Project description:The establishment of ad/abaxial polarity is a fundamental event in plant development. It is critical for correct polar development of the leaf (the upper portion of the leaf is chloroplast rich and optimized for light capture while the lower portion is optimized for gas exchange) and for creating an environment that allows the formation of new meristems (centers of stem cell growth). Class III homeodomain-leucine zipper (HD-ZIPIII) proteins are conserved plant proteins that act as potent regulators of ad/abaxial polarity. HD-ZIPIII protein activity promotes the development of upper (adaxial) leaf fates and meristem formation; in its absence lower (abaxial) leaf fates develop and meristems fail to form. A network of regulatory factors controls the establishment of ad/abaxial leaf fates. However, this network is incomplete and how these factors control one another is poorly understood. Here we report a new family of plant genes, the LITTLE ZIPPER (ZPR) genes (consisting principally of a stretch of leucine zipper similar to the leucine zipper in HD-ZIPIII proteins) that are transcriptionally up-regulated by HD-ZIPIII activity. Furthermore, we show that the ZPR proteins interact with and repress HD-ZIPIII activity, thus forming a negative feedback loop. Our results suggest that HD-ZIPIII proteins exist in active homodimers and, together with the ZPR proteins, in inactive heterodimers. The newly discovered HD-ZIPIII/ZPR regulatory module would not only serve to dampen the effect of fluctuations in HD-ZIPIII protein levels but more importantly would provide a point of regulation - control over the ratio of inactive heterodimers to active homodimers - that could be influenced by other components of the pathway. For instance, the binding of a small hydrophobic molecule to the conserved (yet little understood) START domain present in the HD-ZIPIII proteins may influence the type of dimer formed. Published in: Wenkel, S., Emery, J., Hou, B.-H., Evans, M.M.S. and M.K. Barton, 2007, A Feedback Regulatory Module Formed by LITTLE ZIPPER and HD-ZIPIII Genes. Plant Cell 2007 Keywords: screen for downstream genes, transcription factor induction
Project description:A key question in developmental biology is how cells exchange positional information for proper patterning during organ development. In plant roots the radial tissue organization is highly conserved with a central vascular cylinder in which two water conducting cell types, protoxylem and metaxylem, are patterned centripetally. We show that this patterning occurs through crosstalk between the vascular cylinder and the surrounding endodermis mediated by cell-to-cell movement of a transcription factor in one direction and microRNAs in the other. SHORT ROOT, produced in the vascular cylinder, moves into the endodermis to activate SCARECROW. Together these transcription factors activate MIR165a and 166b. Endodermally produced miR165/6 then acts to degrade its target mRNAs encoding class III homeodomain-leucine zipper transcription factors in the endodermis and stele periphery. The resulting differential distribution of target mRNA in the vascular cylinder determines xylem cell types in a dosage dependent manner.
Project description:The establishment of ad/abaxial polarity is a fundamental event in plant development. It is critical for correct polar development of the leaf (the upper portion of the leaf is chloroplast rich and optimized for light capture while the lower portion is optimized for gas exchange) and for creating an environment that allows the formation of new meristems (centers of stem cell growth). Class III homeodomain-leucine zipper (HD-ZIPIII) proteins are conserved plant proteins that act as potent regulators of ad/abaxial polarity. HD-ZIPIII protein activity promotes the development of upper (adaxial) leaf fates and meristem formation; in its absence lower (abaxial) leaf fates develop and meristems fail to form. A network of regulatory factors controls the establishment of ad/abaxial leaf fates. However, this network is incomplete and how these factors control one another is poorly understood. Here we report a new family of plant genes, the LITTLE ZIPPER (ZPR) genes (consisting principally of a stretch of leucine zipper similar to the leucine zipper in HD-ZIPIII proteins) that are transcriptionally up-regulated by HD-ZIPIII activity. Furthermore, we show that the ZPR proteins interact with and repress HD-ZIPIII activity, thus forming a negative feedback loop. Our results suggest that HD-ZIPIII proteins exist in active homodimers and, together with the ZPR proteins, in inactive heterodimers. The newly discovered HD-ZIPIII/ZPR regulatory module would not only serve to dampen the effect of fluctuations in HD-ZIPIII protein levels but more importantly would provide a point of regulation - control over the ratio of inactive heterodimers to active homodimers - that could be influenced by other components of the pathway. For instance, the binding of a small hydrophobic molecule to the conserved (yet little understood) START domain present in the HD-ZIPIII proteins may influence the type of dimer formed. Published in:; Wenkel, S., Emery, J., Hou, B.-H., Evans, M.M.S. and M.K. Barton, 2007, A Feedback Regulatory Module Formed by LITTLE ZIPPER and HD-ZIPIII Genes. Plant Cell 2007 Experiment Overall Design: One of the five Arabidopsis HD-ZIPIII proteins, REVOLUTA, was placed under glucocorticoid control by fusing the glucocorticoid receptor (GR) domain to its amino terminus. This gene fusion was placed under the control of the highly and constitutively expressed viral CaMV35S promoter. Since the HD-ZIPIII genes are controlled by microRNAs, it was also necessary to introduce mutations that disrupted the microRNA complementary sequence. Transgenic Arabidopsis seedlings carrying this construct (GR-REV) as well as wild-type seedlings were grown in liquid culture for ten days. At this point, one flask each of wt and transgenic seedlings was treated with carrier alone (ethanol) and one flask each of wt and transgenic seedlings was treated with dexamethasone. RNA was harvested from these samples after one hour, labeled and hybridized to Affymetrix microarrays.
Project description:A key question in developmental biology is how cells exchange positional information for proper patterning during organ development. In plant roots the radial tissue organization is highly conserved with a central vascular cylinder in which two water conducting cell types, protoxylem and metaxylem, are patterned centripetally. We show that this patterning occurs through crosstalk between the vascular cylinder and the surrounding endodermis mediated by cell-to-cell movement of a transcription factor in one direction and microRNAs in the other. SHORT ROOT, produced in the vascular cylinder, moves into the endodermis to activate SCARECROW. Together these transcription factors activate MIR165a and 166b. Endodermally produced miR165/6 then acts to degrade its target mRNAs encoding class III homeodomain-leucine zipper transcription factors in the endodermis and stele periphery. The resulting differential distribution of target mRNA in the vascular cylinder determines xylem cell types in a dosage dependent manner. To investigate the global regulation of SHR and SCR in the root, direct and indirect target genes of SHR and SCR were cross-compared with selected cell type specific expression profiles in the Arabidopsis root published in Birnbaum et al. (2003), Nawy et al. (2005), Lee et al. (2006), and Brady et al. (2007) [pubmed IDs: 14671301, 15937229, 16581911, 17975066]. Furthermore, the mature endodermis specific transcript profiles were generated using a GFP marker line, E30, and used together with published profiles in this comparison.
Project description:Unlike the situation in animals, the final morphology of the plant body is highly modulated by the environment. During Arabidopsis development, intrinsic factors provide the framework for basic patterning processes. CLASS III HOMEODOMAIN LEUCINE ZIPPER (HD‐ZIPIII) transcription factors are involved in embryo, shoot and root patterning and during vegetative growth regulate several polarity set‐up processes such as in leaves and the vascular system. Here we show that besides being involved in basic patterning, HD‐ZIPIII transcription factors also have a critical role in controlling elongation growth that is induced when plants experience shade. By using a ChIP‐Seq approach, we have identified several direct target genes of the HD‐ZIPIII transcription factor REVOLUTA (REV). We show that REV acts upstream of auxin biosynthesis and directly regulates several HAT transcription factors that control shade avoidance responses in Arabidopsis. Plants in which HD‐ZIPIII genes are mutated show altered responses to shade revealing that the basic patterning machinery also regulates adaptive development. Thus, HD-ZIPIII transcription factors contribute to shade signaling and act upstream of both auxin and HAT activation. A. thaliana REVOLUTA ChIP-seq w control
Project description:Unlike the situation in animals, the final morphology of the plant body is highly modulated by the environment. During Arabidopsis development, intrinsic factors provide the framework for basic patterning processes. CLASS III HOMEODOMAIN LEUCINE ZIPPER (HD‐ZIPIII) transcription factors are involved in embryo, shoot and root patterning and during vegetative growth regulate several polarity set‐up processes such as in leaves and the vascular system. Here we show that besides being involved in basic patterning, HD‐ZIPIII transcription factors also have a critical role in controlling elongation growth that is induced when plants experience shade. By using a ChIP‐Seq approach, we have identified several direct target genes of the HD‐ZIPIII transcription factor REVOLUTA (REV). We show that REV acts upstream of auxin biosynthesis and directly regulates several HAT transcription factors that control shade avoidance responses in Arabidopsis. Plants in which HD‐ZIPIII genes are mutated show altered responses to shade revealing that the basic patterning machinery also regulates adaptive development. Thus, HD-ZIPIII transcription factors contribute to shade signaling and act upstream of both auxin and HAT activation.
Project description:The class III HD-ZIPtranscription factors regulate vascular patterning in Arabidopsis thaliana roots. In this expression study we compare the expression profile in root tips upon miR165 induction, after 6h, 10h and 24h. The results are presented in PHABULOSA mediates an auxin signaling loop to regulate vascular patterning in Arabidopsis by Christina Joy Müller, Ana Elisa Valdés, Guodong Wang, Prashanth Ramachandran, Lisa Beste, Daniel Uddenberg, and Annelie Carlsbecker, accepted for publication in Plant Physiology Nov. 2015. Plant vascular tissues, xylem and phloem, differentiate in distinct patterns from procambial cells as an integral transport system for water, sugars and signaling molecules. Procambium formation is promoted by high auxin levels activating class III homeodomain leucine zipper (HD-ZIP III) transcription factors (TFs). In the root of Arabidopsis thaliana, HD-ZIP III TFs dose-dependently govern the patterning of the xylem axis, with higher levels promoting metaxylem cell identity in the central axis and lower levels protoxylem at its flanks. It is, however, unclear by what mechanisms the HD-ZIP III TFs control xylem axis patterning. Here we present data suggesting that an important mechanism is their ability to moderate auxin response. We found that changes in HD-ZIP III TF levels affect the expression of genes encoding core auxin response molecules. We show that one of the HD-ZIP III TFs, PHABULOSA, directly binds the promoter of both MONOPTEROS/AUXIN RESPONSE FACTOR5 (MP/ARF5), a key factor in vascular formation, and IAA20, encoding an AUX/IAA protein which is stable in the presence of auxin and able to interact with and repress MP activity. The double mutant of IAA20 and its closest homologue IAA30 forms ectopic protoxylem, while overexpression of IAA30 causes discontinuous protoxylem and occasional ectopic metaxylem, similar to a weak loss-of-function mp-mutant. Our results provide evidence that HD-ZIP III TFs directly affect auxin response and mediate a feed forward loop formed by MP and IAA20 that may focus and stabilize auxin response during vascular patterning and differentiation of xylem cell types.
Project description:The shoot apical meristem (SAM) comprises a group of undifferentiated cells that divide to maintain the plant meristem and also give rise to all shoot organs. SAM fate is specified by class III HOMEODOMAIN-LEUCINE ZIPPER (HD-ZIP III) transcription factors, which are targets of miR166/165. In Arabidopsis, AGO10 is a critical regulator of SAM maintenance, and here we demonstrate that AGO10 specifically interacts with miR166/165. The association is determined by a distinct structure of the miR166/165 duplex. Deficient loading of miR166 into AGO10 results in a defective SAM. Notably, the miRNA-binding ability of AGO10, but not its catalytic activity, is required for SAM development, and AGO10 has a higher binding affinity for miR166 than does AGO1, a principal contributor to miRNA-mediated silencing. We propose that AGO10 functions as a decoy for miR166/165 to maintain the SAM, preventing their incorporation into AGO1 complexes and the subsequent repression of HD-ZIP III gene expression.
Project description:In addition to apical growth, plants undergo radial growth to increase girth, a particularly prominent feature in trees. During both apical and radial growth the phytohormones auxin and cytokinins form symmetries that govern further growth patterns. But whereas the gene regulatory networks interpreting the hormonal fields during apical growth are well established, such networks are not known for the radial growth. We show here that the initiation of radial growth occurs around early protophloem sieve element (PSE) cell files of the root procambial tissue in Arabidopsis. In this domain cytokinin signalling promotes expression of a pair of novel mobile transcription factors, PHLOEM EARLY DOF (PEAR1, PEAR2) and their four homologs (OBP2, DOF6, TMO6 and HCA2), collectively called PEAR proteins. The PEAR proteins form a short-range concentration gradient peaking at PSE and activating gene expression that promotes radial growth. The expression and function of PEAR proteins are antagonized by well-established polarity transcription factors, HD-ZIP III, whose expression is concentrated in the more internal domain of radially non-dividing procambial cells by the function of auxin and mobile miR165/166. The PEAR proteins locally promote transcription of their inhibitory HD-ZIP III genes, thereby establishing a negative feedback loop that forms a sharp boundary demarking the zone of cell divisions. Taken together, we have established a network, in which the PEAR - HD-ZIP III module integrates the spatial information of the hormonal domains and miRNA gradients during root procambial development, to provide a pre-pattern with actively dividing and more quiescent zones, thus priming radial growth.