In plants and animals, small peptide ligands that signal in cell-cell communication have been suggested to be a crucial component of development. A bioassay of single-cell transdifferentation demonstrates that a dodecapeptide with two hydroxyproline residues is the functional product of genes from the CLE family, which includes CLAVATA3 in Arabidopsis. The dodecapeptide suppresses xylem cell development at a concentration of 10(-11) M and promotes cell division. An application, corresponding to all 26 Arabidopsis CLE protein family members, of synthetic dodecapeptides reveals two counteracting signaling pathways involved in stem cell fate.

Inductive cell-cell interactions are essential for controlling cell fate determination in both plants and animals; however, the chemical basis of inductive signals in plants remains little understood. A proteoglycan-like factor named xylogen mediates local and inductive cell-cell interactions required for xylem differentiation in Zinnia cells cultured in vitro. Here we describe the purification of xylogen and cloning of its complementary DNA, and present evidence for its role in planta. The polypeptide backbone of xylogen is a hybrid-type molecule with properties of both arabinogalactan proteins and nonspecific lipid-transfer proteins. Xylogen predominantly accumulates in the meristem, procambium and xylem. In the xylem, xylogen has a polar localization in the cell walls of differentiating tracheary elements. Double knockouts of Arabidopsis lacking both genes that encode xylogen proteins show defects in vascular development: discontinuous veins, improperly interconnected vessel elements and simplified venation. Our results suggest that the polar secretion of xylogen draws neighbouring cells into the pathway of vascular differentiation to direct continuous vascular development, thereby identifying a molecule that mediates an inductive cell-cell interaction involved in plant tissue differentiation.

Edited by Shou-Wei DingKeywords: RNA silencing Suppressor of gene silencing 3 RNA-dependent RNA polymerase 6 Dicer-like 4 Processing body Arabidopsis a b s t r a c t Suppressor of gene silencing 3 (SGS3) is involved in RNA-dependent RNA polymerase 6 (RDR6)-dependent small-interfering RNA (siRNA) pathways in Arabidopsis. However, the roles of SGS3 in those pathways are unclear. Here, we show that SGS3 interacts and colocalizes with RDR6 in cytoplasmic granules. Interestingly, the granules containing SGS3 and RDR6 (named SGS3/RDR6-bodies) were distinct from the processing bodies where mRNAs are decayed and/or stored. Microscopic analyses and complementation experiments using SGS3-deletion mutants suggested that proper localization of SGS3 is important for its function. These results provide novel insights into RDR6-dependent siRNA formation in plants.

SummaryTo study cellular morphogenesis genetically, we isolated loss-of-function mutants of Arabidopsis thaliana, designated ibo1. The ibo1 mutations cause local outgrowth in the middle of epidermal cells of the hypocotyls and petioles, resulting in the formation of a protuberance. In Arabidopsis, the hypocotyl epidermis differentiates into two alternate cell files, the stoma cell file and the non-stoma cell file, by a mechanism involving TRANSPARENT TESTA GLABRA1 (TTG1) and GLABRA2 (GL2). The ectopic protuberances of the ibo1 mutants were preferentially induced in the non-stoma cell files, which express GL2. TTG1-dependent epidermal patterning is required for protuberance formation in ibo1, suggesting that IBO1 functions downstream from epidermal cell specification. Pharmacological and genetic analyses demonstrated that ethylene promotes protuberance formation in ibo1, implying that IBO1 acts antagonistically to ethylene to suppress radial outgrowth. IBO1 is identical to NEK6, which encodes a Never In Mitosis A (NIMA)-related protein kinase (Nek) with sequence similarity to Neks involved in microtubule organization in fungi, algae, and animals. The ibo1-1 mutation, in which a conserved Glu residue in the activation loop is substituted by Arg, completely abolishes its kinase activity. The intracellular localization of GFP-tagged NEK6 showed that NEK6 mainly accumulates in cytoplasmic spots associated with cortical microtubules and with a putative component of the c-tubulin complex. The localization of NEK6 is regulated by the C-terminal domain, which is truncated in the ibo1-2 allele. These results suggest that the role of NEK6 in the control of cellular morphogenesis is dependent on its kinase action and association with the cortical microtubules.

Although decapping is an important process in eukaryotic mRNA turnover, little is known about this process in plants. Here, we identified Arabidopsis thaliana decapping proteins AtDCP1 and AtDCP2 and showed that (I) AtDCP2 is an active decapping enzyme, (II) AtDCP1 interacts with itself, (III) AtDCP1 and AtDCP2 are localized to cytoplasmic foci (putative Arabidopsis processing body), and (IV) AtDCP1 and AtDCP2 are essential for post-embryonic development. Our findings provide new insights into the role of decapping-dependent mRNA turnover.

The plant hormone auxin is a key morphogenetic signal that controls many aspects of plant growth and development. Cellular auxin levels are coordinately regulated by multiple processes, including auxin biosynthesis and the polar transport and metabolic pathways. The auxin concentration gradient determines plant organ positioning and growth responses to environmental cues. Auxin transport systems play crucial roles in the spatiotemporal regulation of the auxin gradient. This auxin gradient has been analyzed using SCF-type E3 ubiquitin-ligase complex-based auxin biosensors in synthetic auxin-responsive reporter lines. However, the contributions of auxin biosynthesis and metabolism to the auxin gradient have been largely elusive. Additionally, the available information on subcellular auxin localization is still limited. Here we designed fluorescently labeled auxin analogs that remain active for auxin transport but are inactive for auxin signaling and metabolism. Fluorescent auxin analogs enable the selective visualization of the distribution of auxin by the auxin transport system. Together with auxin biosynthesis inhibitors and an auxin biosensor, these analogs indicated a substantial contribution of local auxin biosynthesis to the formation of auxin maxima at the root apex. Moreover, fluorescent auxin analogs mainly localized to the endoplasmic reticulum in cultured cells and roots, implying the presence of a subcellular auxin gradient in the cells. Our work not only provides a useful tool for the plant chemical biology field but also demonstrates a new strategy for imaging the distribution of smallmolecule hormones.auxin transporter | subcellular localization

SUMMARYNimA-related kinase 6 (NEK6) has been implicated in microtubule regulation to suppress the ectopic outgrowth of epidermal cells; however, its molecular functions remain to be elucidated. Here, we analyze the function of NEK6 and other members of the NEK family with regard to epidermal cell expansion and cortical microtubule organization. The functional NEK6-green fluorescent protein fusion localizes to cortical microtubules, predominantly in particles that exhibit dynamic movement along microtubules. The kinasedead mutant of NEK6 (ibo1-1) exhibits a disturbance of the cortical microtubule array at the site of ectopic protrusions in epidermal cells. Pharmacological studies with microtubule inhibitors and quantitative analysis of microtubule dynamics indicate excessive stabilization of cortical microtubules in ibo1/nek6 mutants. In addition, NEK6 directly binds to microtubules in vitro and phosphorylates b-tubulin. NEK6 interacts and co-localizes with NEK4 and NEK5 in a transient expression assay. The ibo1-3 mutation markedly reduces the interaction between NEK6 and NEK4 and increases the interaction between NEK6 and NEK5. NEK4 and NEK5 are required for the ibo1/nek6 ectopic outgrowth phenotype in epidermal cells. These results demonstrate that NEK6 homodimerizes and forms heterodimers with NEK4 and NEK5 to regulate cortical microtubule organization possibly through the phosphorylation of b-tubulins.

Highlights d An hyper-response to mechanical stress can lead to increased phenotypic variability d NEK6 exhibits a bipolar recruitment on microtubules that align with tensile stress d The nek6 wavy phenotype is consistent with a hyperproprioceptive response d Growth rate uncouples shape-and growth-derived stress response in nek6