Evolution and regulatory logic of an enhancer underlying a novel pigmentation pattern in Drosophila wings

Novel morphological traits originate largely from the novel expression patterns of genes controlled by enhancers during development. Enhancers bind and integrate the spatial-temporal activity of transcription factors, and their combinatorial interplay determines the time, location and levels of transcriptional output. New enhancers can arise through enhancer co-option by reusing some of the regulatory information from a preexisting enhancer. While enhancer co-option is thought to be a fast and likely way to evolve new enhancers, its genetic and molecular mechanisms still remain elusive. In this context, this thesis investigates the genetic origin of a novel enhancer and the regulatory logic underlying its function. I used the spot enhancer of the gene yellow as a model, which underlies the evolution of a morphological trait, the wing spot in Drosophila biarmipes. I sought to understand how the novel spot enhancer has evolved and what regulatory logic governs its function. Specifically: In the first chapter, I examined the evolutionary mechanism of spot enhancer in the context of the preexisting wing blade enhancer. By revisiting the entire D. biarmipes yellow 5´ region with a comprehensive and quantitative method, I mapped the full activities of the novel spot and preexisting wing blade enhancers to a much larger region (3.5 kb) than previously described (1.1 kb together). Within the region, the regulatory information necessary and sufficient for the spot activity was inseparable from, and extensively overlapping with the wing blade activity. Further dissection of the shared core region revealed a pleiotropic binding site that contributed to both activities by regulating the local chromatin accessibility. I therefore confirmed that the novel spot activity originates from the co-option of the preexisting wing blade activity. The pleiotropic site for chromatin accessibility suggests a possible model where a new enhancer could evolve by co-option of chromatin accessibility input from the ancestral element, and that might facilitate the emergence and diversification of morphological traits. In the second chapter, I investigated how the various aspects of regulatory information encoded in the spot enhancer sequences influenced its activity. Through introducing systematic mutations along the enhancer sequences and implementing a quantitative framework, the spatial activities on the wing of all the mutant enhancers were measured. The analysis showed an unexpected density of regulatory information within the spot enhancer. Moreover, it reveals an unanticipated regulatory logic underlying the activity of this enhancer and how it reads the wing trans-regulatory landscape to encode a spatial pattern. The gene yellow is required for black pigment production and its expression in late pupal stage prefigures the adult wing spot pigmentation pattern. Therefore, understanding the dynamics of yellow expression is essential to elucidate the process of yellow enhancer regulation as well as pigment formation during development. Chapter three investigates the process of pigment formation in space and time using the pigment gene yellow in D. melanogaster. Firstly, a fluorescent protein-tagged yellow allele was generated, then the dynamics of yellow expression and cellular localization in relationship to the process of pigment formation was examined during development. It was found that yellow is expressed in a few neurons in the brain and the ventral nerve chord from the second larval instar to adult stage, indicating a neuro-developmental function of yellow. In addition, the results mainly showed how yellow expression in the adult cuticle is determined by regulated developmental processes affecting the body color, and suggested a structural role of Yellow in the establishment of pigmentation patterns

Identification and Expression Analysis of an Atypical Alkaline Phosphatase in Emiliania huxleyi

Emiliania huxleyi, a cosmopolitan coccolithophore in the modern ocean, plays an important role in the carbon cycle and local climate feedback as it can form extensive blooms, calcify, and produce dimethylsulfoniopropionate (DMSP) leading to the generation of dimethyl sulfide (DMS) which affects climate when oxidized in the atmosphere. It is known to be able to utilize dissolved organic phosphorus (DOP) by expressing a specific type of alkaline phosphatase (EHAP1) under phosphorus-limited conditions. In this study, we identified a new alkaline phosphatase (EH-PhoAaty) in this species, which we found belongs to the newly classified PhoAaty family. The expression of this atypical phosphatase was up-regulated under P-depleted conditions at both the transcriptional and translational levels, suggesting that E. huxleyi is able to express this AP to cope with phosphorus limitation. Comparative analysis revealed different transcriptional expression dynamics between eh-PhoAaty and ehap1, although both genes exhibited inducible expression under phosphate deficiency. In addition, after AP activity was eliminated by using EDTA to chelate metal ions, we found that AP activity was recovered with the supplement of Ca2+ and Zn2+, indicative of the adoption of Ca2+ as the cofactor under Zn-P co-limited conditions, likely a result of adaptation to oceanic environments where Zn2+ is often limiting

Survival in Patients With Metastatic Prostate Cancer Undergoing Radiotherapy: The Importance of Prostate-Specific Antigen-Based Stratification

ObjectivesTo explore the effectiveness of radiotherapy in mPCa patients with different PSA stratifications based on the cancer database of a large population.BackgroundScreening criteria for patients with metastatic prostate cancer, who are candidates for radiotherapy, are rarely reported.Patients and MethodsWe identified 22,604 patients with metastatic prostate cancer in the Surveillance, Epidemiology, and End Results database and divided them into a radiotherapy group and a control group. Patients with metastatic prostate cancer were divided into subgroups according to their levels of prostate-specific antigen to evaluate the efficacy of radiotherapy. They were also divided into six subgroups according to their prostate-specific antigen levels. We used multivariate Cox analysis to evaluate overall survival and cancer-specific survival. After 1:1 propensity score matching, Kaplan-Meier analysis was used to explore the difference in overall survival and cancer-specific survival in the radiotherapy and control group.ResultsIn all, 5,505 patients received radiotherapy, compared to 17,099 in the control group. In the multivariate Cox analysis, radiotherapy improved overall survival (hazard ratio [HR]: 0.730, 95% confidence interval [CI]: 0.636–0.838; P<0.001) and cancer-specific survival (HR: 0.764, 95% CI: 0.647–0.903; P=0.002) in patients with a PSA level of 4–10 ng/mL. Similar results were obtained by Kaplan-Meier analysis after 1:1 propensity score matching. In patients with prostate-specific antigen levels between 4–10 ng/mL, the overall survival (P<0.001) and cancer-specific survival (P<0.05) in the radiotherapy group was significantly better than those in the control group.ConclusionThe result of this large population-based study shows that rigorous selection of appropriate metastatic prostate cancer patients for radiotherapy can benefit prognosis significantly. This can be the basis for future prospective trials

Comparative metatranscriptomic profiling and microRNA sequencing to reveal active metabolic pathways associated with a dinoflagellate bloom.

Harmful algal blooms (HABs) have increased as a result of global climate and environmental changes, exerting increasing impacts on the aquatic ecosystem, coastal economy, and human health. Despite great research efforts, our understanding on the drivers of HABs is still limited in part because HAB species’ physiology is difficult to probe in situ. Here, we used molecular ecological analyses to characterize a dinoflagellate bloom at Xiamen Harbor, China. Prorocentrum donghaiense was identified as the culprit, which nutrient bioassays showed were not nutrient-limited. Metatranscriptome profiling revealed that P. donghaiense highly expressed genes related to N- and P-nutrient uptake, phagotrophy, energy metabolism (photosynthesis, oxidative phophorylation, and rhodopsin) and carbohydrate metabolism (glycolysis/gluconeogenesis, TCA cycle and pentose phosphate) during the bloom. Many genes in P. donghaiense were up-regulated at night, including phagotrophy and environmental communication genes, and showed active expression in mitosis. Eight microbial defense genes were up-regulated in the bloom compared with previously analyzed laboratory cultures. Furthermore, 76 P. donghaiense microRNA were identified from the bloom, and their target genes exhibited marked differences in amino acid metabolism between the bloom and cultures and the potential of up-regulated antibiotic and cell communication capabilities. These findings, consistent with and complementary to recent reports, reveal major metabolic processes in P. donghaiense potentially important for bloom formation and provide a gene repertoire for developing bloom markers in future research

Metatranscriptomic Signatures Associated With Phytoplankton Regime Shift From Diatom Dominance to a Dinoflagellate Bloom

Diatoms and dinoflagellates dominate coastal marine phytoplankton communities as major players of marine biogeochemical cycles and their seasonal succession often leads to harmful algal blooms (HABs). What regulates their respective dominances and the development of the HABs remains elusive. Here we conducted time-sequential metatranscriptomic profiling on a natural assemblage that evolved from diatom dominance to a dinoflagellate bloom to interrogate the underlying major metabolic and ecological drivers. Data reveals similarity between diatoms and dinoflagellates in exhibiting high capacities of energy production, nutrient acquisition, and stress protection in their respective dominance stages. The diatom-to-dinoflagellate succession coincided with an increase in turbidity and sharp declines in silicate and phosphate availability, concomitant with the transcriptomic shift from expression of silicate uptake and urea utilization genes in diatoms to that of genes for light harvesting, diversified phosphorus acquisition and autophagy-based internal nutrient recycling in dinoflagellates. Furthermore, the diatom-dominant community featured strong potential to carbohydrate metabolism and a strikingly high expression of trypsin potentially promoting frustule building. In contrast, the dinoflagellate bloom featured elevated expression of xanthorhodopsin, and antimicrobial defensin genes, indicating potential importance of energy harnessing and microbial defense in bloom development. This study sheds light on mechanisms potentially governing diatom- and dinoflagellate-dominance and regulating bloom development in the natural environment and raises new questions to be addressed in future studies

Comparative metatranscriptomic profiling and microRNA sequencing to reveal active metabolic pathways associated with a dinoflagellate bloom

Abstract(#br)Harmful algal blooms (HABs) have increased as a result of global climate and environmental changes, exerting increasing impacts on the aquatic ecosystem, coastal economy, and human health. Despite great research efforts, our understanding on the drivers of HABs is still limited in part because HAB species’ physiology is difficult to probe in situ . Here, we used molecular ecological analyses to characterize a dinoflagellate bloom at Xiamen Harbor, China. Prorocentrum donghaiense was identified as the culprit, which nutrient bioassays showed were not nutrient-limited. Metatranscriptome profiling revealed that P. donghaiense highly expressed genes related to N- and P-nutrient uptake, phagotrophy, energy metabolism (photosynthesis, oxidative phophorylation, and rhodopsin) and carbohydrate metabolism (glycolysis/gluconeogenesis, TCA cycle and pentose phosphate) during the bloom. Many genes in P. donghaiense were up-regulated at night, including phagotrophy and environmental communication genes, and showed active expression in mitosis. Eight microbial defense genes were up-regulated in the bloom compared with previously analyzed laboratory cultures. Furthermore, 76 P. donghaiense microRNA were identified from the bloom, and their target genes exhibited marked differences in amino acid metabolism between the bloom and cultures and the potential of up-regulated antibiotic and cell communication capabilities. These findings, consistent with and complementary to recent reports, reveal major metabolic processes in P. donghaiense potentially important for bloom formation and provide a gene repertoire for developing bloom markers in future research

Metformin improves polycystic ovary syndrome in mice by inhibiting ovarian ferroptosis

Background and objectivePCOS is a common metabolic disorder in women of reproductive age, which pathogenesis is very complex. The role of ferroptosis in PCOS is a novel finding, and the mechanistic studies are not clear. Metformin is a commonly used drug of PCOS but few studies on whether metformin can improve the follicle development and ovarian function in PCOS. We aims to use PCOS mouse model to study the effect of metformin on PCOS based on the ovarian function and explored the regulation of metformin in PCOS mice by intervening in ferroptosis pathway.Materials and methodsC57 BL/6J female mice aged 4-5 weeks were purchased and gavaged with letrozole (1 mg/kg/day) combined with high-fat diet for 21days to establish PCOS model, and control group was set up. After modeling, the mice were divided into PCOS model group and metformin treatment group (Met) (n=6).The Met group were gavaged metformin (200 mg/kg/day) for 28 days. The body weight, estrous cycle, glucose tolerance test (OGTT)and insulin resistance test (ITT) were monitored. Then, The mice were euthanized to collect serum and ovaries. Elisa was used to detect changes in related serum hormones (E2, LH, FSH, TP). Ovaries used for molecular biology experiments to detect changes in GPX4, SIRT3, AMPK/p-AMPK, and mTOR/p-mTOR by Western blot and qPCR.ResultsCompared with the model group mice, body weight was significantly reduced, and their estrous cycle was restored in Met group. The results of OGTT and ITT showed an improvment of glucose tolerance and insulin resistance. Morphological results showed that after metformin treatment, polycystic lesions in ovaries were reduced, the ovarian function was restored, and the expressions of SIRT3 and GPX4 were elevated. WB results demonstrated that the expressions of p-mTOR and p-AMPK in ovaries were significantly reduced in Model group, but reversed in MET group.ConclusionOur study confirmed metformin could not only improve body weight and metabolism disorders, but also improve ovarian dysfunction in PCOS mice.In addition, we explored metformin could regulate ferroptosis to improve PCOS via the SIRT3/AMPK/mTOR pathway. Our study complements the mechanisms by which metformin improves PCOS

Electroacupuncture Alleviates Neuroinflammation by Inhibiting the HMGB1 Signaling Pathway in Rats with Sepsis-Associated Encephalopathy

Sepsis-Associated Encephalopathy (SAE) is common in sepsis patients, with high mortality rates. It is believed that neuroinflammation is an important mechanism involved in SAE. High mobility group box 1 protein (HMGB1), as a late pro-inflammatory factor, is significantly increased during sepsis in different brain regions, including the hippocampus. HMGB1 causes neuroinflammation and cognitive impairment through direct binding to advanced glycation end products (RAGE) and Toll-like receptor 4 (TLR4). Electroacupuncture (EA) at Baihui (GV20) and Zusanli (ST36) is beneficial for neurological diseases and experimental sepsis. Our study used EA to treat SAE induced by lipopolysaccharide (LPS) in male Sprague–Dawley rats. The Y maze test was performed to assess working memory. Immunofluorescence (IF) and Western blotting (WB) were used to determine neuroinflammation and the HMGB1 signaling pathway. Results showed that EA could improve working memory impairment in rats with SAE. EA alleviated neuroinflammation by downregulating the hippocampus’s HMGB1/TLR4 and HMGB1/RAGE signaling, reducing the levels of pro-inflammatory factors, and relieving microglial and astrocyte activation. However, EA did not affect the tight junctions’ expression of the blood–brain barrier (BBB) in the hippocampus

A Fatal Pneumonia due to Coinfection of Pseudomonas putida and Staphylococcus pseudintermedius in a Laboratory Beagle Dog

Background: Pseudomonas putida (P. putida) is widely distributed in the environment, and sometimes caused nosocomial infections in human beings, but no case of infection has been reported in beagle dogs. Staphylococcus pseudintermedius (S. pseudintermedius) is a natural cutaneous bacterium in dogs and occasionally causes purulent infections of the skin and rarely causes pneumonia. Both bacteria are opportunistic pathogens. Dogs, even well-controlled laboratory beagle dogs, maybe infected by the bacterium in certain conditions like this report. In order to provide information and give suggestion to veterinarians involved in dogs study, a complete profile of the coinfection was drawn in this report.Case: It is presented a case of an 8-month-old beagle dog, weighing 6 kg that suffered from coinfection of P. putida and S. pseudintermedius during a treatment of chemotherapy. The animal was confirmed as normal by appearance, physical examination, and laboratory tests before arrival according to the applicable guidelines. After 14-day acclimation period, the animal was administrated with a tyrosinase inhibitor once daily via oral gavage. From Day 8, coughing, decreased activity, hyporeflexia, squinting, shortness of breath (abdominal breathing), and discharge around the nose as well as crackles in the lung and rapid heart rate were noted. Since the poor conditions progressed rapidly and have not been improved by treatment of ceftriaxone and dexamethasone. On Day 9, the animal was euthanized for humanitarian reasons due to rapid progress and poor condition. To define the pathogen, hilar lymph node and thoracic swab were collected for bacteria isolation and purification in special mediums, and at last characterized by Gram staining and 16s rRNA gene sequence analysis and positive PCR-restriction fragment length polymorphism. In clinical pathological examination, an increase in WBC, neutrophils, lymphocytes, monocytes, cholesterol, triglyceride, total protein, globulin, and lactate dehydrogenase, as well as a decrease in RBC, hemoglobin, hematocrit, platelets, sodium ion, chloride ion, and albumin were noted. At necropsy, dark red and enlarged lymph nodes were noted in the hilum of lung, multiple abscesses with yellow pus and multifocal hemorrhage were noted in the lung, and a large amount of frothy yellow fluid were noted in the trachea. In pathological examinations, severe neutrophilic inflammation, diffuse and moderate macrophage aggregation, mild hemorrhage, and moderate alveolar emphysema were noted in the lung, and severe sinusoidal stasis were noted in portal lymph nodes.Discussion: The current case presented a profile of the appearance, treatment, hematological examination, coagulation examination, clinical chemistry, macroscopic and histological changes in the lung. Multiple purulent abscesses, infiltration of neutrophils, macrophage, and hemorrhage, were correlated to the increase in WBC, neutrophils, lymphocytes, and monocytes, and the decrease in RBC, hemoglobin, hematocrit, and platelets. In the coagulation examination, an increase in Fbg concentration was noted. This change may be induced by the coagulase effect of the S. pseudintermedius, yet no effect on PT or APTT was noted, indicating the coagulation function has not been affected. In the clinical chemistry, the increase of creatine kinase and lactate dehydrogenase may indicate tissue cell damages. Significant increase of globulin may be caused by the inflammatory status. In conclusion, the findings in this case indicate that both Pseudomonas putida and Staphylococcus pseudintermedius can induce infections in laboratory beagle dogs under certain conditions, and might result in a fatal pneumonia which could progress very fast within several days

Energy risk assessment based on PCA-LZNMAP model

10 risk factors are obtained from the view of energy risk management. The weight of each risk factor is decided by the sequence of PCA-LZNMAP model. Risk coordinate diagram, which intuitively reflects the attention degree of the energy risk factors and the measures which should be taken, was drawn based on these weights. At the same time, it is concluded that the evaluation value of energy risk is 6.33465 in 2008, and the state is between attention and danger