PHOSPHORUS-STARVATION TOLERANCE 1 (OsPSTOL1) is prevalent in upland rice and enhances root growth and hastens low phosphate signaling in wheat.

PHOSPHORUS-STARVATION TOLERANCE 1 (OsPSTOL1) is a variably present gene that benefits crown root growth and phosphorus (P) sufficiency in rice (Oryza sativa). To explore the ecophysiological importance of this gene, we performed a biogeographic survey of landraces and cultivars, confirming that functional OsPSTOL1 alleles prevail in low nutrient and drought-prone rainfed ecosystems, whereas loss-of-function and absence haplotypes predominate in control-irrigated paddy varieties of east Asia. An evolutionary history analysis of OsPSTOL1 and related genes in cereal, determined it and other genes are kinase-only domain derivatives of membrane-associated receptor like kinases. Finally, to evaluate the potential value of this kinase of unknown function in another Gramineae, wheat (Triticum aestivum) lines overexpressing OsPSTOL1 were evaluated under field and controlled low P conditions. OsPSTOL1 enhances growth, crown root number, and overall root plasticity under low P in wheat. Survey of root and shoot crown transcriptomes at two developmental stages identifies transcription factors that are differentially regulated in OsPSTOL1 wheat that are similarly controlled by the gene in rice. In wheat, OsPSTOL1 alters the timing and amplitude of regulators of root development in dry soils and hastens induction of the core P-starvation response. OsPSTOL1 and related genes may aid more sustainable cultivation of cereal crops.

Arabidopsis bZIP11 Is a Susceptibility Factor During Pseudomonas syringae Infection.

The induction of plant nutrient secretion systems is critical for successful pathogen infection. Some bacterial pathogens (e.g., Xanthomonas spp.) use transcription activator-like (TAL) effectors to induce transcription of SWEET sucrose efflux transporters. Pseudomonas syringae pv. tomato strain DC3000 lacks TAL effectors yet is able to induce multiple SWEETs in Arabidopsis thaliana by unknown mechanisms. Because bacteria require other nutrients in addition to sugars for efficient reproduction, we hypothesized that Pseudomonas spp. may depend on host transcription factors involved in secretory programs to increase access to essential nutrients. Bioinformatic analyses identified the Arabidopsis basic-leucine zipper transcription factor bZIP11 as a potential regulator of nutrient transporters, including SWEETs and UmamiT amino acid transporters. Inducible downregulation of bZIP11 expression in Arabidopsis resulted in reduced growth of P. syringae pv. tomato strain DC3000, whereas inducible overexpression of bZIP11 resulted in increased bacterial growth, supporting the hypothesis that bZIP11-regulated transcription programs are essential for maximal pathogen titer in leaves. Our data are consistent with a model in which a pathogen alters host transcription factor expression upstream of secretory transcription networks to promote nutrient efflux from host cells.[Formula: see text] Copyright © 2021 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Sugar flux and signaling in plant-microbe interactions.

Plant breeders have developed crop plants that are resistant to pests, but the continual evolution of pathogens creates the need to iteratively develop new control strategies. Molecular tools have allowed us to gain deep insights into disease responses, allowing for more efficient, rational engineering of crops that are more robust or resistant to a greater number of pathogen variants. Here we describe the roles of SWEET and STP transporters, membrane proteins that mediate transport of sugars across the plasma membrane. We discuss how these transporters may enhance or restrict disease through controlling the level of nutrients provided to pathogens and whether the transporters play a role in sugar signaling for disease resistance. This review indicates open questions that require further research and proposes the use of genome editing technologies for engineering disease resistance.

Fertility induction in hypogonadotropic hypogonadal men.

Men with hypogonadotropic hypogonadism (HH) are typically azoospermic, and yet HH is one of the few treatable forms of male infertility. Sperm induction protocols using gonadotrophins aim to replicate the natural endocrine control of spermatogenesis. Previously virilised men with adult-onset HH and normal testicular volume respond well to monotherapy in which human chorionic gonadotrophin (hCG) acts as a long-acting LH-analogue stimulating spermatogenesis. However, this approach is rarely successful for men with congenital HH (CHH) (eg, Kallmann syndrome), for whom combined gonadotrophin therapy (hCG + follicle-stimulating hormone [FSH]) is an absolute requirement to maximise fertility potential. Key baseline predictors of successful spermatogenesis-induction include prior spontaneous testicular development (ie, testicular volume [TV] > 4 mL), serum inhibin B (IB ) concentration >60 pg/mL and no history of maldescended testes (cryptorchidism).

Proteomic Analysis of GLUT4 Storage Vesicles Reveals Tumor Suppressor Candidate 5 (TUSC5) as a Novel Regulator of Insulin Action in Adipocytes.

Insulin signaling augments glucose transport by regulating glucose transporter 4 (GLUT4) trafficking from specialized intracellular compartments, termed GLUT4 storage vesicles (GSVs), to the plasma membrane. Proteomic analysis of GSVs by mass spectrometry revealed enrichment of 59 proteins in these vesicles. We measured reduced abundance of 23 of these proteins following insulin stimulation and assigned these as high confidence GSV proteins. These included established GSV proteins such as GLUT4 and insulin-responsive aminopeptidase, as well as six proteins not previously reported to be localized to GSVs. Tumor suppressor candidate 5 (TUSC5) was shown to be a novel GSV protein that underwent a 3.7-fold increase in abundance at the plasma membrane in response to insulin. siRNA-mediated knockdown of TUSC5 decreased insulin-stimulated glucose uptake, although overexpression of TUSC5 had the opposite effect, implicating TUSC5 as a positive regulator of insulin-stimulated glucose transport in adipocytes. Incubation of adipocytes with TNFα caused insulin resistance and a concomitant reduction in TUSC5. Consistent with previous studies, peroxisome proliferator-activated receptor (PPAR) γ agonism reversed TNFα-induced insulin resistance. TUSC5 expression was necessary but insufficient for PPARγ-mediated reversal of insulin resistance. These findings functionally link TUSC5 to GLUT4 trafficking, insulin action, insulin resistance, and PPARγ action in the adipocyte. Further studies are required to establish the exact role of TUSC5 in adipocytes.

Direction pathway analysis of large-scale proteomics data reveals novel features of the insulin action pathway.

MOTIVATION: With the advancement of high-throughput techniques, large-scale profiling of biological systems with multiple experimental perturbations is becoming more prevalent. Pathway analysis incorporates prior biological knowledge to analyze genes/proteins in groups in a biological context. However, the hypotheses under investigation are often confined to a 1D space (i.e. up, down, either or mixed regulation). Here, we develop direction pathway analysis (DPA), which can be applied to test hypothesis in a high-dimensional space for identifying pathways that display distinct responses across multiple perturbations. RESULTS: Our DPA approach allows for the identification of pathways that display distinct responses across multiple perturbations. To demonstrate the utility and effectiveness, we evaluated DPA under various simulated scenarios and applied it to study insulin action in adipocytes. A major action of insulin in adipocytes is to regulate the movement of proteins from the interior to the cell surface membrane. Quantitative mass spectrometry-based proteomics was used to study this process on a large-scale. The combined dataset comprises four separate treatments. By applying DPA, we identified that several insulin responsive pathways in the plasma membrane trafficking are only partially dependent on the insulin-regulated kinase Akt. We subsequently validated our findings through targeted analysis of key proteins from these pathways using immunoblotting and live cell microscopy. Our results demonstrate that DPA can be applied to dissect pathway networks testing diverse hypotheses and integrating multiple experimental perturbations. AVAILABILITY AND IMPLEMENTATION: The R package 'directPA' is distributed from CRAN under GNU General Public License (GPL)-3 and can be downloaded from: http://cran.r-project.org/web/packages/directPA/index.html CONTACT: jean.yang@sydney.edu.au SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Re-fraction: a machine learning approach for deterministic identification of protein homologues and splice variants in large-scale MS-based proteomics.

A key step in the analysis of mass spectrometry (MS)-based proteomics data is the inference of proteins from identified peptide sequences. Here we describe Re-Fraction, a novel machine learning algorithm that enhances deterministic protein identification. Re-Fraction utilizes several protein physical properties to assign proteins to expected protein fractions that comprise large-scale MS-based proteomics data. This information is then used to appropriately assign peptides to specific proteins. This approach is sensitive, highly specific, and computationally efficient. We provide algorithms and source code for the current version of Re-Fraction, which accepts output tables from the MaxQuant environment. Nevertheless, the principles behind Re-Fraction can be applied to other protein identification pipelines where data are generated from samples fractionated at the protein level. We demonstrate the utility of this approach through reanalysis of data from a previously published study and generate lists of proteins deterministically identified by Re-Fraction that were previously only identified as members of a protein group. We find that this approach is particularly useful in resolving protein groups composed of splice variants and homologues, which are frequently expressed in a cell- or tissue-specific manner and may have important biological consequences.

Comparison of 3D ultrasound, 2D ultrasound and 3D Doppler in the diagnosis of endometrial carcinoma in patients with uterine bleeding: A systematic review and meta-analysis.

Endometrial cancer is a common malignancy affecting women worldwide. Usually, it clinically manifests with uterine bleeding, although identical clinical manifestations occur in benign conditions as well, with several endometrial biopsies being conducted unnecessarily. Therefore, an accurate, non-invasive diagnostic test is needed for first-line assessment, so as unnecessary biopsies are limited as much as possible. This systematic review aims to assess the diagnostic accuracy of three-dimensional ultrasound, a relatively novel method in gynecologic assessment, compared to two-dimensional ultrasound and three-dimensional Doppler in the prediction of uterine malignancy in women with abnormal uterine bleeding. The accuracy of endometrial volume, as a diagnostic parameter assessed by three-dimensional ultrasound is compared to diagnostic parameters from the other two methods, namely endometrial thickness and 3D Doppler indices (vascularization index, flow index and vascularization flow index). Articles relevant to our research question were systematically sought in the Web of Science, Scopus and MEDLINE/PubMed databases and underwent rigorous evaluation for inclusion according to the PRISMA 2020 guidelines. Eligible studies were thoroughly assessed for risk of bias and relevant data was extracted and analyzed. Studies were heterogenous and extracted data varied from study to study. Data on endometrial volume was compared to other diagnostic parameters. Forest plots with pooled percentages and hierarchical summary receiver operating characteristic curves were constructed for each comparison. Relative sensitivity and specificity ratios were calculated for each comparison to test for statistical significance. Endometrial volume and thickness comparison showed sensitivity 83% for both parameters and specificity 75% and 69% respectively, with volume being more specific than thickness (p < 0.05). Endometrial volume and Doppler indices comparison showed that sensitivity was 73%, 82%, 81% and 82%, while specificity was 72%, 76%, 75% and 76% for endometrial volume, vascularization index, flow index and vascularization-flow index respectively. All three Doppler indices were significantly more sensitive in the diagnosis of malignancy compared to endometrial volume (p < 0.05) While endometrial thickness remains a reliable predictor of uterine malignancy, endometrial volume appears promising as a method with higher specificity and more reliable measurements. Similarly, vascular indices seem as competent and even more sensitive than endometrial volume as predictors, with the added advantage of semi-automated and reproducible measurements that reflect the whole organ. More comparative studies with standardized protocols should be established, so as reliable cut-off values can be determined and thus standardize and streamline the diagnostic algorithm via the implementation of the three-dimensional modalities in the settings that they are available.

Gene regulatory networks shape developmental plasticity of root cell types under water extremes in rice.

Understanding how roots modulate development under varied irrigation or rainfall is crucial for development of climate-resilient crops. We established a toolbox of tagged rice lines to profile translating mRNAs and chromatin accessibility within specific cell populations. We used these to study roots in a range of environments: plates in the lab, controlled greenhouse stress and recovery conditions, and outdoors in a paddy. Integration of chromatin and mRNA data resolves regulatory networks of the following: cycle genes in proliferating cells that attenuate DNA synthesis under submergence; genes involved in auxin signaling, the circadian clock, and small RNA regulation in ground tissue; and suberin biosynthesis, iron transporters, and nitrogen assimilation in endodermal/exodermal cells modulated with water availability. By applying a systems approach, we identify known and candidate driver transcription factors of water-deficit responses and xylem development plasticity. Collectively, this resource will facilitate genetic improvements in root systems for optimal climate resilience.

Quantitative proteomic analysis of the adipocyte plasma membrane.

The adipocyte is a key regulator of mammalian metabolism. To advance our understanding of this important cell, we have used quantitative proteomics to define the protein composition of the adipocyte plasma membrane (PM) in the presence and absence of insulin. Using this approach, we have identified a high confidence list of 486 PM proteins, 52 of which potentially represent novel cell surface proteins, including a member of the adiponectin receptor family and an unusually high number of hydrolases with no known function. Several novel insulin-responsive proteins including the sodium/hydrogen exchanger, NHE6 and the collagens III and VI were also identified, and we provide evidence of PM-ER association suggestive of a unique functional association between these two organelles in the adipocyte. Together these studies provide a wealth of potential therapeutic targets for the manipulation of adipocyte function and a valuable resource for metabolic research and PM biology.

Benign paroxysmal positional vertigo nurse-led follow-up clinic.

Patients with benign paroxysmal positional vertigo (BPPV) often require multiple appointments for treatment with Epley manoeuvres. Waiting times for medical follow up can be very long. To reduce waiting times and increase availability of ENT outpatients' appointments, a nurse-led dizziness clinic (NLDC) to follow up BPPV patients was established. Prospective audit of 99 consecutive patients attending the NLDC, at which patients are assessed and treated, was conducted. Non-responders are redirected for further medical review. 99 patients were seen in 200 appointments in the NLDC from July 2007 to May 2009. The mean time to NLDC was 16 days. 67 patients were discharged from the NLDC free of symptoms. Cost analysis revealed savings of £3,800. A survey of NLDC attendees revealed that the care they received was rated as excellent, very good or good by 92% of patients. In conclusion, the NLDC is an innovation which increases availability of ENT outpatient appointments. This is acceptable to patients and is a natural extension of the roles of ENT nurse practitioners which could be implemented in other ENT departments.

A Patient-Initiated Digital COVID-19 Contact Notification Tool (TellYourContacts): Evaluation Study.

BACKGROUND: Contact notification is a method used to control the spread of infectious disease. In this process, a patient who tests positive for an infectious disease and public health officials work to identify the patient's close contacts, notify them of their risk of possible exposure to the disease, and provide resources to facilitate the decreased spreading of disease. Contact notification can be done physically in person, via phone call, or digitally through the use of media such as SMS text messages and email. When alerts are made through the latter, it is called digital contact notification. OBJECTIVE: For this study, we aim to perform a preliminary evaluation of the use of the TellYourContacts website, a digital contact notification tool for COVID-19 that can be used confidentially and anonymously. We will gather information about the number of website users and message senders, the types of messages sent, and the geographic distribution of senders. METHODS: Patients who chose to get tested for COVID-19 and subsequently tested positive for the disease were alerted of their positive results through Curative Inc (a COVID-19 testing laboratory) and Healthvana (a results disclosure app). Included in the notification was a link to the TellYourContacts website and a message encouraging the person who tested positive for COVID-19 to use the website to alert their close contacts of exposure risk. Over the course of three months, from May 18, 2020, to August 17, 2020, we used Google Analytics and Microsoft Excel to record data on the number of website users and message senders, types of messages sent, and geographic distribution of the senders. RESULTS: Over the course of three months, 9130 users accessed the website and 1474 unique senders sent a total of 1957 messages, which included 1820 (93%) SMS text messages and 137 (7%) emails. Users sent messages from 40 US states, with the majority of US senders residing in California (49%). CONCLUSIONS: We set out to determine if individuals who test positive for COVID-19 will use the TellYourContacts website to notify their close contacts of COVID-19 exposure risk. Our findings reveal that, during the observation period, each unique sender sent an average of 1.33 messages. The TellYourContacts website offers an additional method that individuals can and will use to notify their close contacts about a recent COVID-19 diagnosis.

NDRG2, a novel regulator of myoblast proliferation, is regulated by anabolic and catabolic factors.

Skeletal muscle tissue undergoes adaptive changes in response to stress and the genes that control these processes are incompletely characterised. NDRG2 (N-myc downstream-regulated gene 2), a stress- and growth-related gene, was investigated in skeletal muscle growth and adaption. While NDRG2 expression levels were found to be up-regulated in both differentiated human and mouse myotubes compared with undifferentiated myoblasts, the suppression of NDRG2 in C2C12 myoblasts resulted in slowed myoblast proliferation. The increased expression levels of the cell cycle inhibitors, p21 Waf1/Cip1 and p27 Kip1, and of various muscle differentiation markers in NDRG2-deficient myoblasts indicate that a lack of NDRG2 promoted cell cycle exiting and the onset of myogenesis. Furthermore, the analysis of NDRG2 regulation in C2C12 myotubes treated with catabolic and anabolic agents and in skeletal muscle from human subjects following resistance exercise training revealed NDRG2 gene expression to be down-regulated during hypertrophic conditions, and conversely, up-regulated during muscle atrophy. Together, these data demonstrate that NDRG2 expression is highly responsive to different stress conditions in skeletal muscle and suggest that the level of NDRG2 expression may be critical to myoblast growth and differentiation.

Priorities for research in miscarriage: a priority setting partnership between people affected by miscarriage and professionals following the James Lind Alliance methodology.

OBJECTIVES: To identify and prioritise important research questions for miscarriage. DESIGN: A priority setting partnership using prospective surveys and consensus meetings following methods advocated by the James Lind Alliance. SETTING: UK. PARTICIPANTS: Women and those affected by miscarriage working alongside healthcare professionals. RESULTS: In the initial survey, 1093 participants (932 women who have experienced miscarriage, 8 partners, 17 family members, friends or colleagues, 104 healthcare professionals and eight charitable organisations) submitted 3279 questions. A review of existing literature identified a further 64. Non-questions were removed, and the remaining questions were categorised and summarised into 58 questions. In an interim electronic survey, 2122 respondents chose their top 10 priorities from the 58 summary questions. The 25 highest ranked in the survey were prioritised at a final face-to-face workshop. In summary, the top 10 priorities were ranked as follows: research into preventative treatment, emotional aspects in general, investigation, relevance of pre-existing medical conditions, emotional support as a treatment, importance of lifestyle factors, importance of genetic and chromosomal causes, preconception tests, investigation after different numbers of miscarriage and male causal factors. CONCLUSIONS: These results should be the focus of future miscarriage research. Presently, studies are being conducted to address the top priority; however, many other priorities, especially psychological and emotional support, are less well researched areas. We hope our results will encourage both researchers and funders to focus on these priorities.

What shape do UK trainees want their training to be? Results of a cross-sectional study.

OBJECTIVES: The British Government is acting on recommendations to overhaul postgraduate training to meet the needs of the changing population, to produce generalist doctors undergoing shorter broad-based training (Greenaway Review). Only 45 doctors in training were involved in the consultation process. This study aims to obtain a focused perspective on the proposed reforms by doctors in training from across specialities. DESIGN: Prospective, questionnaire-based cross-sectional study. SETTING/PARTICIPANTS: Following validation, a 31-item electronic questionnaire was distributed via trainee organisations and Postgraduate Local Education and Training Board (LETB) mailing lists. Throughout the 10-week study period, the survey was publicised on several social media platforms. RESULTS: Of the 3603 demographically representative respondents, 69% knew about proposed changes. Of the respondents, 73% expressed a desire to specialise, with 54% keen to provide general emergency cover. A small proportion (12%) stated that current training pathway length is too long, although 86% felt that it is impossible to achieve independent practitioner-level proficiency in a shorter period of time than is currently required. Opinions regarding credentialing were mixed, but tended towards disagreement. The vast majority (97%) felt credentialing should not be funded by doctors in training. Respondents preferred longer placement lengths with increasing career progression. Doctors in training value early generalised training (65%), with suggestions for further improvement. CONCLUSIONS: This is the first large-scale cross-specialty study regarding the Shape of Training Review. Although there are recommendations which trainees support, it is clear that one size does not fit all. Most trainees are keen to provide a specialist service on an emergency generalist background. Credentialing is a contentious issue; however, we believe removing aspects from curricula into post-Certificate of Completion of Training (CCT) credentialing programmes with shortened specialty training routes only degrades the current consultant expertise, and does not serve the population. Educational needs, not political winds, should drive changes in postgraduate medical education and all stakeholders should be involved.

Src homology 3-domain growth factor receptor-bound 2-like (endophilin) interacting protein 1, a novel neuronal protein that regulates energy balance.

To identify genes involved in the central regulation of energy balance, we compared hypothalamic mRNA from lean and obese Psammomys obesus, a polygenic model of obesity, using differential display PCR. One mRNA transcript was observed to be elevated in obese, and obese diabetic, P. obesus compared with lean animals and was subsequently found to be increased 4-fold in the hypothalamus of lethal yellow agouti (A(y)/a) mice, a murine model of obesity and diabetes. Intracerebroventricular infusion of antisense oligonucleotide targeted to this transcript selectively suppressed its hypothalamic mRNA levels and resulted in loss of body weight in both P. obesus and Sprague Dawley rats. Reductions in body weight were mediated by profoundly reduced food intake without a concomitant reduction in metabolic rate. Yeast two-hybrid screening, and confirmation in mammalian cells by bioluminescence resonance energy transfer analysis, demonstrated that the protein it encodes interacts with endophilins, mediators of synaptic vesicle recycling and receptor endocytosis in the brain. We therefore named this transcript Src homology 3-domain growth factor receptor-bound 2-like (endophilin) interacting protein 1 (SGIP1). SGIP1 encodes a large proline-rich protein that is expressed predominantly in the brain and is highly conserved between species. Together these data suggest that SGIP1 is an important and novel member of the group of neuronal molecules required for the regulation of energy homeostasis.

Effect of bacterial endotoxin and middle ear effusion on ciliary activity: implications for otitis media.

OBJECTIVES/HYPOTHESIS: Otitis media with effusion (OME) is the most common cause of childhood deafness. The pathogenesis is not fully understood, especially the reasons for failure of mucociliary clearance of the middle ear. It is not clear whether the cilia function normally in the middle ear and eustachian tube in the chronic phase of otitis media with effusion. However, impaired ciliary function in primary ciliary dyskinesia is known to be frequently associated with the development of otitis media with effusion. We hypothesized that endotoxin or the bacterial products in middle ear fluid in otitis media with effusion would adversely affect ciliary activity, thereby contributing to the pathogenesis of the disease. STUDY DESIGN: Laboratory-based study of human ciliary activity with reference to otitis media with effusion. METHODS: We have studied the activity of human adenoidal cilia under various conditions. Ciliary activity in the presence of Haemophilus influenzae endotoxin additions (at varying concentrations) to cultured adenoidal explants has been measured. In addition, ciliary activity of these explants was also observed after addition of middle ear effusion aspirated from patients. RESULTS: We have shown that endotoxin in concentrations far in excess of those found in the middle ear with chronic otitis media with effusion had no effect on ciliary activity. Furthermore, ciliary activity was completely unaffected by the presence of middle ear effusion. CONCLUSION: There is no evidence that ciliary activity is reduced by the constituents of middle ear fluid in chronic otitis media with effusion.

Reverse genetics in Chlamydomonas: a platform for isolating insertional mutants.

A method was developed to identify insertional mutants of Chlamydomonas reinhardtii disrupted for selected target genes. The approach relies on the generation of thousands of transformants followed by PCR-based screenings that allow for identification of strains harboring the introduced marker gene within specific genes of interest. Our results highlight the strengths and limitations of two independent screens that differed in the nature of the marker DNA used (PCR-amplified fragment containing the plasmid-free marker versus entire linearized plasmid with the marker) and in the strategies used to maintain and store transformants.