Allosteric interactions prime androgen receptor dimerization and activation.

The androgen receptor (AR) is a nuclear receptor that governs gene expression programs required for prostate development and male phenotype maintenance. Advanced prostate cancers display AR hyperactivation and transcriptome expansion, in part, through AR amplification and interaction with oncoprotein cofactors. Despite its biological importance, how AR domains and cofactors cooperate to bind DNA has remained elusive. Using single-particle cryo-electron microscopy, we isolated three conformations of AR bound to DNA, showing that AR forms a non-obligate dimer, with the buried dimer interface utilized by ancestral steroid receptors repurposed to facilitate cooperative DNA binding. We identify novel allosteric surfaces which are compromised in androgen insensitivity syndrome and reinforced by AR's oncoprotein cofactor, ERG, and by DNA-binding motifs. Finally, we present evidence that this plastic dimer interface may have been adopted for transactivation at the expense of DNA binding. Our work highlights how fine-tuning AR's cooperative interactions translate to consequences in development and disease.

Single-cell analysis of treatment-resistant prostate cancer: Implications of cell state changes for cell surface antigen-targeted therapies.

Targeting cell surface molecules using radioligand and antibody-based therapies has yielded considerable success across cancers. However, it remains unclear how the expression of putative lineage markers, particularly cell surface molecules, varies in the process of lineage plasticity, wherein tumor cells alter their identity and acquire new oncogenic properties. A notable example of lineage plasticity is the transformation of prostate adenocarcinoma (PRAD) to neuroendocrine prostate cancer (NEPC)-a growing resistance mechanism that results in the loss of responsiveness to androgen blockade and portends dismal patient survival. To understand how lineage markers vary across the evolution of lineage plasticity in prostate cancer, we applied single-cell analyses to 21 human prostate tumor biopsies and two genetically engineered mouse models, together with tissue microarray analysis on 131 tumor samples. Not only did we observe a higher degree of phenotypic heterogeneity in castrate-resistant PRAD and NEPC than previously anticipated but also found that the expression of molecules targeted therapeutically, namely PSMA, STEAP1, STEAP2, TROP2, CEACAM5, and DLL3, varied within a subset of gene-regulatory networks (GRNs). We also noted that NEPC and small cell lung cancer subtypes shared a set of GRNs, indicative of conserved biologic pathways that may be exploited therapeutically across tumor types. While this extreme level of transcriptional heterogeneity, particularly in cell surface marker expression, may mitigate the durability of clinical responses to current and future antigen-directed therapies, its delineation may yield signatures for patient selection in clinical trials, potentially across distinct cancer types.

Author Correction: FOXA1 mutations alter pioneering activity, differentiation and prostate cancer phenotypes.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

FOXA1 mutations alter pioneering activity, differentiation and prostate cancer phenotypes.

Mutations in the transcription factor FOXA1 define a unique subset of prostate cancers but the functional consequences of these mutations and whether they confer gain or loss of function is unknown1-9. Here, by annotating the landscape of FOXA1 mutations from 3,086 human prostate cancers, we define two hotspots in the forkhead domain: Wing2 (around 50% of all mutations) and the highly conserved DNA-contact residue R219 (around 5% of all mutations). Wing2 mutations are detected in adenocarcinomas at all stages, whereas R219 mutations are enriched in metastatic tumours with neuroendocrine histology. Interrogation of the biological properties of wild-type FOXA1 and fourteen FOXA1 mutants reveals gain of function in mouse prostate organoid proliferation assays. Twelve of these mutants, as well as wild-type FOXA1, promoted an exaggerated pro-luminal differentiation program, whereas two different R219 mutants blocked luminal differentiation and activated a mesenchymal and neuroendocrine transcriptional program. Assay for transposase-accessible chromatin using sequencing (ATAC-seq) of wild-type FOXA1 and representative Wing2 and R219 mutants revealed marked, mutant-specific changes in open chromatin at thousands of genomic loci and exposed sites of FOXA1 binding and associated increases in gene expression. Of note, ATAC-seq peaks in cells expressing R219 mutants lacked the canonical core FOXA1-binding motifs (GTAAAC/T) but were enriched for a related, non-canonical motif (GTAAAG/A), which was preferentially activated by R219-mutant FOXA1 in reporter assays. Thus, FOXA1 mutations alter its pioneering function and perturb normal luminal epithelial differentiation programs, providing further support for the role of lineage plasticity in cancer progression.

Optical Control of Cytokine Production Using Photoswitchable Galactosylceramides.

α-Galactosylceramides are glycosphingolipids that show promise in cancer immunotherapy. After presentation by CD1d, they activate natural killer T cells (NKT), which results in the production of a variety of pro-inflammatory and immunomodulatory cytokines. Herein, we report the synthesis and biological evaluation of photochromic derivatives of KRN-7000, the activity of which can be modulated with light. Based on established structure-activity relationships, we designed photoswitchable analogues of this glycolipid that control the production of pro-inflammatory cytokines, such as IFN-γ. The azobenzene derivative α-GalACer-4 proved to be more potent than KRN-7000 itself when activated with 370 nm light. Photolipids of this type could improve our mechanistic understanding of cytokine production and could open new directions in photoimmunotherapy.

IL-14 and IL-16 are expressed in the thyroid of patients with either Graves' disease or Hashimoto's thyroiditis.

OBJECTIVES: Cytokines have an important role in orchestrating the pathophysiology in autoimmune thyroid disease. The aim of the current study was to analyse the expression of interleukin (IL)-14 and IL-16 in the thyroid tissue of patients with Graves' disease (GD), Hashimoto's thyroiditis (HT) or multinodular goitre (MNG) and in that of normal individuals, in patients' intrathyroidal CD4(+) and CD8(+) T cells, and in patient and normal cultured thyroid follicular cells. METHODS: The expression of IL-14 and IL-16 mRNA and protein was investigated using reverse transcription-polymerase chain reaction (RT-PCR) amplification, and Western blotting and ELISAs, respectively. RESULTS: IL-14 mRNA expression was detected in thyroid tissue from 8/9 GD, 3/4 HT and 3/13 MNG patients and 1/6 normal individuals, and IL-16 mRNA expression in thyroid tissue from 9/9 GD, 4/4 HT and 9/13 MNG patients and 4/6 normal individuals. IL-14 mRNA expression was detected in intrathyroidal CD4(+) and CD8(+) T cells from 2/2 GD and 2/2 HT patients, while IL-16 mRNA was detected in samples from 1/2 HT patients but not in those from either patient with GD. IL-14 and IL-16 mRNA expression was found in thyroid follicular cells derived from 2/2 patient with GD and 1/1 normal individual. IL-14 protein was detected in thyroid tissue from 6/6 GD, 1/1 HT and 0/6 MNG patients and 0/6 normal individuals, and IL-16 protein in thyroid tissue from 6/6 GD, 1/1 HT and 1/6 MNG patients and 0/6 normal individuals. Expression of IL-14 protein was stimulated in thyroid follicular cells derived from two patients with GD and one normal individual by peripheral blood mononuclear cell (PBMC)-conditioned medium. Treatment of thyrocytes from two patients with GD and one normal individual with PBMC-conditioned medium and tumour necrosis factor (TNF)-α stimulated IL-16 protein expression. In normal thyrocytes, IL-16 protein synthesis was induced also by IL-1ß, IL-17A, IL-4 and transforming growth factor (TGF)-ß. CONCLUSIONS: The data provide evidence that the intrathyroidal production of IL-14 and IL-16 is associated with the pathogenesis of autoimmune thyroid disease. Thyroid follicular cells display the ability to express IL-14 and IL-16 mRNA and can be stimulated to express IL-16 protein, by a panel of cytokines, and IL-14 protein, by as yet unidentified factors.

Neuropsychological profiles of patients with juvenile myoclonic epilepsy and their siblings: An extended study.

OBJECTIVE: To examine executive function, intelligence, visuospatial skills, language, memory, attention, reaction time, anxiety, depression, and emotional and behavioral traits most frequently associated with executive dysfunction in patients with juvenile myoclonic epilepsy (JME) compared with a sibling and a normal control group under video-electroencephalography (video-EEG) conditions. METHODS: Twenty-two sibling pairs, one with JME, were compared with 44 controls matched for age, gender, and educational level. All participants were administered a comprehensive set of neuropsychological and questionnaire measures during and without video-EEG recording. RESULTS: The JME group differed significantly from controls in measures of phonemic and semantic verbal fluency. They scored significantly higher on the dysexecutive self-rating questionnaire, being more likely to report traits associated with executive dysfunction than both siblings and controls. Patients with JME reported significantly low mood than both controls and their siblings. Unaffected siblings differed significantly from controls on psychomotor speed, phonemic verbal fluency and were considered to exhibit traits associated with executive dysfunction by others. Qualitative inspection of data suggested a convincing trend for patients with JME and their siblings to perform worse than controls on most measures. SIGNIFICANCE: This study supports the existence of a distinct neuropsychological profile among patients with JME and their siblings, which is likely to be genetically determined. The similarity of neuropsychological profiles between JME patients and their siblings is independent of antiepileptic drug effects or subclinical EEG activity. The significant differences between the sibling and controls suggests that there is a neurocognitive endophenotype for JME.

The effect of TG2-inhibitory monoclonal antibody zampilimab on tissue fibrosis in human in vitro and primate in vivo models of chronic kidney disease.

Fibrotic remodeling is the primary driver of functional loss in chronic kidney disease, with no specific anti-fibrotic agent available for clinical use. Transglutaminase 2 (TG2), a wound response enzyme that irreversibly crosslinks extracellular matrix proteins causing dysregulation of extracellular matrix turnover, is a well-characterized anti-fibrotic target in the kidney. We describe the humanization and characterization of two anti-TG2 monoclonal antibodies (zampilimab [hDC1/UCB7858] and BB7) that inhibit crosslinking by TG2 in human in vitro and rabbit/cynomolgus monkey in vivo models of chronic kidney disease. Determination of zampilimab half-maximal inhibitory concentration (IC50) against recombinant human TG2 was undertaken using the KxD assay and determination of dissociation constant (Kd) by surface plasmon resonance. Efficacy in vitro was established using a primary human renal epithelial cell model of tubulointerstitial fibrosis, to assess mature deposited extracellular matrix proteins. Proof of concept in vivo used a cynomolgus monkey unilateral ureteral obstruction model of chronic kidney disease. Zampilimab inhibited TG2 crosslinking transamidation activity with an IC50 of 0.25 nM and Kd of <50 pM. In cell culture, zampilimab inhibited extracellular TG2 activity (IC50 119 nM) and dramatically reduced transforming growth factor-ß1-driven accumulation of multiple extracellular matrix proteins including collagens I, III, IV, V, and fibronectin. Intravenous administration of BB7 in rabbits resulted in a 68% reduction in fibrotic index at Day 25 post-unilateral ureteral obstruction. Weekly intravenous administration of zampilimab in cynomolgus monkeys with unilateral ureteral obstruction reduced fibrosis at 4 weeks by >50%, with no safety signals. Our data support the clinical investigation of zampilimab for the treatment of kidney fibrosis.

Targeting prostate tumor low-molecular weight tyrosine phosphatase for oxidation-sensitizing therapy.

Protein tyrosine phosphatases (PTPs) play major roles in cancer and are emerging as therapeutic targets. Recent reports suggest low-molecular weight PTP (LMPTP)-encoded by the ACP1 gene-is overexpressed in prostate tumors. We found ACP1 up-regulated in human prostate tumors and ACP1 expression inversely correlated with overall survival. Using CRISPR-Cas9-generated LMPTP knockout C4-2B and MyC-CaP cells, we identified LMPTP as a critical promoter of prostate cancer (PCa) growth and bone metastasis. Through metabolomics, we found that LMPTP promotes PCa cell glutathione synthesis by dephosphorylating glutathione synthetase on inhibitory Tyr270. PCa cells lacking LMPTP showed reduced glutathione, enhanced activation of eukaryotic initiation factor 2-mediated stress response, and enhanced reactive oxygen species after exposure to taxane drugs. LMPTP inhibition slowed primary and bone metastatic prostate tumor growth in mice. These findings reveal a role for LMPTP as a critical promoter of PCa growth and metastasis and validate LMPTP inhibition as a therapeutic strategy for treating PCa through sensitization to oxidative stress.

Single Cell Analysis of Treatment-Resistant Prostate Cancer: Implications of Cell State Changes for Cell Surface Antigen Targeted Therapies.

Targeting cell surface molecules using radioligand and antibody-based therapies has yielded considerable success across cancers. However, it remains unclear how the expression of putative lineage markers, particularly cell surface molecules, varies in the process of lineage plasticity, wherein tumor cells alter their identity and acquire new oncogenic properties. A notable example of lineage plasticity is the transformation of prostate adenocarcinoma (PRAD) to neuroendocrine prostate cancer (NEPC)--a growing resistance mechanism that results in the loss of responsiveness to androgen blockade and portends dismal patient survival. To understand how lineage markers vary across the evolution of lineage plasticity in prostate cancer, we applied single cell analyses to 21 human prostate tumor biopsies and two genetically engineered mouse models, together with tissue microarray analysis (TMA) on 131 tumor samples. Not only did we observe a higher degree of phenotypic heterogeneity in castrate-resistant PRAD and NEPC than previously anticipated, but also found that the expression of molecules targeted therapeutically, namely PSMA, STEAP1, STEAP2, TROP2, CEACAM5, and DLL3, varied within a subset of gene-regulatory networks (GRNs). We also noted that NEPC and small cell lung cancer (SCLC) subtypes shared a set of GRNs, indicative of conserved biologic pathways that may be exploited therapeutically across tumor types. While this extreme level of transcriptional heterogeneity, particularly in cell surface marker expression, may mitigate the durability of clinical responses to novel antigen-directed therapies, its delineation may yield signatures for patient selection in clinical trials, potentially across distinct cancer types.

A qualitative study of primary care professionals' views of case finding for depression in patients with diabetes or coronary heart disease in the UK.

BACKGROUND: Routinely conducting case finding (also commonly referred to as screening) in patients with chronic illness for depression in primary care appears to have little impact. We explored the views and experiences of primary care nurses, doctors and managers to understand how the implementation of case finding/screening might impact on its effectiveness. METHODS: Two complementary qualitative focus group studies of primary care professionals including nurses, doctors and managers, in five primary care practices and five Community Health Partnerships, were conducted in Scotland. RESULTS: We identified several features of the way case finding/screening was implemented that may lead to systematic under-detection of depression. These included obstacles to incorporating case finding/screening into a clinical review consultation; a perception of replacing individualised care with mechanistic assessment, and a disconnection for nurses between management of physical and mental health. Far from being a standardised process that encouraged detection of depression, participants described case finding/screening as being conducted in a way which biased it towards negative responses, and for nurses, it was an uncomfortable task for which they lacked the necessary skills to provide immediate support to patients at the time of diagnosis. CONCLUSION: The introduction of case finding/screening for depression into routine chronic illness management is not straightforward. Routinized case finding/screening for depression can be implemented in ways that may be counterproductive to engagement (particularly by nurses), with the mental health needs of patients living with long term conditions. If case finding/screening or engagement with mental health problems is to be promoted, primary care nurses require more training to increase their confidence in raising and dealing with mental health issues and GPs and nurses need to work collectively to develop the relational work required to promote cognitive participation in case finding/screening.

Constitutively active androgen receptor splice variants expressed in castration-resistant prostate cancer require full-length androgen receptor.

Androgen receptor (AR) splice variants lacking the ligand binding domain (ARVs), originally isolated from prostate cancer cell lines derived from a single patient, are detected in normal and malignant human prostate tissue, with the highest levels observed in late stage, castration-resistant prostate cancer. The most studied variant (called AR-V7 or AR3) activates AR reporter genes in the absence of ligand and therefore, could play a role in castration resistance. To explore the range of potential ARVs, we screened additional human and murine prostate cancer models using conventional and next generation sequencing technologies and detected several structurally diverse AR isoforms. Some, like AR-V7/AR3, display gain of function, whereas others have dominant interfering activity. We also find that ARV expression increases acutely in response to androgen withdrawal, is suppressed by testosterone, and in some models, is coupled to full-length AR (AR-FL) mRNA production. As expected, constitutively active, ligand-independent ARVs such as AR-V7/AR3 are sufficient to confer anchorage-independent (in vitro) and castration-resistant (in vivo) growth. Surprisingly, this growth is blocked by ligand binding domain-targeted antiandrogens, such as MDV3100, or by selective siRNA silencing of AR-FL, indicating that the growth-promoting effects of ARVs are mediated through AR-FL. These data indicate that the increase in ARV expression in castrate-resistant prostate cancer is an acute response to castration rather than clonal expansion of castration or antiandrogen-resistant cells expressing gain of function ARVs, and furthermore, they provide a strategy to overcome ARV function in the clinic.

Who visits a national park and what do they get out of it?: a joint visitor cluster analysis and travel cost model for Yellowstone National Park.

Yellowstone National Park visitor data were obtained from a survey collected for the National Park Service by the Park Studies Unit at the University of Idaho. Travel cost models have been conducted for national parks in the United States; however, this study builds on these studies and investigates how benefits vary by types of visitors who participate in different activities while at the park. Visitor clusters were developed based on activities in which a visitor participated while at the park. The clusters were analyzed and then incorporated into a travel cost model to determine the economic value (consumer surplus) that the different visitor groups received from visiting the park. The model was estimated using a zero-truncated negative binomial regression corrected for endogenous stratification. The travel cost price variable was estimated using both 1/3 and 1/4 the wage rate to test for sensitivity to opportunity cost specification. The average benefit across all visitor cluster groups was estimated at between $235 and $276 per person per trip. However, per trip benefits varied substantially across clusters; from $90 to $103 for the "value picnickers," to $185-$263 for the "backcountry enthusiasts," $189-$278 for the "do it all adventurists," $204-$303 for the "windshield tourists," and $323-$714 for the "creature comfort" cluster group.

A crucial sequence for transglutaminase type 2 extracellular trafficking in renal tubular epithelial cells lies in its N-terminal beta-sandwich domain.

Transglutaminase type 2 (TG2) catalyzes the formation of an ε-(γ-glutamyl)-lysine isopeptide bond between adjacent peptides or proteins including those of the extracellular matrix (ECM). Elevated extracellular TG2 leads to accelerated ECM deposition and reduced clearance that underlie tissue scarring and fibrosis. The extracellular trafficking of TG2 is crucial to its role in ECM homeostasis; however, the mechanism by which TG2 escapes the cell is unknown as it has no signal leader peptide and therefore cannot be transported classically. Understanding TG2 transport may highlight novel mechanisms to interfere with the extracellular function of TG2 as isoform-specific TG2 inhibitors remain elusive. Mammalian expression vectors were constructed containing domain deletions of TG2. These were transfected into three kidney tubular epithelial cell lines, and TG2 export was assessed to identify critical domains. Point mutation was then used to highlight specific sequences within the domain required for TG2 export. The removal of ß-sandwich domain prevented all TG2 export. Mutations of Asp(94) and Asp(97) within the N-terminal ß-sandwich domain were identified as crucial for TG2 externalization. These form part of a previously identified fibronectin binding domain ((88)WTATVVDQQDCTLSLQLTT(106)). However, siRNA knockdown of fibronectin failed to affect TG2 export. The sequence (88)WTATVVDQQDCTLSLQLTT(106) within the ß-sandwich domain of TG2 is critical to its export in tubular epithelial cell lines. The extracellular trafficking of TG2 is independent of fibronectin.

Cognitive and emotional abnormalities in systemic lupus erythematosus: evidence for amygdala dysfunction.

Systemic lupus erythematosus (SLE) is a multi-system autoimmune disorder characterized by the production of autoantibodies. Approximately 30-50 % of patients produce autoantibodies directed against N-Methyl-D-aspartic acid receptors (NMDARs). Once they have gained access to brain tissue, these autoantibodies bind to the NR2A subunit of the NMDARs and synergize with glutamate to cause excitatory, non-inflammatory cell death or alter neuron function. Both humans with SLE and animal models of SLE have shown structural and functional damage to the amygdala. The amygdala is a brain region important for processing the emotional relevance of stimuli in the environment. It also serves to modulate perception, attention, and memory to facilitate the processing and learning of relevant stimuli. Research has linked amygdala damage to deficits in emotional memory and emotional behavior. Individuals with SLE often exhibit emotional dysregulation, such as lability and depression; however, the behavioral impact of possible amygdala dysfunction has yet to be studied in this population. The purpose of this review is to 1) examine possible associations between SLE, anti-NMDAR antibodies, amygdala damage, and emotional processing deficits and 2) to identify the clinical, social, and treatment implications for individuals with SLE who suffer from deficits in emotional processing.

Adaptive coded-aperture imaging with subpixel superresolution.

We describe an adaptive coded-aperture imager operating in the midwave IR. This consists of a coded-aperture mask, a set of optics, and a 4k×4k focal plane array (FPA). This system can produce images with a resolution better than the detector pixel limit by combining multiple frames of data recorded with different coding. This superresolution capability has been demonstrated both in the laboratory and with targets placed outside, the highest resolution being one-half of the FPA pixel pitch.

Retrospective study of sensitivity and specificity of EEG in the elderly compared with younger age groups.

Following a recommendation from a Good Practice Guide published in Scotland stating that EEG should not be routinely used in the diagnosis of epilepsy in the elderly, we conducted a retrospective study to ascertain the effects this recommendation had. We found that predating the recommendation, there had already been a decline in the use of EEG in people aged 65 and over. Detailed examination of a 3.5-year epoch which straddled 2 years before the recommendation and 1.5 years after its publication revealed no evidence of a change in the type of referrals but just in the number of referrals. Comparison with 2 younger cohorts showed that EEG in the elderly had the same specificity and sensitivity as in the younger age groups and was of particular use in picking up previously unsuspected non-convulsive status. We conclude the EEG remains an important diagnostic adjunct in the elderly.

Androgen-dependent mammary carcinogenesis in rats transgenic for the Neu proto-oncogene.

Transgenic rats were created with overexpression of the Neu proto-oncogene in the mammary gland of both sexes, yet only males developed mammary cancer in an androgen-dependent fashion. Transgenic females only developed mammary cancer if treated with androgens. These tumors were positive for androgen receptor (AR), but negative for estrogen and progesterone receptors. Extensive analysis failed to detect mutations anywhere within the neu transgene from mammary carcinomas. Established mammary carcinomas eventually escaped their dependency on androgens. Transgenic long-term gonadectomized rats did not develop mammary cancer, but Neu overexpression stimulated the growth of their mammary glands. Our results suggest crosstalk between the Neu proto-oncogene and AR signaling pathways in the growth of both the normal and cancerous mammary epithelium.

Changes in Distilled Spirits Sales and Preferences During COVID-19.

OBJECTIVE: COVID-19 and the associated policies created a large shift in alcohol sales. A change in availability and consumer preferences caused a shift from wholesale to retail sales in many areas. This study estimates the magnitude of the changes in wholesale and retail sales, and the persistence of these changes. METHOD: Highly detailed sales data are used to analyze trends in distilled spirts sales during the COVID-19 pandemic in the state of Idaho. A total of 810,000 unique observations that cover 58 types of distilled spirts are used in a regression analysis to find the determinants of distilled spirts sales. RESULTS: In March 2020, there was a 27.2% increase in sales compared with the previous March because of a 42.3% increase in retail sales and a 41.1% decrease in wholesale sales. Increased sales continued until August 2020. The regression analysis shows changes in demand during 2020 varied by the type of distilled spirits; demand increased more for distilled spirts types with higher ethanol early in March 2020, but from April to July demand increased more for expensive distilled spirts with a lower ethanol level. CONCLUSIONS: Examination of the types of distilled spirts purchased during the early stages of the pandemic shows us that consumers bought different types of distilled spirts for various characteristics, such as price and ethanol level. We find that consumers initially purchased cheap distilled spirts with high alcohol content. By August, distilled spirts sales were similar to previous years. These results will be useful to policymakers in determining the effects of distilled spirts restrictions.

Characterization of Cytochrome P450s with Key Roles in Determining Herbicide Selectivity in Maize.

Safeners such as metcamifen and benoxacor are widely used in maize to enhance the selectivity of herbicides through the induction of key detoxifying enzymes, notably cytochrome P450 monooxygenases (CYPs). Using a combination of transcriptomics, proteomics, and functional assays, the safener-inducible CYPs responsible for herbicide metabolism in this globally important crop have been identified. A total of 18 CYPs belonging to clans 71, 72, 74, and 86 were safener-induced, with the respective enzymes expressed in yeast and screened for activity toward thiadiazine (bentazon), sulfonylurea (nicosulfuron), and triketone (mesotrione and tembotrione) chemistries. Herbicide metabolism was largely restricted to family CYP81A members from clan 71, notably CYP81A9, CYP81A16, and CYP81A2. Quantitative transcriptomics and proteomics showed that CYP81A9/CYP81A16 were dominant enzymes in safener-treated field maize, whereas only CYP81A9 was determined in sweet corn. The relationship between CYP81A sequence and activities were investigated by splicing CYP81A2 and CP81A9 together as a series of recombinant chimeras. CYP81A9 showed wide ranging activities toward the three herbicide chemistries, while CYP81A2 uniquely hydroxylated bentazon in multiple positions. The plasticity in substrate specificity of CYP81A9 toward multiple herbicides resided in the second quartile of its N terminal half. Further phylogenetic analysis of CYP81A9 showed that the maize enzyme was related to other CYP81As linked to agrochemical metabolism in cereals and wild grasses, suggesting this clan 71 CYP has a unique function in determining herbicide selectivity in arable crops.