Slow release of a synthetic auxin induces formation of adventitious roots in recalcitrant woody plants.

Clonal propagation of plants by induction of adventitious roots (ARs) from stem cuttings is a requisite step in breeding programs. A major barrier exists for propagating valuable plants that naturally have low capacity to form ARs. Due to the central role of auxin in organogenesis, indole-3-butyric acid is often used as part of commercial rooting mixtures, yet many recalcitrant plants do not form ARs in response to this treatment. Here we describe the synthesis and screening of a focused library of synthetic auxin conjugates in Eucalyptus grandis cuttings and identify 4-chlorophenoxyacetic acid-L-tryptophan-OMe as a competent enhancer of adventitious rooting in a number of recalcitrant woody plants, including apple and argan. Comprehensive metabolic and functional analyses reveal that this activity is engendered by prolonged auxin signaling due to initial fast uptake and slow release and clearance of the free auxin 4-chlorophenoxyacetic acid. This work highlights the utility of a slow-release strategy for bioactive compounds for more effective plant growth regulation.

Chemical induction of hypocotyl rooting reveals extensive conservation of auxin signalling controlling lateral and adventitious root formation.

Upon exposure to light, etiolated Arabidopsis seedlings form adventitious roots (AR) along the hypocotyl. While processes underlying lateral root formation are studied intensively, comparatively little is known about the molecular processes involved in the initiation of hypocotyl AR. AR and LR formation were studied using a small molecule named Hypocotyl Specific Adventitious Root INducer (HYSPARIN) that strongly induces AR but not LR formation. HYSPARIN does not trigger rapid DR5-reporter activation, DII-Venus degradation or Ca2+ signalling. Transcriptome analysis, auxin signalling reporter lines and mutants show that HYSPARIN AR induction involves nuclear TIR1/AFB and plasma membrane TMK auxin signalling, as well as multiple downstream LR development genes (SHY2/IAA3, PUCHI, MAKR4 and GATA23). Comparison of the AR and LR induction transcriptome identified SAURs, AGC kinases and OFP transcription factors as specifically upregulated by HYSPARIN. Members of the SAUR19 subfamily, OFP4 and AGC2 suppress HYS-induced AR formation. While SAUR19 and OFP subfamily members also mildly modulate LR formation, AGC2 regulates only AR induction. Analysis of HYSPARIN-induced AR formation uncovers an evolutionary conservation of auxin signalling controlling LR and AR induction in Arabidopsis seedlings and identifies SAUR19, OFP4 and AGC2 kinase as novel regulators of AR formation.

Intrinsic disorder and conformational coexistence in auxin coreceptors.

AUXIN/INDOLE 3-ACETIC ACID (Aux/IAA) transcriptional repressor proteins and the TRANSPORT INHIBITOR RESISTANT 1/AUXIN SIGNALING F-BOX (TIR1/AFB) proteins to which they bind act as auxin coreceptors. While the structure of TIR1 has been solved, structural characterization of the regions of the Aux/IAA protein responsible for auxin perception has been complicated by their predicted disorder. Here, we use NMR, CD and molecular dynamics simulation to investigate the N-terminal domains of the Aux/IAA protein IAA17/AXR3. We show that despite the conformational flexibility of the region, a critical W-P bond in the core of the Aux/IAA degron motif occurs at a strikingly high (1:1) ratio of cis to trans isomers, consistent with the requirement of the cis conformer for the formation of the fully-docked receptor complex. We show that the N-terminal half of AXR3 is a mixture of multiple transiently structured conformations with a propensity for two predominant and distinct conformational subpopulations within the overall ensemble. These two states were modeled together with the C-terminal PB1 domain to provide the first complete simulation of an Aux/IAA. Using MD to recreate the assembly of each complex in the presence of auxin, both structural arrangements were shown to engage with the TIR1 receptor, and contact maps from the simulations match closely observations of NMR signal-decreases. Together, our results and approach provide a platform for exploring the functional significance of variation in the Aux/IAA coreceptor family and for understanding the role of intrinsic disorder in auxin signal transduction and other signaling systems.

Are Patients With Morbid Obesity at Increased Risk of Pulmonary Embolism or Proximal Deep Vein Thrombosis After Lower Limb Arthroplasty? A Large-database Study.

BACKGROUND: Whether increased BMI is associated with an increased risk of venous thromboembolism (VTE) is controversial. Despite this, BMI > 40 kg/m 2 remains a common cutoff for lower limb arthroplasty eligibility. Current United Kingdom national guidelines list obesity as a risk factor for VTE, but these are based on evidence that has largely failed to differentiate between potentially minor (distal deep vein thrombosis [DVT]), and more harmful (pulmonary embolism [PE] and proximal DVT) diagnoses. Determining the association between BMI and the risk of clinically important VTE is needed to improve the utility of national risk stratification tools. QUESTIONS/PURPOSES: (1) In patients undergoing lower limb arthroplasty, is BMI 40 kg/m 2 or higher (morbid obesity) associated with an increased risk of PE or proximal DVT within 90 days of surgery, compared with patients with BMI less than 40 kg/m 2 ? (2) What proportion of investigations ordered for PE and proximal DVT were positive in patients with morbid obesity who underwent lower limb arthroplasty compared with those with BMI less than 40 kg/m 2 ? METHODS: Data were collected retrospectively from the Northern Ireland Electronic Care Record, a national database recording patient demographics, diagnoses, encounters, and clinical correspondence. Between January 2016 and December 2020, 10,217 primary joint arthroplasties were performed. Of those, 21% (2184 joints) were excluded; 2183 were in patients with multiple arthroplasties and one had no recorded BMI. All 8033 remaining joints were eligible for inclusion, 52% of which (4184) were THAs, 44% (3494) were TKAs, and 4% (355) were unicompartmental knee arthroplasties; all patients had 90 days of follow-up. The Wells score was used to guide the investigations. Indications for CT pulmonary angiography for suspected PE included pleuritic chest pain, reduced oxygen saturations, dyspnea, or hemoptysis. Indications for ultrasound scans for suspected proximal DVT included leg swelling, pain, warmth, or erythema. Distal DVTs were recorded as negative scans because we do not treat them with modified anticoagulation. The division of categories was set at BMI 40 kg/m 2 , a common clinical cutoff used in surgical eligibility algorithms. Patients were grouped according to WHO BMI categories to assess for the following confounding variables: sex, age, American Society of Anesthesiologists grade, joint replaced, VTE prophylaxis, grade of operative surgeon, and implant cement status. RESULTS: We found no increase in the odds of PE or proximal DVT in any WHO BMI category. When comparing patients with BMI less than 40 kg/m 2 with those with a BMI of 40 kg/m 2 or higher, there was no difference in the odds of PE (0.8% [58 of 7506] versus 0.8% [four of 527]; OR 1.0 [95% CI 0.4 to 2.8]; p > 0.99) or proximal DVT (0.4% [33 of 7506] versus 0.2% [one of 527]; OR 2.3 [95% CI 0.3 to 17.0]; p = 0.72). Of those who received diagnostic imaging, 21% (59 of 276) of CT pulmonary angiograms and 4% (34 of 718) of ultrasounds were positive for patients with BMI less than 40 kg/m 2 compared with 14% (four of 29; OR 1.6 [95% CI 0.6 to 4.5]; p = 0.47) and 2% (one of 57; OR 2.7 [95% CI 0.4 to 18.6]; p = 0.51) for patients with BMI 40 kg/m 2 or higher. There was no difference in the percentage of CT pulmonary angiograms ordered (4% [276 of 7506] versus 5% [29 of 527]; OR 0.7 [95% CI 0.5 to 1.0]; p = 0.07) or ultrasounds ordered (10% [718 of 7506] versus 11% [57 of 527]; OR 0.9 [95% CI 0.7 to 1.2]; p = 0.49) for BMI less than 40 kg/m 2 and BMI 40 kg/m 2 or higher. CONCLUSION: Increased BMI should not preclude individuals from lower limb arthroplasty based on suspected risk of clinically important VTE. National VTE risk stratification tools should be based on evidence assessing clinically relevant VTE (specifically, proximal DVT, PE, or death of thromboembolism) only. LEVEL OF EVIDENCE: Level III, therapeutic study.

PIN structures shed light on their mechanism of auxin efflux.

Polar auxin transport is a quintessential feature of higher plant physiology and it has been known for many years that some of the primary drivers of polar auxin transport are the PIN-formed (PIN) auxin efflux proteins. Formative research established many key biochemical features of the transport system and discovered inhibitors such as 1-naphthylphthalamic acid (NPA), but the mechanism of action of PINs has remained elusive. This changed in 2022 with the publication of high-resolution structures of the membrane-spanning domains of three PIN proteins. The atomic structures and associated activity assays reveal that PINs use an elevator mechanism to transport auxin anions out of the cell. NPA was shown to be a competitive inhibitor that traps PINs in their inward-open conformation. The secrets of the hydrophilic cytoplasmic loop of PIN proteins remain to be discovered.

Non-specific effects of the CINNAMATE-4-HYDROXYLASE inhibitor piperonylic acid.

Chemical inhibitors are often implemented for the functional characterization of genes to overcome the limitations associated with genetic approaches. Although it is well established that the specificity of the compound is key to success of a pharmacological approach, off-target effects are often overlooked or simply neglected in a complex biological setting. Here we illustrate the cause and implications of such secondary effects by focusing on piperonylic acid (PA), an inhibitor of CINNAMATE-4-HYDROXYLASE (C4H) that is frequently used to investigate the involvement of lignin during plant growth and development. When supplied to plants, we found that PA is recognized as a substrate by GRETCHEN HAGEN 3.6 (GH3.6), an amido synthetase involved in the formation of the indole-3-acetic acid (IAA) conjugate IAA-Asp. By competing for the same enzyme, PA interferes with IAA conjugation, resulting in an increase in IAA concentrations in the plant. In line with the broad substrate specificity of the GH3 family of enzymes, treatment with PA increased not only IAA levels but also those of other GH3-conjugated phytohormones, namely jasmonic acid and salicylic acid. Finally, we found that interference with the endogenous function of GH3s potentially contributes to phenotypes previously observed upon PA treatment. We conclude that deregulation of phytohormone homeostasis by surrogate occupation of the conjugation machinery in the plant is likely a general phenomenon when using chemical inhibitors. Our results hereby provide a novel and important basis for future reference in studies using chemical inhibitors.

Step-Growth Glycopolymers with a Defined Tacticity for Selective Carbohydrate-Lectin Recognition.

Glycopolymers are potent candidates for biomedical applications by exploiting multivalent carbohydrate-lectin interactions. Owing to their specific recognition capabilities, glycosylated polymers can be utilized for targeted drug delivery to certain cell types bearing the corresponding lectin receptors. A fundamental challenge in glycopolymer research, however, is the specificity of recognition to receptors binding to the same sugar unit (e.g., mannose). Variation of polymer backbone chirality has emerged as an effective method to distinguish between lectins on a molecular level. Herein, we present a facile route toward producing glycopolymers with a defined tacticity based on a step-growth polymerization technique using click chemistry. A set of polymers have been fabricated and further functionalized with mannose moieties to enable lectin binding to receptors relevant to the immune system (mannose-binding lectin, dendritic cell-specific intercellular adhesion molecule-3-grabbing non-integrin, and dendritic and thymic epithelial cell-205). Surface plasmon resonance spectrometry was employed to determine the kinetic parameters of the step-growth glycopolymers. The results highlight the importance of structural complexity in advancing glycopolymer synthesis, yet multivalency remains a main driving force in lectin recognition.

Nucleic acid aptamers as aptasensors for plant biology.

Our knowledge of cell- and tissue-specific quantification of phytohormones is heavily reliant on laborious mass spectrometry techniques. Genetically encoded biosensors have allowed spatial and some temporal quantification of phytohormones intracellularly, but there is still limited information on their intercellular distributions. Here, we review nucleic acid aptamers as an emerging biosensing platform for the detection and quantification of analytes with high affinity and specificity. Options for DNA aptamer technology are explained through selection, sequencing analysis and techniques for evaluating affinity and specificity, and we focus on previously developed DNA aptamers against various plant analytes. We suggest how these tools might be applied in planta for quantification of molecules of interest both intracellularly and intercellularly.

The differential binding and biological efficacy of auxin herbicides.

BACKGROUND: Auxin herbicides have been used for selective weed control for 75 years and they continue to be amongst the most widely used weed control agents globally. The auxin herbicides fall into five chemical classes, with two herbicides not classified, and in all cases it is anticipated that recognition in the plant starts with binding to the Transport Inhibitor Response 1 (TIR1) family of auxin receptors. There is evidence that some classes of auxins act selectively with certain clades of receptors, although a comprehensive structure-activity relationship has not been available. RESULTS: Using purified receptor proteins to measure binding efficacy we have conducted quantitative structure activity relationship (qSAR) assays using representative members of the three receptor clades in Arabidopsis, TIR1, AFB2 and AFB5. Complementary qSAR data for biological efficacy at the whole-plant level using root growth inhibition and foliar phytotoxicity assays have also been analyzed for each family of auxin herbicides, including for the afb5-1 receptor mutant line. CONCLUSIONS: Comparisons of all these assays highlight differences in receptor selectivity and some systematic differences between results for binding in vitro and activity in vivo. The results could provide insights into weed spectrum differences between the different classes of auxin herbicides, as well as the potential resistance and cross-resistance implications for this herbicide class. © 2022 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Is Cementless Total Knee Arthroplasty Safe in Women Over 75 Y of Age?

BACKGROUND: Cementless total knee arthroplasty (TKA) is the subject of renewed interest. Previous concerns about survivorship have been addressed and there is an appeal in terms of biological fixation and surgical efficiency. However, even surgeon advocates have concerns about the risk of marked subsidence when using this technology in older patients at risk for osteoporosis. METHODS: This was a retrospective analysis of 1,000 consecutive fully cementless mobile bearing TKAs performed at a single institution on women over 75 years of age who had postoperative and 1-year x-rays. The primary outcome was the incidence of subsidence. RESULTS: There were three asymptomatic cases with definite subsidence and change in alignment. In a fourth symptomatic case, the femoral component subsided into varus and the tibia into valgus, thus maintaining alignment which facilitated nonoperative treatment in a 92-year-old. Overall, at 1 year, there were two- liner revisions for infection without recurrence. Five patients had further surgery, of which three were washouts and two were for periprosthetic fractures sustained postoperatively within 1 year. Seven patients had further anesthesia, of which five were manipulations and two were nonrecurrent closed reductions for spinouts. CONCLUSION: Cementless TKA did not have a high risk of subsidence in this at-risk population. In the hands of experienced surgeons, these procedures can be used safely irrespective of bone quality.

Polymer nanoparticles pass the plant interface.

As agriculture strives to feed an ever-increasing number of people, it must also adapt to increasing exposure to minute plastic particles. To learn about the accumulation of nanoplastics by plants, we prepared well-defined block copolymer nanoparticles by aqueous dispersion polymerisation. A fluorophore was incorporated via hydrazone formation and uptake into roots and protoplasts of Arabidopsis thaliana was investigated using confocal microscopy. Here we show that uptake is inversely proportional to nanoparticle size. Positively charged particles accumulate around root surfaces and are not taken up by roots or protoplasts, whereas negatively charged nanoparticles accumulate slowly and become prominent over time in the xylem of intact roots. Neutral nanoparticles penetrate rapidly into intact cells at the surfaces of plant roots and into protoplasts, but xylem loading is lower than for negative nanoparticles. These behaviours differ from those of animal cells and our results show that despite the protection of rigid cell walls, plants are accessible to nanoplastics in soil and water.

Adenylate cyclase activity of TIR1/AFB auxin receptors in plants.

The phytohormone auxin is the major coordinative signal in plant development1, mediating transcriptional reprogramming by a well-established canonical signalling pathway. TRANSPORT INHIBITOR RESPONSE 1 (TIR1)/AUXIN-SIGNALING F-BOX (AFB) auxin receptors are F-box subunits of ubiquitin ligase complexes. In response to auxin, they associate with Aux/IAA transcriptional repressors and target them for degradation via ubiquitination2,3. Here we identify adenylate cyclase (AC) activity as an additional function of TIR1/AFB receptors across land plants. Auxin, together with Aux/IAAs, stimulates cAMP production. Three separate mutations in the AC motif of the TIR1 C-terminal region, all of which abolish the AC activity, each render TIR1 ineffective in mediating gravitropism and sustained auxin-induced root growth inhibition, and also affect auxin-induced transcriptional regulation. These results highlight the importance of TIR1/AFB AC activity in canonical auxin signalling. They also identify a unique phytohormone receptor cassette combining F-box and AC motifs, and the role of cAMP as a second messenger in plants.

Chemical inhibition of the auxin inactivation pathway uncovers the roles of metabolic turnover in auxin homeostasis.

The phytohormone auxin, indole-3-acetic acid (IAA), plays a prominent role in plant development. Auxin homeostasis is coordinately regulated by auxin synthesis, transport, and inactivation; however, the physiological contribution of auxin inactivation to auxin homeostasis has not been determined. The GH3 IAA-amino acid conjugating enzymes play a central role in auxin inactivation. Chemical inhibition of GH3 proteins in planta is challenging because the inhibition of these enzymes leads to IAA overaccumulation that rapidly induces GH3 expression. Here, we report the characterization of a potent GH3 inhibitor, kakeimide, that selectively targets IAA-conjugating GH3 proteins. Chemical knockdown of the auxin inactivation pathway demonstrates that auxin turnover is very rapid (about 10 min) and indicates that both auxin biosynthesis and inactivation dynamically regulate auxin homeostasis.

An in-frame deletion mutation in the degron tail of auxin coreceptor IAA2 confers resistance to the herbicide 2,4-D in Sisymbrium orientale.

The natural auxin indole-3-acetic acid (IAA) is a key regulator of many aspects of plant growth and development. Synthetic auxin herbicides such as 2,4-D mimic the effects of IAA by inducing strong auxinic-signaling responses in plants. To determine the mechanism of 2,4-D resistance in a Sisymbrium orientale (Indian hedge mustard) weed population, we performed a transcriptome analysis of 2,4-D-resistant (R) and -susceptible (S) genotypes that revealed an in-frame 27-nucleotide deletion removing nine amino acids in the degron tail (DT) of the auxin coreceptor Aux/IAA2 (SoIAA2). The deletion allele cosegregated with 2,4-D resistance in recombinant inbred lines. Further, this deletion was also detected in several 2,4-D-resistant field populations of this species. Arabidopsis transgenic lines expressing the SoIAA2 mutant allele were resistant to 2,4-D and dicamba. The IAA2-DT deletion reduced binding to TIR1 in vitro with both natural and synthetic auxins, causing reduced association and increased dissociation rates. This mechanism of synthetic auxin herbicide resistance assigns an in planta function to the DT region of this Aux/IAA coreceptor for its role in synthetic auxin binding kinetics and reveals a potential biotechnological approach to produce synthetic auxin-resistant crops using gene-editing.

Using a ring to locate femoral head centre in total hip arthroplasty.

AIMS: Traditional methods of determining femoral head centre (FHC) during total hip arthroplasty (THA) rely on measuring the distance from a fixed point on the femur or using a calliper. The aim of this experiment was to investigate how accurately a simple circular ring could locate FHC. METHODS: 144 consecutively available femoral heads (FHs) were collected from patients undergoing THA. Each FH was orientated and mounted on a Sawbone, to create a model of its position on a proximal femur. The ring was applied to the posterior aspect of the FH and a head-centre pin (HCP) was then drilled into the FH and the ring removed, leaving the HCP in place.Each FH was then photographed normal to the axis of the HCP. A MATLAB analysis program then assessed the accuracy of the ring in locating FHC. RESULTS: Mean location accuracy for FHC was 1.77 (range 0.07-5.83) mm with 97.2% within 4 mm and all but 1 within 5 mm. CONCLUSIONS: This ring device located FHC to within 4 mm in 97% of a series of osteoarthritic FHs. This indicates that the posterior aspect of the FH maintains its sphericity late into the osteoarthritic process. Having a simple FHC location device during THA would be of value to control leg length and offset when using the posterior approach.

Hyaluronan (HA)-inspired glycopolymers as molecular tools for studying HA functions.

Hyaluronic acid (HA), the only non-sulphated glycosaminoglycan, serves numerous structural and biological functions in the human body, from providing viscoelasticity in tissues to creating hydrated environments for cell migration and proliferation. HA is also involved in the regulation of morphogenesis, inflammation and tumorigenesis through interactions with specific HA-binding proteins. Whilst the physicochemical and biological properties of HA have been widely studied for decades, the exact mechanisms by which HA exerts its multiple functions are not completely understood. Glycopolymers offer a simple and precise synthetic platform for the preparation of glycan analogues, being an alternative to the demanding synthetic chemical glycosylation. A library of homo, statistical and alternating HA glycopolymers were synthesised by reversible addition-fragmentation chain transfer polymerisation and post-modification utilising copper alkyne-azide cycloaddition to graft orthogonal pendant HA monosaccharides (N-acetyl glucosamine: GlcNAc and glucuronic acid: GlcA) onto the polymer. Using surface plasmon resonance, the binding of the glycopolymers to known HA-binding peptides and proteins (CD44, hyaluronidase) was assessed and compared to carbohydrate-binding proteins (lectins). These studies revealed potential structure-binding relationships between HA monosaccharides and HA receptors and novel HA binders, such as Dectin-1 and DEC-205 lectins. The inhibitory effect of HA glycopolymers on hyaluronidase (HAase) activity was also investigated suggesting GlcNAc- and GlcA-based glycopolymers as potential HAase inhibitors.

Generating aptamers towards human sperm cells using massively parallel sequencing.

Determining the presence of sperm cells on an item or swab is often a crucial component of sexual offence investigation. However, traditional histological staining techniques used for the morphological identification of spermatozoa lack both specificity and sensitivity, making analysis a complex and time-consuming process. New methods for the detection of sperm cells based on aptamer recognition may be able to overcome these issues. In this work, we present the selection of ssDNA aptamers against human sperm cells using Cell-SELEX and massively parallel sequencing technologies. A total of 14 rounds of selection were performed following a modified Cell-SELEX protocol, which included additional steps for the isolation of spermatozoa from seminal fluid. Massively parallel sequencing using the Illumina Miseq platform was conducted on enriched aptamer pools to elucidate the structure of potential binders. A custom bioinformatics pipeline was also developed using Galaxy for the automated processing of sequencing datasets. This data revealed several promising aptamer candidates, which were shown to selectively bind sperm cells through both microscale thermophoresis and enzyme-linked oligonucleotide assays. These aptamers have the potential to increase the efficiency of sexual offence casework by facilitating sperm detection.

Are patient-reported outcomes the same following second-side surgery in primary hip and knee arthroplasty?

AIMS: Up to one in five patients undergoing primary total hip (THA) and knee arthroplasty (TKA) require contralateral surgery. This is frequently performed as a staged procedure. This study aimed to determine if outcomes, as determined by the Oxford Hip Score (OHS) and Knee Score (OKS) differed following second-side surgery. METHODS: Over a five-year period all patients who underwent staged bilateral primary THA or TKA utilizing the same type of implants were studied. Eligible patients had both preoperative and one year Oxford scores and had their second procedure completed within a mean (2 SDs) of the primary surgery. Patient demographics, radiographs, and OHS and OKS were analyzed. RESULTS: A total of 236 patients met the inclusion criteria, of which 122 were THAs and 114 TKAs. The mean age was 66.5 years (SD 9.4), with a 2:1 female:male ratio. THAs showed similar significant improvements in outcomes following first- and second-side surgery, regardless of sex. In contrast for TKAs, although male patients demonstrated the same pattern as the THAs, female TKAs displayed significantly less improvement in both OKS and its pain component following second-side surgery. CONCLUSION: Female patients undergoing second-side TKA showed less improvement in Oxford and pain scores compared to the first-side. This difference in outcome following second-side surgery did not apply to male patients undergoing TKA, or to either sex undergoing THA. Cite this article: Bone Jt Open 2021;2(4):243-254.

The Story of Auxin-Binding Protein 1 (ABP1).

The auxin-binding protein 1 (ABP1) has endured a history of undulating prominence as a candidate receptor for this important phytohormone. Its capacity for binding auxin has not been in doubt, a feature adequately explained by its crystal structure, but any relevance of this to auxin signaling and plant development has been far more demanding to define. Over its research lifetime, it has been associated with many auxin-induced activities, including ion fluxes across the plasma membrane, rearrangement of the cytoskeleton and cell shape, and the abundance of PIN proteins at the plasma membrane via control of endocytosis, all of which required its presence in the apoplast. Yet, ABP1 has a KDEL sequence that targets it to the endoplasmic reticulum, where most of it remains. This mismatch has been more than adequately compensated for by the need for an auxin receptor to account for responses far too rapid to be executed through transcription and translation and the TIR1/AuxIAA coreceptor system. However, discoveries showing that abp1-null mutants are not compromised for auxin signaling or development, that TIR1 or AFB1 are necessarily involved with very rapid responses at the plasma membrane, and that these rapid responses are mediated with intracellular auxin all suggest that ABP1's auxin-binding capacity is not physiologically relevant. Nevertheless, ABP1 is ubiquitous in higher plants and throughout plant tissues. We need to complete its history by defining its function inside plant cells.

Seedling developmental defects upon blocking CINNAMATE-4-HYDROXYLASE are caused by perturbations in auxin transport.

The phenylpropanoid pathway serves a central role in plant metabolism, providing numerous compounds involved in diverse physiological processes. Most carbon entering the pathway is incorporated into lignin. Although several phenylpropanoid pathway mutants show seedling growth arrest, the role for lignin in seedling growth and development is unexplored. We use complementary pharmacological and genetic approaches to block CINNAMATE-4-HYDROXYLASE (C4H) functionality in Arabidopsis seedlings and a set of molecular and biochemical techniques to investigate the underlying phenotypes. Blocking C4H resulted in reduced lateral rooting and increased adventitious rooting apically in the hypocotyl. These phenotypes coincided with an inhibition in AUX transport. The upstream accumulation in cis-cinnamic acid was found to be likely to cause polar AUX transport inhibition. Conversely, a downstream depletion in lignin perturbed phloem-mediated AUX transport. Restoring lignin deposition effectively reestablished phloem transport and, accordingly, AUX homeostasis. Our results show that the accumulation of bioactive intermediates and depletion in lignin jointly cause the aberrant phenotypes upon blocking C4H, and demonstrate that proper deposition of lignin is essential for the establishment of AUX distribution in seedlings. Our data position the phenylpropanoid pathway and lignin in a new physiological framework, consolidating their importance in plant growth and development.