only shoots with trichome-bearing leaves under this condition. Based on these observations, the mixed accomplishment of shoot organogenesis and somatic embryogenesis is hypothesized in the Arabidopsis root explants cultured under hormone-free conditions following cytokinin induction. Using whole seedlings instead of root explants in the same experimental set up, no regenerates were formed on the roots. Applying the auxin transport inhibitor TIBA to the root-to-shoot junction of the seedlings, the regeneration ability of the root could be restored. The observations indicate that shoot-derived endogenous auxin blocks the cytokinin-induced regeneration process in the roots of whole seedlings. The expression of the wound-induced transcription factor WIND1 could be detected in the roots of unwounded seedlings if the shoot-to-root auxin transport was inhibited. Manipulating the exogenous cytokinin level together with the endogenous shoot-to-root auxin transport therefore could mimic the effect of wounding (removal of shoot) on plant regeneration from roots. Transferring root explants from high cytokinin to hormone-free conditions resulted in the expression of embryogenic markers. Inhibiting the shoot-to-root auxin transport had similar effect on regeneration as wounding.

Plant nucleosome assembly protein-related proteins (NRPs) are histone chaperons involved in nucleosome turnover. Despite this basic cellular function, the Arabidopsis nrp1-1 nrp2-1 knock out mutant has been reported to exhibit only mild seedling root phenotypes and to significantly affect the expression of only few hundred genes Zhu et al. (2006). Here we report that NRP loss-of-function as well as the ectopic overexpression of At NRP1 significantly affected the growth, development, and the pathogen response of Arabidopsis plants under short day conditions. The nrp1-1 nrp2-1 mutant grew faster and flowered weeks earlier than the wild type and the overexpressor. The latter developed slower and flowered at a lower number of leaves than the mutant and the wild type. Moreover, the mutant was more sensitive, the overexpressor was more tolerant to pathogen-induced necrosis correlating with their more adult and juvenile character, respectively. Transcriptomic comparison of mature non-bolting plants agreed with the phenotypes. The presented and other published data indicate that although NRPs might not be absolutely required for normal plant growth and development, their level needs to be controlled to allow the epigenetic coordination of metabolic, growth, defence and developmental processes during the acclimation to unfavourable growth conditions such as short days.

It is widely believed that all cells of a plant are totipotent and can regenerate the whole organism. This view is supported by uncountable experimental observations demonstrating the regrowth of whole plants from various explants, even from single cells. However, recent investigations have demonstrated that plant regeneration may proceed via transdifferentiation of meristems or root meristem-like callus tissues due to adult stem cells present all over the plants. These pathways do not start from single totipotent cells. There is a strong argument for plant cell totipotency, however, and that is somatic embryogenesis. During this process, differentiated somatic cells change their fate to develop into an embryo. Animal embryos can develop only from the totipotent zygote and its direct descendants (this cell state can also be artificially produced by injecting a somatic cell nucleus into an egg cell cytoplasm during cloning). Plant cells have to be induced to start somatic embryogenesis and not all of them are competent to respond properly to the induction. In conclusion, plant cells cannot be considered as totipotent per se, but some of them can regain totipotency under appropriate conditions. In addition, accumulating evidence supports the view that even somatic embryo development can follow various initial steps not necessarily requiring cellular totipotency. Although, there are experimental observations to support the progression of somatic embryogenesis through a zygote-like state in certain experimental systems, in other instances the reorganization of several cells into an embryo was described. The direct release/induction of the embryo development program in vegetative plant cells may represent a third pathway of somatic embryo development. In this chapter, a brief literature review is provided to support the above view on plant cell totipotency as well as on the various ways to start somatic embryogenesis.