Correction: A preliminary study for the development of cleavable linkers using activatable fluorescent probes targeting leucine aminopeptidase.

Correction for 'A preliminary study for the development of cleavable linkers using activatable fluorescent probes targeting leucine aminopeptidase' by Julie Kang et al., Analyst, 2022, https://doi.org/10.1039/d2an01145j.

A preliminary study for the development of cleavable linkers using activatable fluorescent probes targeting leucine aminopeptidase.

Ligand-targeted drugs (LTDs) such as antibody-drug conjugates (ADCs) are currently attracting great attention as an alternative class of therapeutics to conventional chemotherapy for the clinical treatment of cancer. The linker is one of important factors determining the efficacy and toxicity of LTDs. The linker for LTDs should have enough stability during blood circulation, effectively release the payload, and leave no polar moieties in the released payload. However, the drug release activity and plasma stability of cleavable linkers are generally evaluated by complex and sophisticated in vivo techniques containing LC-MS, and the designing of new clinically applicable linkers remains a challenge. In this work, leucine aminopeptidase (LAP)-responsive fluorescent probes were designed as a simple preliminary model to verify whether a peptidase-responsive fluorescent probe can be used as a facile tool for the development of cleavable linkers although LAP is an exopeptidase and can't be a real target for cleavable linkers. LAP-responsive fluorescent probes were prepared by conjugation of a leucine to several xanthene fluorophores through a few linkages with a p-aminobenzyl spacer. The stability tests, kinetic study and live cell imaging of LAP-responsive activatable fluorescent probes demonstrated that the chemical stability and intrinsic activity of the linker for the release of drug can be easily evaluated by a fluorogenic assay. The ex vivo plasma stability test using mice suggested that an enzyme-responsive activatable fluorescent probe can be used as a feasible platform to evaluate the plasma stability of cleavable linkers during blood circulation.

Leaf form diversification in an ornamental heirloom tomato results from alterations in two different HOMEOBOX genes.

Domesticated plants display diverse phenotypic traits. However, the influence of breeding effort on this phenotypic diversity remains unknown. Here, we demonstrate that a single nucleotide deletion in the homeobox motif of BIPINNATA, a BEL-LIKE HOMEODOMAIN gene, led to a highly complex leaf phenotype in an heirloom tomato (Solanum lycopersicum), Silvery Fir Tree (SiFT), which is used as a landscaping and ornamental plant. A comparative gene network analysis revealed that repression of SOLANIFOLIA, the ortholog of WUSCHEL RELATED HOMEOBOX 1, caused the narrow leaflet phenotype seen in SiFT. Comparative genomics indicated that the bip mutation in SiFT likely arose de novo and is unique to SiFT and not introgressed from other tomato genomes. These results provide new insights into the natural variation in phenotypic traits introduced into crops during improvement processes after domestication and establish homeobox genes as evolutionary hotspots.

A Fluorogenic Assay: Analysis of Chemical Modification of Lysine and Arginine to Control Proteolytic Activity of Trypsin.

The chemical modification of amino acids plays an important role in the modulation of proteins or peptides and has useful applications in the activation and stabilization of enzymes, chemical biology, shotgun proteomics, and the production of peptide-based drugs. Although chemoselective modification of amino acids such as lysine and arginine via the insertion of respective chemical moieties as citraconic anhydride and phenyl glyoxal is important for achieving desired application objectives and has been extensively reported, the extent and chemoselectivity of the chemical modification of specific amino acids using specific chemical agents (blocking or modifying agents) has yet to be sufficiently clarified owing to a lack of suitable assay methodologies. In this study, we examined the utility of a fluorogenic assay method, based on a fluorogenic tripeptide substrate (FP-AA1-AA2-AA3) and the proteolytic enzyme trypsin, in determinations of the extent and chemoselectivity of the chemical modification of lysine or arginine. As substrates, we used two fluorogenic tripeptide probes, MeRho-Lys-Gly-Leu(Ac) (lysine-specific substrate) and MeRho-Arg-Gly-Leu(Ac) (arginine-specific substrate), which were designed, synthesized, and evaluated for chemoselective modification of specific amino acids (lysine and arginine) using the fluorogenic assay. The results are summarized in terms of half-maximal inhibitory concentrations (IC50) for the extent of modification and ratios of IC50 values (IC50arginine/IC50lysine and IC50lysine/IC50arginine) as a measure of the chemoselectivity of chemical modification for amino acids lysine and arginine. This novel fluorogenic assay was found to be rapid, precise, and reproducible for determinations of the extent and chemoselectivity of chemical modification.

Bisoprolol-based 18F-PET tracer: Synthesis and preliminary in vivo validation of ß1-blocker selectivity for ß1-adrenergic receptors in the heart.

The selectivity of a drug toward various isoforms of the target protein family is important in terms of toxicology. Typically, drug or candidate selectivity is assessed by in vitro assays, but in vivo investigations are currently lacking. Positron emission tomography (PET) allows the non-invasive determination of the in vivo distribution of a radiolabeled drug, which can provide in vivo data regarding drug selectivity. Since the discovery of propranolol, a non-selective ß-blocker inhibiting both ß1- and ß2-adrenoreceptors (ß-ARs), various selective ß1-blockers, including bisoprolol, have been developed to overcome disadvantages associated with ß2-AR inhibition. As a proof of concept, we performed an in vivo PET study to understand the selectivity and efficacy of bisoprolol as a selective ß-blocker toward ß1-AR, as the heart and peripheral smooth muscles demonstrate distinct populations of ß1- and ß2-ARs. Biodistribution of 18F-labeled bisoprolol (1, [18F]bisoprolol) showed the retention of its uptake in the heart compared with other ß-AR-rich organs at late time points post-injection. The competitive blocking assay using unlabeled bisoprolol exhibited no inhibition of [18F]bisoprolol uptake in any organ but exhibited significantly rapid loss of radioactivity between two different time points in ß1-AR-rich organs such as the heart and brain. Furthermore, the organ-to-blood ratio revealed the slow excretion and better accumulation of [18F]bisoprolol inside the heart. Collectively, the ex vivo biodistribution and blocking study presented insightful evidence to better comprehend the in vivo distribution pattern of bisoprolol as a selective inhibitor targeting ß1-ARs in the heart and provided the possibility of PET as an in vivo technique for evaluating drug selectivity.

Beyond reirradiation: Efficacy and safety of three or more courses of radiation for head and neck malignancies.

PURPOSE: Recurrent head and neck cancers are associated with significant morbidity and mortality. Outcomes of multiple courses of radiation have not yet been described. METHODS AND MATERIALS: A single institution database was queried to retrospectively review treatment plans and select patients who underwent ≥ 3 courses of radiation to the head and neck region. RESULTS: Thirty-three patients were found to have ≥ 3 courses of radiation with overlapping fields. Median local recurrence free survival after last course of reirradiation was 9.1 months and median overall survival was 10 months. Grade 3 and above toxicities were reported in 15 patients (45%). Grade 4 and above toxicities were reported in seven patients (21%). There was no grade 5 toxicity. 20 patients (61%) underwent subsequent therapies following completion of repeat reirradiation. CONCLUSIONS: Repeat reirradiation to the head and neck region is feasible and carries significant risks that are most appropriately managed with a multi-disciplinary team and must be balanced against the potential for local control and opportunities for emerging systemic therapies.

Synthesis of imatinib, a tyrosine kinase inhibitor, labeled with carbon-14.

Imatinib (Gleevec) is a multiple tyrosine kinase inhibitor that decreases the activity of the fusion oncogene called BCR-ABL (breakpoint cluster region protein-Abelson murine leukemia viral oncogene homolog) and is clinically used for the treatment of chronic myelogenous leukemia and acute lymphocytic leukemia. Small molecule drugs, such as imatinib, can bind to several cellular proteins including the target proteins in the cells, inducing undesirable effects along with the effects against the disease. In this study, we report the synthetic optimization for 14 C-labeling and radiosynthesis of [14 C]imatinib to analyze binding with cellular proteins using accelerator mass spectroscopy. 14 C-labeling of imatinib was performed by the synthesis of 14 C-labeld 2-aminopyrimidine intermediate using [14 C]guanidine·HCl, which includes an in situ reduction of an inseparable byproduct for easy purification by HPLC, followed by a cross-coupling reaction with aryl bromide precursor. The radiosynthesis of [14 C]imatinib (specific activity, 631 MBq/mmol; radiochemical purity, 99.6%) was achieved in six steps with a total chemical yield of 29.2%.

Platinum-based regimens versus cetuximab in definitive chemoradiation for human papillomavirus-unrelated head and neck cancer.

For patients ineligible for cisplatin with definitive radiotherapy (CP-CRT) for locally advanced head and neck squamous cell carcinoma (LA-HNSCC), concurrent cetuximab (C225-RT) is a popular substitute. Carboplatin-based chemoradiation (CB-CRT) is another option; however, relative efficacies of CP-CRT, CB-CRT and C225-RT are unclear, particularly in the human papillomavirus (HPV)-unrelated population. We identified 316 patients with stage III-IVB cancers of the oropharynx (24.7%), larynx (58.2%) and hypopharynx (17.1%) undergoing definitive C225-RT (N = 61), CB-CRT (N = 74) or CP-CRT (N = 181). Kaplan-Meier and cumulative incidence functions were generated to estimate overall survival (OS), locoregional failure (LRF) and distant metastasis (DM). Cox proportional hazards were used to determine the association of survival endpoints with clinical characteristics. Respectively, 3-year cumulative incidences for CP-CRT, CB-CRT and C225-RT were: LRF (0.19, 0.18 and 0.48, p ≤ 0.001), DM (0.17, 0.12 and 0.25, p = 0.32). Kaplan-Meier estimates for 3 year OS were: CP-CRT: 71%; CB-CRT: 59% and C225-RT: 54%; p = 0.0094. CP-CRT (hazard ratio [HR] 0.336; 95% confidence interval [CI] 0.203-0.557, p < 0.01) and CB-CRT (HR 0.279; 95% CI 0.141-0.551, p < 0.01) were associated with reduced hazard for LRF on multivariable analysis. CP-CRT (HR 0.548; 95% CI 0.355-0.845, p < 0.01) and CB-CRT (HR 0.549; 95% CI 0.334-0.904, p = 0.02) were associated with a reduced hazard for death on multivariable analysis. Propensity matching confirmed reduced hazards with a combined CP/CB-CRT group compared to C225-RT for LRF: HR 0.384 (p = 0.018) and OS: HR 0.557 (p = 0.045) and CB-CRT group compared to C225-RT for LRF: HR 0.427 (p = 0.023). In conclusion, CB-CRT is an effective alternative to CP-CRT in HPV-unrelated LA-HNSCC with superior locoregional control and OS compared to C225-RT.

Trends and Disparities of Proton Therapy Use among Patients with Head and Neck Cancer: Analysis from the National Cancer Database (2005-14).

PURPOSE: The purpose of this study was to analyze national trends and disparities in proton therapy use among patients with head and neck cancer receiving radiotherapy to primary disease sites. PATIENTS AND METHODS: Using the National Cancer Database, we identified patients diagnosed with any nonmetastatic head and neck primary malignancy between 2005 and 2014 who were treated with radiation therapy or proton therapy directed specifically at the primary disease site. Distributions of patient and clinical factors between the two groups were evaluated. Multivariable logistic regression was used to correlate factors associated with proton therapy use compared with other modalities of radiation therapy. RESULTS: There were 220 491 patients who received any radiation therapy as part of their initial treatment course, only 417 (0.2%) of whom received proton therapy. The use of protons underwent a small increase from 0.13% in 2005-06 to 0.41% by 2013-14 (P < .001). The most common primary sites treated with proton therapy were the nasal cavity/nasopharynx (n = 151, 36.2%) and the oral cavity (n = 98, 23.5%). Most patients had T4 disease (n = 94, 31.0%). On multivariable logistic regression, all primary sites compared with hypopharynx/larynx sites (odds ratio [OR], 2.53-10.53; P < .001), treatment at an academic facility (OR, 2.54; P < .001), ≥ 13-mile distance from the treating facility (OR, 1.94; P < .001), and highest median household income quartile (> $63 000; OR, 2.52; P = .002) were associated with an increased likelihood of receiving proton therapy. CONCLUSION: Proton use has undergone an incremental increase in the United States but remains an uncommon modality for the treatment of primary head and neck cancer.

Asymmetric and Reduced Xanthene Fluorophores: Synthesis, Photochemical Properties, and Application to Activatable Fluorescent Probes for Detection of Nitroreductase.

Xanthene fluorophores, including fluorescein, rhodol, and rhodamines, are representative classes of fluorescent probes that have been applied in the detection and visualization of biomolecules. "Turn on" activatable fluorescent probes, that can be turned on in response to enzymatic reactions, have been developed and prepared to reduce the high background signal of "always-on" fluorescent probes. However, the development of activity-based fluorescent probes for biological applications, using simple xanthene dyes, is hampered by their inefficient synthetic methods and the difficulty of chemical modifications. We have, thus, developed a highly efficient, versatile synthetic route to developing chemically more stable reduced xanthene fluorophores, based on fluorescein, rhodol, and rhodamine via continuous Pd-catalyzed cross-coupling. Their fluorescent nature was evaluated by monitoring fluorescence with variation in the concentration, pH, and solvent. As an application to activatable fluorescent probe, nitroreductase (NTR)-responsive fluorescent probes were also developed using the reduced xanthene fluorophores, and their fluorogenic properties were evaluated.

Gaze position modulates the effectiveness of forward collision warnings for drowsy drivers.

Advanced driver assistance systems (ADAS) have the potential to prevent crashes and reduce their severity. Forward collision warnings (FCW) are quickly becoming standard across vehicle lineups and may prevent frontal crashes by alerting drivers. Previous research has demonstrated the effectiveness of FCW for distracted drivers, but their effectiveness for other types of impairment remains unknown. Like distraction, drowsiness can impair driver response time and lead to crashes. The goal of the present study was to evaluate the effectiveness of FCW for moderately and severely drowsy drivers using a high-fidelity driving simulator. Drowsy drivers were divided into three warning conditions during a revealed stop vehicle forward collision event: An auditory alert, a haptic seat vibration, and a no warning baseline. Results indicate that FCW were effective at speeding drowsy driver response, but only when the drowsy drivers were looking away from the forward roadway at the onset of the event. These results have important implications for ADAS technology and driver state monitoring systems.

Comparing Kadish, TNM, and the modified Dulguerov staging systems for esthesioneuroblastoma.

BACKGROUND: Esthesioneuroblastoma (ENB) is a rare neuroendocrine tumor. The purpose of this study was to compare the Kadish, tumor-node-metastasis (TNM), and Dulguerov's modified TNM staging in order to determine the impact of the stage on primary surgical treatment selection, margin status, and survival. METHODS: The National Cancer Database (NCDB) was used to identify patients diagnosed with ENB between 2004 to 2015. Patients were excluded based on the ability to properly stage their disease as well as the availability of treatment data. RESULTS: Eight-hundred eighty-three patients had sufficient data for analysis. On multivariate analysis, age and government insurance were associated with primary surgical treatment, whereas tumor stage, gender, race, hospital type and volume, and comorbidity score were not. Age, charlson-deyo comorbidity (CDCC) score, hospital volume, and nodal status were found to be predictors of survival. Multivariate-analysis controlling for stage failed to demonstrate clear survival differences between staging in both TNM and Kadish systems. T-stage and the presence of regional nodal metastasis were associated with an increased risk of positive margins on multivariate analysis. CONCLUSION: Although primary surgical management and positive margins can be predicted by certain patient and tumor factors, clinical staging systems for ENB poorly predict prognosis over a 10-year horizon.

Radiosynthesis, biological evaluation and preliminary microPET study of 18F-labeled 5-resorcinolic triazolone derivative based on ganetespib targeting HSP90.

Heat-shock protein 90 (HSP90) is a molecular chaperone that activates oncogenic transformation in several solid tumors, including lung and breast cancers. Ganetespib, a most promising candidate among several HSP90 inhibitors under clinical trials, has entered Phase III clinical trials for cancer therapy. Despite numerous evidences validating HSP90 as a target of anticancer, there are few studies on PET agents targeting oncogenic HSP90. In this study, we synthesized and biologically evaluated a novel 18F-labeled 5-resorcinolic triazolone derivative (1, [18F]PTP-Ganetespib) based on ganetespib. [18F]PTP-Ganetespib was labeled by click chemistry of Ganetespib-PEG-Alkyne (10) and [18F]PEG-N3 (11) with 37.3 ±â€¯5.11% of radiochemical yield and 99.7 ±â€¯0.09% of radiochemical purity. [18F]PTP-Ganetespib showed proper LogP (0.96 ±â€¯0.06) and good stability in human serum over 97% for 2 h. [18F]PTP-Ganetespib showed high uptakes in breast cancer cells containing triple negative breast cancer (TNBC) MDA-MB-231 and Her2-negative MCF-7 cells, which are target breast cancer cell lines of HSP90 inhibitor, ganetespib, as an anticancer. Blocking of HSP90 by the pretreatment of ganetespib exhibited significantly decreased accumulation of [18F]PTP-Ganetespib in MDA-MB-231 and MCF-7 cells, indicating the specific binding of [18F]PTP-Ganetespib to MDA-MB-231 and MCF-7 cells with high HSP90 expression. In the biodistribution and microPET imaging studies, the initial uptake into tumor was weaker than in other thoracic and abdominal organs, but [18F]PTP-Ganetespib was retained relatively longer in the tumor than other organs. The uptake of [18F]PTP-Ganetespib in tumors was not sufficient for further development as a tumor-specific PET imaging agent by itself, but this preliminary PET imaging study of [18F]PTP-Ganetespib can be basis for developing new PET imaging agents based on HSP90 inhibitor, ganetespib.

Impact of smoke-free housing policy lease exemptions on compliance, enforcement and smoking behavior: A qualitative study.

This paper investigates the impacts of smoke-free housing policies on compliance, enforcement and smoking behavior. From 2012 to 2014, we studied two affordable housing providers in Canada with comprehensive smoke-free policies: Waterloo Regional Housing that required new leases to be non-smoking and exempted existing leases, and Yukon Housing Corporation that required all leases (existing and new) to be non-smoking. Focus groups and key informant interviews were conducted with 31 housing and public health staff involved in policy development and implementation, and qualitative interviews with 56 tenants. Both types of smoke-free policies helped tenants to reduce and quit smoking. However, exempting existing tenants from the policy created challenges for monitoring compliance and enforcing the policy, and resulted in ongoing tobacco smoke exposure. Moreover, some new tenants were smoking in exempted units, which undermined the policy and maintained smoking behavior. Our findings support the implementation of complete smoke-free housing policies that do not exempt existing leases to avoid many of the problems experienced by staff and tenants. In jurisdictions where exempting existing leases is still required by law, adequate staff resources for monitoring and enforcement, along with consistent and clear communication (particularly regarding balconies, patios and outdoor spaces) will encourage compliance.

Survival Impact of Cardiac Dose Following Lung Stereotactic Body Radiotherapy.

INTRODUCTION: The purpose of this study was to determine the impact of radiation dose to substructures of the heart in lung stereotactic body radiotherapy (SBRT) patients on non-cancer-related deaths. METHODS: Patients treated with lung SBRT at a single institution from 2005 to 2013 were included. The heart and its substructures were contoured, and dose was calculated including mean, max, and max 10 cc dose. Clinical variables including stage, histology, age, gender, Charlson comorbidity index (CCI), preexisting cardiac disease, pulmonary function (forced expiratory volume in 1 second, diffusion capacity), and smoking status were explored for association with non-cancer-related deaths in univariable (UVA) and multivariable (MVA) analyses. Heart dosimetric parameters were correlated with the risk of radiation pneumonitis (RP) using UVA and MVA. RESULTS: A total of 189 patients were included with median age of 76 years (range, 48-93 years). Of these patients, 45.5% were female, 27.5% were T2, 16.9% were current smokers, 64% had preexisting cardiac risk factors, and 34.5% had CCI score of ≥ 3. Mean lung dose ± SD was 456 ± 231 cGy. Heart max, mean, and 10 cc doses were 1867 ± 1712 cGy, 265 ± 269 cGy, and 1150 ± 1075 cGy, respectively. There were 14 (7.4%) ≥ Grade 2 RP and 3 (1.6%) were ≥ Grade 3. The median overall survival was 37.3 months (95% confidence interval, 29.8-45.3 months). On UVA, female gender (P < .01), higher Eastern Cooperative Oncology Group (P = .01), cardiac risk (P < .01), CCI (P < .01), and bilateral ventricles max dose (P = .02) were associated with non-cancer-related deaths; on MVA, bilateral ventricles max dose was significant (P = .05). No heart parameters were associated with RP. CONCLUSIONS: Higher bilateral ventricles max dose is associated with poorer survival. Heart dose parameters should be considered when planning patients for SBRT.

Evaluating driver drowsiness countermeasures.

OBJECTIVE: Driver drowsiness contributes to a substantial number of fatal and nonfatal crashes, with recent estimates attributing up to 21% of fatal crashes to drowsiness. This article describes recent NHTSA research on in-vehicle drowsiness countermeasures. Recent advances in technology and state detection algorithms have shown success in detecting drowsiness using a variety of data sources, including camera-based eye tracking, steering wheel position, yaw rate, and vehicle lane position. However, detection is just the first step in reducing drowsy driving crashes. Countermeasures are also needed to provide feedback to the driver, modify driver behavior, and prevent crashes. The goal of this study was to evaluate the effectiveness of in-vehicle drowsiness countermeasures in reducing drowsy lane departures. The tested countermeasures included different warning modalities in either a discrete or staged interface. METHODS: Data were collected from 72 young adult drivers (age 21-32) in the high-fidelity full-motion National Advanced Driving Simulator. Drivers completed a 45-min simulated nighttime drive at 2 time points, late night and early morning, where drowsiness was manipulated by continuous hours awake. Forty-eight drivers were exposed to one of 6 countermeasures that varied along 2 dimensions, type and modality. The countermeasures relied on a steering-based drowsiness detection algorithm developed in prior NHTSA research. Twenty-four drivers received no countermeasure and were used as a baseline comparison. System effectiveness was measured by lane departures and standard deviation in lateral position (SDLP). RESULTS: There was a reduction in drowsy lane departure frequency and lane position variability for drivers with countermeasures compared to the baseline no-countermeasure group. Importantly, the data suggest that multistage alerts, which provide an indication of increasing urgency, were more effective in reducing drowsy lane departures than single-stage discrete alerts, particularly during early morning drives when drivers were drowsier. CONCLUSIONS: The results indicate that simple in-vehicle countermeasures, such as an auditory-visual coffee cup icon, can reduce the frequency of drowsy lane departures in the context of relatively short drives. An important next step is to evaluate the impact of drowsiness countermeasures in the context of longer, multiple-hour drives. In these cases, it may not be possible to keep drivers awake via feedback warnings and it is important to understand whether countermeasures prompt drivers to stop to rest. The next phase of this research project will examine the role of drowsiness countermeasures over longer drives using a protocol that replicates the motivational conditions of drowsy driving.

Climate and Developmental Plasticity: Interannual Variability in Grapevine Leaf Morphology.

The shapes of leaves are dynamic, changing over evolutionary time between species, within a single plant producing different shaped leaves at successive nodes, during the development of a single leaf as it allometrically expands, and in response to the environment. Notably, strong correlations between the dissection and size of leaves with temperature and precipitation exist in both the paleorecord and extant populations. Yet, a morphometric model integrating evolutionary, developmental, and environmental effects on leaf shape is lacking. Here, we continue a morphometric analysis of >5,500 leaves representing 270 grapevines of multiple Vitis species between two growing seasons. Leaves are paired one-to-one and vine-to-vine accounting for developmental context, between growing seasons. Linear discriminant analysis reveals shape features that specifically define growing season, regardless of species or developmental context. The shape feature, a more pronounced distal sinus, is associated with the colder, drier growing season, consistent with patterns observed in the paleorecord. We discuss the implications of such plasticity in a long-lived woody perennial, such as grapevine (Vitis spp.), with respect to the evolution and functionality of plant morphology and changes in climate.

KNOX1 genes regulate lignin deposition and composition in monocots and dicots.

Plant secondary cell walls are deposited mostly in vascular tissues such as xylem vessels, tracheids, and fibers. These cell walls are composed of a complex matrix of compounds including cellulose, hemicellulose, and lignin. Lignin functions primarily to maintain the structural and mechanical integrity of both the transport vessel and the entire plant itself. Since lignin has been identified as a major source of biomass for biofuels, regulation of secondary cell wall biosynthesis has been a topic of much recent investigation. Biosynthesis and patterning of lignin involves many developmental and environmental cues including evolutionarily conserved transcriptional regulatory modules and hormonal signals. Here, we investigate the role of the class I Knotted1-like-homeobox (KNOX) genes and gibberellic acid in the lignin biosynthetic pathway in a representative monocot and a representative eudicot. Knotted1 overexpressing mutant plants showed a reduction in lignin content in both maize and tobacco. Expression of four key lignin biosynthesis genes was analyzed and revealed that KNOX1 genes regulate at least two steps in the lignin biosynthesis pathway. The negative regulation of lignin both in a monocot and a eudicot by the maize Kn1 gene suggests that lignin biosynthesis may be preserved across large phylogenetic distances. The evolutionary implications of regulation of lignification across divergent species are discussed.

The plant vascular system: evolution, development and functions.

The emergence of the tracheophyte-based vascular system of land plants had major impacts on the evolution of terrestrial biology, in general, through its role in facilitating the development of plants with increased stature, photosynthetic output, and ability to colonize a greatly expanded range of environmental habitats. Recently, considerable progress has been made in terms of our understanding of the developmental and physiological programs involved in the formation and function of the plant vascular system. In this review, we first examine the evolutionary events that gave rise to the tracheophytes, followed by analysis of the genetic and hormonal networks that cooperate to orchestrate vascular development in the gymnosperms and angiosperms. The two essential functions performed by the vascular system, namely the delivery of resources (water, essential mineral nutrients, sugars and amino acids) to the various plant organs and provision of mechanical support are next discussed. Here, we focus on critical questions relating to structural and physiological properties controlling the delivery of material through the xylem and phloem. Recent discoveries into the role of the vascular system as an effective long-distance communication system are next assessed in terms of the coordination of developmental, physiological and defense-related processes, at the whole-plant level. A concerted effort has been made to integrate all these new findings into a comprehensive picture of the state-of-the-art in the area of plant vascular biology. Finally, areas important for future research are highlighted in terms of their likely contribution both to basic knowledge and applications to primary industry.

Interspecific RNA interference of SHOOT MERISTEMLESS-like disrupts Cuscuta pentagona plant parasitism.

Infection of crop species by parasitic plants is a major agricultural hindrance resulting in substantial crop losses worldwide. Parasitic plants establish vascular connections with the host plant via structures termed haustoria, which allow acquisition of water and nutrients, often to the detriment of the infected host. Despite the agricultural impact of parasitic plants, the molecular and developmental processes by which host/parasitic interactions are established are not well understood. Here, we examine the development and subsequent establishment of haustorial connections by the parasite dodder (Cuscuta pentagona) on tobacco (Nicotiana tabacum) plants. Formation of haustoria in dodder is accompanied by upregulation of dodder KNOTTED-like homeobox transcription factors, including SHOOT MERISTEMLESS-like (STM). We demonstrate interspecific silencing of a STM gene in dodder driven by a vascular-specific promoter in transgenic host plants and find that this silencing disrupts dodder growth. The reduced efficacy of dodder infection on STM RNA interference transgenics results from defects in haustorial connection, development, and establishment. Identification of transgene-specific small RNAs in the parasite, coupled with reduced parasite fecundity and increased growth of the infected host, demonstrates the efficacy of interspecific small RNA-mediated silencing of parasite genes. This technology has the potential to be an effective method of biological control of plant parasite infection.