Stem cells self-renew and repeatedly produce differentiated cells during development and growth. The differentiated cells can be converted into stem cells in some metazoans and land plants with appropriate treatments. After leaves of the moss Physcomitrella patens are excised, leaf cells reenter the cell cycle and commence tip growth, which is characteristic of stem cells called chloronema apical cells. To understand the underlying molecular mechanisms, a digital gene expression profiling method using mRNA 5′-end tags (5′-DGE) was established. The 5′-DGE method produced reproducible data with a dynamic range of four orders that correlated well with qRT-PCR measurements. After the excision of leaves, the expression levels of 11% of the transcripts changed significantly within 6 h. Genes involved in stress responses and proteolysis were induced and those involved in metabolism, including photosynthesis, were reduced. The later processes of reprogramming involved photosynthesis recovery and higher macromolecule biosynthesis, including of RNA and proteins. Auxin and cytokinin signaling pathways, which are activated during stem cell formation via callus in flowering plants, are also activated during reprogramming in P. patens, although no exogenous phytohormone is applied in the moss system, suggesting that an intrinsic phytohormone regulatory system may be used in the moss.

Both land plants and metazoa have the capacity to reprogram differentiated cells to stem cells. Here we show that the moss Physcomitrella patens Cold-Shock Domain Protein 1 (PpCSP1) regulates reprogramming of differentiated leaf cells to chloronema apical stem cells and shares conserved domains with the induced pluripotent stem cell factor Lin28 in mammals. PpCSP1 accumulates in the reprogramming cells and is maintained throughout the reprogramming process and in the resultant stem cells. Expression of PpCSP1 is negatively regulated by its 3′-untranslated region (3′-UTR). Removal of the 3′-UTR stabilizes PpCSP1 transcripts, results in accumulation of PpCSP1 protein and enhances reprogramming. A quadruple deletion mutant of PpCSP1 and three closely related PpCSP genes exhibits attenuated reprogramming indicating that the PpCSP genes function redundantly in cellular reprogramming. Taken together, these data demonstrate a positive role of PpCSP1 in reprogramming, which is similar to the function of mammalian Lin28.

DEAD-box RNA helicases compose a large subfamily of RNA helicases found in all eukaryotes and prokaryotes. Functional DEAD-box RNA helicases are considered to be RNA chaperones that modify RNA secondary structure and perhaps three-dimensional structures during different cellular processes involving RNA metabolism. Although a relationship of DEAD-box RNA helicases to abiotic stress adaptation is known, few reports document the involvement of RNA helicases during plant growth and development. In this study, detailed analyses were performed for two cold-responsive DEAD-box RNA helicases, AtRH22 and AtRH52, on the transcript level during different developmental stages of Arabidopsis thaliana. Expression levels of AtRH22 and AtRH52 were up-regulated markedly in response to cold stress and were enriched in shoot apical meristems, floral buds, and siliques. The expression of AtRH22 and AtRH52 was spatiotemporally regulated during vegetative growth, floral transition and embryonic development. To investigate, the functional role of AtRH22 and AtRH52 in A. thaliana, we functionally characterized independent T-DNA insertion mutant lines for both genes. Genotypic analysis of self-fertilized heterozygous lines revealed only heterozygous (WT/T-DNA) and homozygous wild-type progeny for AtRH22 and AtRH52. Self-fertilized heterozygous mutants of AtRH22 and AtRH52 show normal vegetative phenotype, but produced normal-sized and abortive seeds. Collectively, these results demonstrate that the cold-responsive AtRH22 and AtRH52 genes are spatiotemporally regulated during plant development and are essential during Arabidopsis embryogenesis.