PHYB ACTIVATION TAGGED SUPPRESSOR 1 (BAS1) and SUPPRESSOR OF PHYB‐4 7 (SOB7) are two cytochrome P450 enzymes that inactivate brassinosteroids (BRs) in Arabidopsis. The NAC transcription factor (TF) ...ATAF2 (ANAC081) and the core circadian clock regulator CIRCADIAN CLOCK ASSOCIATED 1 (CCA1) both suppress the expression of BAS1 and SOB7 via direct promoter binding. Additionally, BRs cause feedback suppression on ATAF2 expression. Here, we report that two ATAF‐subgroup TFs, ANAC102 and ATAF1 (ANAC002), also contribute to the transcriptional suppression of BAS1 and SOB7. ANAC102 and ATAF1 gene‐knockout mutants exhibit elevated expression of both BAS1 and SOB7, expanded tissue‐level accumulation of their protein products and reduced hypocotyl growth in response to exogenous BR treatments. Similar to ATAF2, both ANAC102 and ATAF1 are transcriptionally suppressed by BRs and white light. Neither BAS1 nor SOB7 expression is further elevated in ATAF double or triple mutants, suggesting that the suppression effect of these three ATAFs is not additive. In addition, ATAF single, double, and triple mutants have similar levels of BR responsiveness with regard to hypocotyl elongation. ATAF2, ANAC102, ATAF1, and CCA1 physically interact with itself and each other, suggesting that they may coordinately suppress BAS1 and SOB7 expression via protein–protein interactions. Despite the absence of CCA1‐binding elements in their promoters, ANAC102 and ATAF1 have similar transcript circadian oscillation patterns as that of CCA1, suggesting that these two ATAF genes may be indirectly regulated by the circadian clock.
The core circadian regulator CCA1 is a direct repressor of brassinosteroid-inactivating genes BAS1 and SOB7, and interacts with another repressor, ATAF2. Their differential suppressing effects are ...regulated by light.
Abstract
Brassinosteroids (BRs) are a group of steroid hormones regulating plant growth and development. Since BRs do not undergo transport among plant tissues, their metabolism is tightly regulated by transcription factors (TFs) and feedback loops. BAS1 (CYP734A1, formerly CYP72B1) and SOB7 (CYP72C1) are two BR-inactivating cytochrome P450s identified in Arabidopsis thaliana. We previously found that a TF ATAF2 (ANAC081) suppresses BAS1 and SOB7 expression by binding to the Evening Element (EE) and CIRCADIAN CLOCK ASSOCIATED 1 (CCA1)-binding site (CBS) on their promoters. Both the EE and CBS are known binding targets of the circadian regulatory protein CCA1. Here, we confirm that CCA1 binds the EE and CBS motifs on BAS1 and SOB7 promoters, respectively. Elevated accumulations of BAS1 and SOB7 transcripts in the CCA1 null mutant cca1-1 indicate that CCA1 is a repressor of their expression. When compared with either cca1-1 or the ATAF2 null mutant ataf2-2, the cca1-1 ataf2-2 double mutant shows higher SOB7 transcript accumulations and a stronger BR-insensitive phenotype of hypocotyl elongation in white light. CCA1 interacts with ATAF2 at both DNA–protein and protein–protein levels. ATAF2, BAS1, and SOB7 are all circadian regulated with distinct expression patterns. These results demonstrate that CCA1 and ATAF2 differentially suppress BAS1- and SOB7-mediated BR inactivation.
Family-centered care (FCC) is a partnership approach to health care decision-making between the family and health care provider. FCC is considered the standard of pediatric health care by many ...clinical practices, hospitals, and health care groups. Despite widespread endorsement, FCC continues to be insufficiently implemented into clinical practice. In this paper we enumerate the core principles of FCC in pediatric health care, describe recent advances applying FCC principles to clinical practice, and propose an agenda for practitioners, hospitals, and health care groups to translate FCC into improved health outcomes, health care delivery, and health care system transformation.
The Arabidopsis thaliana hypocotyl is a robust system for studying the interplay of light and plant hormones, such as brassinosteroids (BRs), in the regulation of plant growth and development. Since ...BRs cannot be transported between plant tissues, their cellular levels must be appropriate for given developmental fates. BR homeostasis is maintained in part by transcriptional feedback regulation loops that control the expression of key metabolic enzymes, including the BR-inactivating enzymes BAS1 (CYP734A1, formerly CYP72B1) and SOB7 (CYP72C1). Here, we find that the NAC transcription factor (TF) ATAF2 binds the promoters of BAS1 and SOB7 to suppress their expression. ATAF2 restricts the tissue-specific expression of BAS1 and SOB7 in planta. ATAF2 loss- and gain-of-function seedlings have opposite BR-response phenotypes for hypocotyl elongation. ATAF2 modulates hypocotyl growth in a light-dependent manner, with the photoreceptor phytochrome A playing a major role. The photomorphogenic phenotypes of ATAF2 loss- and gain-of-function seedlings are suppressed by treatment with the BR biosynthesis inhibitor brassinazole. Moreover, the disruption of BAS1 and SOB7 abolishes the short-hypocotyl phenotype of ATAF2 loss-of-function seedlings in low fluence rate white light, demonstrating an ATAF2-mediated connection between BR catabolism and photomorphogenesis. ATAF2 expression is suppressed by both BRs and light, which demonstrates the existence of an ATAF2-BAS1/SOB7-BR-ATAF2 feedback regulation loop, as well as a light-ATAF2-BAS1/SOB7-BR-photomorphogenesis pathway. ATAF2 also modulates root growth by regulating BR catabolism. As it is known to regulate plant defense and auxin biosynthesis, ATAF2 therefore acts as a central regulator of plant defense, hormone metabolism and light-mediated seedling development.
Recently, it was found that 1% Phytagel plates used to conduct Arabidopsis thaliana seedling phenotypic analysis no longer reproduced previously published results. This Phytagel, which is produced in ...China (Phytagel C), has replace American-made Phytagel (Phytagel), which is no longer commercially available. In this study, we present the impact of Phytagel produced in the United States vs. China on seedling phenotypic analysis. As a part of this study, an alternative gelling agent has been identified that is capable of reproducing previously published seedling morphometrics.
Phytagel and Phytagel C were investigated based on their ability to reproduce the subtle phenotype of the sob3-4 esc-8 double mutant. Fluence-rate-response analysis of seedlings grown on 1% Phytagel C plates failed to replicate the sob3-4 esc-8 subtle phenotype seen on 1% Phytagel. Furthermore, root penetrance analysis showed a significant difference between sob3-4 esc-8 seedlings grown on 1% Phytagel and 1% Phytagel C. It was also found that 1% Phytagel C was significantly harder than 1% Phytagel. As a replacement for Phytagel C, Gellan was tested. 1% Gellan was able to reproduce the subtle phenotype of sob3-4 esc-8. Furthermore, there was no significant difference in root penetration of the wild type or sob3-4 esc-8 seedlings between 1% Phytagel and 1% Gellan. This may be due to the significant reduction in hardness in 1% Gellan plates compared to 1% Phytagel plates. Finally, we tested additional concentrations of Gellan and found that seedlings on 0.6% Gellan looked more uniform while also being able to reproduce previously published results.
Phytagel has been the standard gelling agent for several studies involving the characterization of subtle seedling phenotypes. After production was moved to China, Phytagel C was no longer capable of reproducing these previously published results. An alternative gelling agent, Gellan, was able to reproduce previously published seedling phenotypes at both 1% and 0.6% concentrations. The information provided in this manuscript is beneficial to the scientific community as whole, specifically phenomics labs, as it details key problematic differences between gelling agents that should be performing identically (Phytagel and Phytagel C).
Gestures that accompany speech are an essential part of natural and efficient embodied human communication. The automatic generation of such co‐speech gestures is a long‐standing problem in computer ...animation and is considered an enabling technology for creating believable characters in film, games, and virtual social spaces, as well as for interaction with social robots. The problem is made challenging by the idiosyncratic and non‐periodic nature of human co‐speech gesture motion, and by the great diversity of communicative functions that gestures encompass. The field of gesture generation has seen surging interest in the last few years, owing to the emergence of more and larger datasets of human gesture motion, combined with strides in deep‐learning‐based generative models that benefit from the growing availability of data. This review article summarizes co‐speech gesture generation research, with a particular focus on deep generative models. First, we articulate the theory describing human gesticulation and how it complements speech. Next, we briefly discuss rule‐based and classical statistical gesture synthesis, before delving into deep learning approaches. We employ the choice of input modalities as an organizing principle, examining systems that generate gestures from audio, text and non‐linguistic input. Concurrent with the exposition of deep learning approaches, we chronicle the evolution of the related training data sets in terms of size, diversity, motion quality, and collection method (e.g., optical motion capture or pose estimation from video). Finally, we identify key research challenges in gesture generation, including data availability and quality; producing human‐like motion; grounding the gesture in the co‐occurring speech in interaction with other speakers, and in the environment; performing gesture evaluation; and integration of gesture synthesis into applications. We highlight recent approaches to tackling the various key challenges, as well as the limitations of these approaches, and point toward areas of future development.
The Arabidopsis thaliana genome encodes 29 AT-HOOK MOTIF CONTAINING NUCLEAR LOCALIZED (AHL) genes, which evolved into two phylogenic clades. The AHL proteins contain one or two AT-hook motif(s) and ...one plant and prokaryote conserved (PPC)/domain of unknown function #296 (DUF296) domain. Seedlings lacking both SOB3/AHL29 and ESC/AHL27 confer a subtle long-hypocotyl phenotype compared with the WT or either single-null mutant. In contrast, the missense allele sob3-6 confers a dramatic long-hypocotyl phenotype in the light. In this study, we examined the dominant-negative feature of sob3-6 and found that it encodes a protein with a disrupted AT-hook motif that abolishes binding to AT-rich DNA. A loss-of-function approach demonstrated different, yet redundant, contributions of additional AHL genes in suppressing hypocotyl elongation in the light. We showed that AHL proteins interact with each other and themselves via the PPC/DUF296 domain. AHLs also share interactions with other nuclear proteins, such as transcription factors, suggesting that these interactions also contribute to the functional redundancy within this gene family. The coordinated action of AHLs requires an AT-hook motif capable of binding AT-rich DNA, as well as a PPC/DUF296 domain containing a conserved Gly-Arg-Phe-Glu-Ile-Leu region. Alteration of this region abolished SOB3/AHL29’s physical interaction with transcription factors and resulted in a dominant-negative allele in planta that was phenotypically similar to sob3-6 . We propose a molecular model where AHLs interact with each other and themselves, as well as other nuclear proteins, to form complexes which modulate plant growth and development.
Summary
PCR‐based detection of single nucleotide polymorphisms is a powerful tool for the plant geneticist. Cleaved amplified polymorphic sequence analysis is the most widely used approach for the ...detection of single nucleotide polymorphisms. However, this technique is limited to mutations which create or disrupt a restriction enzyme recognition site. This paper presents a modification of this technique where mismatches in a PCR primer are used to create a polymorphism based on the target mutation. This technique is useful for following known mutations in segregating populations and genetic mapping of isolated DNAs used for positional based cloning of new genes. In addition, a computer program has been developed that facilitates the design of these PCR primers.
The detection of single nucleotide polymorphisms by PCR is necessary for many types of genetic analysis, from mapping genomes to tracking specific mutations. This technique is most commonly used when ...polymorphisms alter restriction endonuclease recognition sites. Here we describe a web-based program, dCAPS Finder 2.0, that facilitates the design of mismatched PCR primers to create or remove a restriction endonuclease recognition site relative to the polymorphism being analyzed.
The detection of single nucleotide polymorphisms by PCR is necessary for many types of genetic analysis, from mapping genomes to tracking specific mutations. A web-based program, dCAPS Finder 2.0, facilitates the design of mismatched PCR primers to aid such analysis.
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