Auxin-induced WUS expression is essential for embryonic stem cell renewal during somatic embryogenesis in Arabidopsis

Somatic embryogenesis requires auxin and establishment of the shoot apical meristem (SAM). WUSCHEL ( WUS ) is critical for stem cell fate determination in the SAM of higher plants. However, regulation of WUS expression by auxin during somatic embryogenesis is poorly understood. Here, we show that expression of several regulatory genes important in zygotic embryogenesis were up-regulated during somatic embryogenesis of Arabidopsis. Interestingly, WUS expression was induced within the embryonic callus at a time when somatic embryos could not be identified morphologically or molecularly. Correct WUS expression, regulated by a defined critical level of exogenous auxin, is essential for somatic embryo induction. Furthermore, it was found that auxin gradients were established in specific regions that could then give rise to somatic embryos. The establishment of auxin gradients was correlated with the induced WUS expression. Moreover, the auxin gradients appear to activate PIN1 polar localization within the embryonic callus. Polarized PIN1 is probably responsible for the observed polar auxin transport and auxin accumulation in the SAM and somatic embryo. Suppression of WUS and PIN1 indicated that both genes are necessary for embryo induction through their regulation of downstream gene expression. Our results reveal that establishment of auxin gradients and PIN1-mediated polar auxin transport are essential for WUS induction and somatic embryogenesis. This study sheds new light on how auxin regulates stem cell formation during somatic embryogenesis.     

The essential gene EMB1611 maintains shoot apical meristem function during Arabidopsis development

The Arabidopsis thaliana genome contains hundreds of genes essential for seed development. Because null mutations in these genes cause embryo lethality, their specific molecular and developmental functions are largely unknown. Here, we identify a role for EMB1611/MEE22 , an essential gene in Arabidopsis, in shoot apical meristem maintenance. EMB1611 encodes a large, novel protein with N-terminal coiled-coil regions and two putative transmembrane domains. We show that the partial loss-of-function emb1611-2 mutation causes a range of pleiotropic developmental phenotypes, most dramatically a progressive loss of shoot apical meristem function that causes premature meristem termination. emb1611-2 plants display disorganization of the shoot meristem cell layers early in development, and an associated stem cell fate change to an organogenic identity. Genetic and molecular analysis indicates that EMB1611 is required for maintenance of the CLV-WUS stem cell regulatory pathway in the shoot meristem, but also has WUS -independent activity. In addition, emb1611-2 plants have reduced shoot and root growth, and their rosette leaves form trichomes with extra branches, a defect we associate with an increase in endoreduplication. Our data indicate that EMB1611 functions to maintain cells, particularly those in the shoot meristem, roots and developing rosette leaves, in a proliferative or uncommitted state.     

WVD2 is a novel microtubule-associated protein in Arabidopsis thaliana

Arabidopsis WAVE-DAMPENED 2 (WVD2) was identified by forward genetics as an activation-tagged allele that causes plant and organ stockiness and inversion of helical root growth handedness on agar surfaces. Plants with high constitutive expression of WVD2 or other members of the WVD2-LIKE (WDL) gene family have stems and roots that are short and thick, have reduced anisotropic cell elongation, are suppressed in a root-waving phenotype, and have inverted handedness of twisting in hypocotyls and roots compared with wild-type. The wvd2-1 mutant shows aberrantly organized cortical microtubules in peripheral root cap cells as well as reduced branching of trichomes, unicellular leaf structures whose development is regulated by microtubule stability. Orthologs of the WVD2/WDL family are found widely throughout the plant kingdom, but are not similar to non-plant proteins with the exception of a C-terminal domain distantly related to the vertebrate microtubule-associated protein TPX2. in vivo, WVD2 and its closest paralog WDL1 are localized to interphase cortical microtubules in leaves, hypocotyls and roots. Recombinant glutathione-S-transferase:WVD2 or maltose binding protein:WVD2 protein bind to and bundle microtubules in vitro. We speculate that a C-terminal domain of TPX2 has been utilised by the WVD2 family for functions critical to the organization of plant microtubules.     

BUD2, encoding an S-adenosylmethionine decarboxylase, is required for Arabidopsis growth and development

Polyamines are implicated in regulating various developmental processes in plants, but their exact roles and how they govern these processes still remain elusive. We report here an Arabidopsis bushy and dwarf mutant, bud2, which results from the complete deletion of one member of the small gene family that encodes S-adenosylmethionine decarboxylases （SAMDCs） necessary for the formation of the indispensable intermediate in the polyamine biosynthetic pathway. The bud2 plant has enlarged vascular systems in inflorescences, roots, and petioles, and an altered homeostasis ofpolyamines. The double mutant of bud2 and samdcl, a knockdown mutant of another SAMDC member, is embryo lethal, demonstrating that SAMDCs are essential for plant embryogenesis. Our results suggest that polyamines are required for the normal growth and development of higher plants.     

A leucine-rich repeat protein is required for growth promotion and enhanced seed production mediated by the endophytic fungus Piriformospora indica in Arabidopsis thaliana

Piriformospora indica, a basidiomycete of the Sebacinaceae family, promotes the growth, development and seed production of a variety of plant species. Arabidopsis plants colonized with the fungus produce 22% more seeds than uncolonized plants. Deactivating the Arabidopsis single-copy gene DMI-1, which encodes an ion carrier required for mycorrihiza formation in legumes, does not affect the beneficial interaction between the two symbiotic partners. We used cellular and molecular responses initiated during the establishment of the interaction between P. indica and Arabidopsis roots to isolate mutants that fail to respond to the fungus. An ethylmethane sulfonate mutant (Piriformospora indica-insensitive-2; pii-2), and a corresponding insertion line, are impaired in a leucine-rich repeat protein (At1g13230). The protein pii-2, which contains a putative endoplasmic reticulum retention signal, is also found in Triton X-100-insoluble plasma membrane microdomains, suggesting that it is present in the endoplasmic reticulum/plasma membrane continuum in Arabidopsis roots. The microdomains also contain an atypical receptor protein (At5g16590) containing leucine-rich repeats, the message of which is transiently upregulated in Arabidopsis roots in response to P. indica. This response is not detectable in At1g13230 mutants, and the protein is not detectable in the At1g13230 mutant microdomains. Partial deactivation of a gene for a sphingosine kinase, which is required for the biosynthesis of sphingolipid found in plasma membrane microdomains, also affects the Arabidopsis/P. indica interaction. Thus, pii-2, and presumably also At5g16590, two proteins present in plasma membrane microdomains, appear to be involved in P. indica-induced growth promotion and enhanced seed production in Arabidopsis thaliana.     

SABRE is required for stabilization of root hair patterning in Arabidopsis thaliana

Patterned differentiation of distinct cell types is essential for the development of multicellular organisms. The root epidermis of Arabidopsis thaliana is composed of alternating files of root hair and non‐hair cells and represents a model system for studying the control of cell‐fate acquisition. Epidermal cell fate is regulated by a network of genes that translate positional information from the underlying cortical cell layer into a specific pattern of differentiated cells. While much is known about the genes of this network, new players continue to be discovered. Here we show that the SABRE (SAB) gene, known to mediate microtubule organization, anisotropic cell growth and planar polarity, has an effect on root epidermal hair cell patterning. Loss of SAB function results in ectopic root hair formation and destabilizes the expression of cell fate and differentiation markers in the root epidermis, including expression of the WEREWOLF (WER) and GLABRA2 (GL2) genes. Double mutant analysis reveal that wer and caprice (cpc) mutants, defective in core components of the epidermal patterning pathway, genetically interact with sab. This suggests that SAB may act on epidermal patterning upstream of WER and CPC. Hence, we provide evidence for a role of SAB in root epidermal patterning by affecting cell‐fate stabilization. Our work opens the door for future studies addressing SAB‐dependent functions of the cytoskeleton during root epidermal patterning.     

SHA1, a novel RING finger protein, functions in shoot apical meristem maintenance in Arabidopsis

Post-embryonic plant growth is dependent on a functional shoot apical meristem (SAM) that provides cells for continuous development of new aerial organs. However, how the SAM is dynamically maintained during vegetative development remains largely unclear. We report here the characterization of a new SAM maintenance mutant, sha1-1 (shoot apical meristem arrest 1-1), that shows a primary SAM-deficient phenotype at the adult stage. The SHA1 gene encodes a novel RING finger protein, and is expressed most intensely in the shoot apex. We show that, in the sha1-1 mutant, the primary SAM develops normally during the juvenile vegetative stage, but cell layer structure becomes disorganized after entering the adult vegetative stage, resulting in a dysfunctional SAM that cannot initiate floral primordia. The sha1-1 SAM terminates completely at the stage when the wild-type begins to bolt, producing adult plants with a primary inflorescence-deficient phenotype. These observations indicate that SHA1, a putative E3 ligase, is required for post-embryonic SAM maintenance by controlling proper cellular organization.     

Different Dicer-like protein components required for intracellular and systemic antiviral silencing in Arabidopsis thaliana

Eukaryotes employ RNA silencing as an innate defense system against invading viruses. Dicer proteins play the most crucial role in initiating this antiviral pathway as they recognize and process incoming viral nucleic acids into small interfering RNAs. Generally, 2 successive infection stages constitute viral infection in plants. First, the virus multiplies in initially infected cells or organs after viral transmission and then the virus subsequently spreads systemically through the vasculature to distal plant tissues or organs. Thus, antiviral silencing in plants must cope with both local and systemic invasion of viruses. In a recent study using 2 sets of different experiments, we clearly demonstrated the differential requirement for Dicer-like 4 (DCL4) and DCL2 proteins in the inhibition of intracellular and systemic infection by potato virus X in Arabidopsis thaliana. Taken together with the results of other studies, here we further discuss the functional specificity of DCL proteins in the antiviral silencing pathway.     

The boundary-expressed EPIDERMAL PATTERNING FACTOR-LIKE2 gene encoding a signaling peptide promotes cotyledon growth during Arabidopsis thaliana embryogenesis

The shoot organ boundaries have important roles in plant growth and morphogenesis. It has been reported that a gene encoding a cysteine-rich secreted peptide of the EPIDERMAL PATTERNING FACTOR-LIKE (EPFL) family, EPFL2, is expressed in the boundary domain between the two cotyledon primordia of Arabidopsis thaliana embryo. However, its developmental functions remain unknown. This study aimed to analyze the role of EPFL2 during embryogenesis. We found that cotyledon growth was reduced in its loss-of-function mutants, and this phenotype was associated with the reduction of auxin response peaks at the tips of the primordia. The reduced cotyledon size of the mutant embryo recovered in germinating seedlings, indicating the presence of a factor that acted redundantly with EPFL2 to promote cotyledon growth in late embryogenesis. Our analysis suggests that the boundary domain between the cotyledon primordia acts as a signaling center that organizes auxin response peaks and promotes cotyledon growth.     

Cell division and morphological changes in the shoot apex of Arabidopsis thaliana during floral transition

Eight-week-old vegetative plants of Arabidopsis thaliana, ecotype Columbia, were induced to flower by a single long day (LD). In this experimental system, it is known that the last component of the floral stimulus moves from the leaves to the apex 24-36 h after the start of the LD, and the first floral meristem is initiated by the shoot apical meristem (SAM) at 44-56 h (Corbesier et al., 1996, The Plant Journal 9: 947-952). Here we show that the rate of cell division is increased at floral transition in all SAM parts but not in the sub-apical pith cells. Mitotic activity starts to increase 24 h after the start of the LD and is two- to three-fold higher at peak times than that in non-induced plants. This activation is followed by the start of SAM enlargement at 44 h, SAM doming at 48 h, and the elongation of apical internodes (bolting) at 52 h.