Palladium-Catalyzed Enantioselective [4+2] Cycloaddition of 4-Vinylbenzodioxinones with Barbiturate-Derived Alkenes: Con-struction of Chiral Spirobarbiturate-Chromanes.

In this paper, Pd-catalyzed [4+2] decarboxylative cycloaddition of 4-vinylbenzodioxinones with barbiturate-derived alkenes has been developed, leading to various spirobarbiturate-chromane derivatives in high yields with excellent diastereo- and enantioselectivities. The scale-up reaction and further derivation of the product were demonstrated. A plausible reaction mechanism was also proposed.

Spatial linear transformer and temporal convolution network for traffic flow prediction.

Accurately obtaining accurate information about the future traffic flow of all roads in the transportation network is essential for traffic management and control applications. In order to address the challenges of acquiring dynamic global spatial correlations between transportation links and modeling time dependencies in multi-step prediction, we propose a spatial linear transformer and temporal convolution network (SLTTCN). The model is using spatial linear transformers to aggregate the spatial information of the traffic flow, and bidirectional temporal convolution network to capture the temporal dependency of the traffic flow. The spatial linear transformer effectively reduces the complexity of data calculation and storage while capturing spatial dependence, and the time convolutional network with bidirectional and gate fusion mechanisms avoids the problems of gradient vanishing and high computational cost caused by long time intervals during model training. We conducted extensive experiments using two publicly available large-scale traffic data sets and compared SLTTCN with other baselines. Numerical results show that SLTTCN achieves the best predictive performance in various error measurements. We also performed attention visualization analysis on the spatial linear transformer, verifying its effectiveness in capturing dynamic global spatial dependency.

Contribution of the LAC Genes in Fruit Quality Attributes of the Fruit-Bearing Plants: A Comprehensive Review.

Laccase genes produce laccase enzymes that play a crucial role in the production of lignin and oxidation reactions within plants. Lignin is a complex polymer that provides structure and toughness to the cell walls of numerous fruit plants. The LAC genes that encode laccase enzymes play vital roles in plant physiology, including the synthesis of pigments like PA that contribute to the colors of fruits, and in defending against pathogens and environmental stresses. They are crucial for fruit development, ripening, structural maintenance in plants, and adaptation to various environmental factors. As such, these genes and enzymes are essential for plant growth and development, as well as for various biotechnological applications in environmental remediation and industrial processes. This review article emphasizes the significance of genes encoding laccase enzymes during fruit growth, specifically pertaining to the strengthening of the endocarp through lignification. This process is crucial for ensuring fruit defense and optimizing seed scattering. The information gathered in this article will aid breeders in producing future fruit-bearing plants that are resistant to disease, cost-effective, and nutrient-rich.

Ring Stripping, Ring Cutting, and Growth Regulators Promote Phase Change and Early Flowering in Pear Seedlings.

Hybrid breeding is the most important means of selecting pear (Pyrus) varieties, but a long juvenile period severely restricts the selection of new varieties. In this study, we used 'Yuluxiang' × 'Akituki' 4-year-old seedling trees to study the effects of plant growth regulators, ring stripping, and ring cutting on the promotion of phase change and flowering to assist in shortening the breeding cycle. A single application of 100 mg/kg 6-BA + 1000 mg/kg PP333 was most effective in promoting phase change and flowering. This treatment effectively inhibited the growth and thickening of annual shoots, significantly increased soluble sugar and protein contents in buds, increased the ABA content by 45.41%, decreased the IAA content by 7.35%, increased the expression of the flower-promoting genes FT and LFY by 2273.41% and 1153.71%, respectively, and decreased the expression of the flower-suppressing gene TFL1 by 74.92%. The flowering plant rate increased by 23.34% compared to the control. Both ring stripping and ring cutting were effective in promoting phase change and flowering, significantly increasing the flowering rate, inflorescence number, and the number of flowering plants. For improving the flowering rate, the ring-stripping treatment had the strongest effect and effectively inhibited the growth and thickening of annual shoots, while also significantly increasing the soluble sugar and protein contents in buds, reducing the contents of IAA and GA3 by 8.73% and 50.12%, respectively, increasing the expression of FT and LFY by 80.01% and 821.14%, respectively, and reducing the expression of the flower-suppressing gene TFL1 by 59.22%. In conclusion, ring stripping, ring cutting, and spraying of 100 mg/kg 6-BA + 1000 mg/kg PP333 were effective in promoting phase change and early flowering in seedling trees.

Inhibition of CtBP-Regulated Proinflammatory Gene Transcription Attenuates Psoriatic Skin Inflammation.

Psoriasis is a chronic immune-mediated disease characterized by excessive proliferation of epidermal keratinocytes and increased immune cell infiltration to the skin. Although it is well-known that psoriasis pathogenesis is driven by aberrant production of proinflammatory cytokines, the mechanisms underlying the imbalance between proinflammatory and anti-inflammatory cytokine expression are incompletely understood. In this study, we report that the transcriptional coregulators CtBP1 and 2 can transactivate a common set of proinflammatory genes both in the skin of imiquimod-induced mouse psoriasis model and in human keratinocytes and macrophages stimulated by imiquimod. We find that mice overexpressing CtBP1 in epidermal keratinocytes display severe skin inflammation phenotypes with increased expression of T helper type 1 and T helper type 17 cytokines. We also find that the expression of CtBPs and CtBP-target genes is elevated both in human psoriatic lesions and in the mouse imiquimod psoriasis model. Moreover, we were able to show that topical treatment with a peptidic inhibitor of CtBP effectively suppresses the CtBP-regulated proinflammatory gene expression and thus attenuates psoriatic inflammation in the imiquimod mouse model. Together, our findings suggest to our knowledge previously unreported strategies for therapeutic modulation of the immune response in inflammatory skin diseases.

Posaconazole and fluconazole prophylaxis during induction therapy for pediatric acute lymphoblastic leukemia.

PURPOSE: To assess and compare the efficacy and safety of posaconazole with fluconazole for the prevention of invasive fungal infections in children who were undergoing induction therapy for acute lymphoblastic leukemia (ALL). To develop an approach to predict invasive fungal infections in ALL patients who accepted posaconazole prophylaxis. METHODS: This was a single-center, retrospective cohort study of patients with newly diagnosed ALL, comparing invasive fungal infections in patients who received no prophylaxis, posaconazole prophylaxis, or fluconazole prophylaxis during induction therapy. A propensity score-weighted logistic regression model was used to adjust for confounders. Hepatotoxicity was assessed according to the Common Terminology Criteria for Adverse Events (CTCAE) criteria. RESULTS: Out of the 155 ALL patients, 60 received no prophylaxis, 70 received posaconazole prophylaxis, and 25 received fluconazole prophylaxis. Posaconazole prophylaxis reduced the odds of invasive fungal infections by > 60%, prolonged infection-free survival significantly, and did not increase the risk of hepatotoxicity. Additionally, we found that the combination of age at diagnosis, clinically documented bacterial infection in the first 15 days of induction therapy, and absolute neutrophil count (ANC) curve enabled significant prediction of the susceptibility to infections after receiving posaconazole prophylaxis. CONCLUSIONS: Our findings supported using targeted prophylaxis with posaconazole in ALL children undergoing induction chemotherapy. Age, clinically documented bacterial infection and ANC are important predictors of invasive fungal infections in patients with posaconazole prophylaxis.

C-terminal binding proteins 1 and 2 in traumatic brain injury-induced inflammation and their inhibition as an approach for anti-inflammatory treatment.

Traumatic brain injury (TBI) induces an acute inflammatory response in the central nervous system that involves both resident and peripheral immune cells. The ensuing chronic neuroinflammation causes cell death and tissue damage and may contribute to neurodegeneration. The molecular mechanisms involved in the maintenance of this chronic inflammation state remain underexplored. C-terminal binding protein (CtBP) 1 and 2 are transcriptional coregulators that repress diverse cellular processes. Unexpectedly, we find that the CtBPs can transactivate a common set of proinflammatory genes both in lipopolysaccharide-activated microglia, astrocytes and macrophages, and in a mouse model of the mild form of TBI. We also find that the expression of these genes is markedly enhanced by a single mild injury in both brain and peripheral blood leukocytes in a severity- and time-dependent manner. Moreover, we were able to demonstrate that specific inhibitors of the CtBPs effectively suppress the expression of the CtBP target genes and thus improve neurological outcome in mice receiving single and repeated mild TBIs. This discovery suggests new avenues for therapeutic modulation of the inflammatory response to brain injury.

Functional connection between the stereotyped behavior and the motor front area in children with autism.

OBJECT: Autism spectrum disorders (ASD) is characterized by stereotyped behavior, attention deficit and/or impaired sensory perception to external stimuli. Its neurobiological mechanisms remain unclear. In this study we examined the resting-state functional connectivity of the premotor area and investigated its correlation with behavioral variables to determine whether connectivity alterations can distinguish ASD from healthy controls. METHODS: 39 children with ASD and 42 healthy children with matched age, sex and intelligence were recruited. All the 81 subjects had behavioral index evaluation and underwent resting-state functional magnetic resonance imaging (fMRI) scans. After MRI data preprocessing, the left and right premotor areas were selected as region of interest (ROI) seeds to perform functional connectivity. Groups were compared, and the correlation between functional connectivity and behavioral indicators was analyzed. RESULTS: Compared with healthy controls, ASD children showed significantly increased functional connectivity between the left premotor area and the posterior cingulate gyrus or anterior lobe of wedge, but functional connectivity between the left premotor area and the left insular lobe was decreased (p < 0.05, FDR correction). In addition, the connectivity between the left premotor area and the left insular lobe was negatively correlated with the behavioral scores (p < 0.05). CONCLUSION: Imbalanced premotor functional connectivity may be one possible mechanism of stereotyped behavior in ASD.

CPP-E1A fusion peptides inhibit CtBP-mediated transcriptional repression.

The carboxyl-terminal binding proteins (CtBP) are transcriptional corepressors that regulate the expression of multiple epithelial-specific and pro-apoptotic genes. Overexpression of CtBP occurs in many human cancers where they promote the epithelial-to-mesenchymal transition, stem cell-like features, and cell survival, while knockdown of CtBP in tumor cells results in p53-independent apoptosis. CtBPs are recruited to their target genes by binding to a conserved PXDLS peptide motif present in multiple DNA-binding transcription factors. Disrupting the interaction between CtBP and its transcription factor partners may be a means of altering CtBP-mediated transcriptional repression and a potential approach for cancer therapies. However, small molecules targeting protein-protein interactions have traditionally been difficult to identify. In this study, we took advantage of the fact that CtBP binds to a conserved peptide motif to explore the feasibility of using peptides containing the PXDLS motif fused to cell-penetrating peptides (CPP) to inhibit CtBP function. We demonstrate that these peptides disrupt the ability of CtBP to interact with its protein partner, E1A, in an AlphaScreen assay. Moreover, these peptides can enter both lung carcinoma and melanoma cells, disrupt the interaction between CtBP and a transcription factor partner, and inhibit CtBP-mediated transcriptional repression. Finally, the constitutive expression of one such peptide, Pep1-E1A-WT, in a melanoma cell line reverses CtBP-mediated oncogenic phenotypes including proliferation, migration, and sphere formation and limits tumor growth in vivo. Together, our results suggest that CPP-fused PXDLS-containing peptides can potentially be developed into a research tool or therapeutic agent targeting CtBP-mediated transcriptional events in various biological pathways.

Altered amygdala-related structural covariance and resting-state functional connectivity in end-stage renal disease patients.

Depression and cognitive control deficits were frequently reported in concurrent end-stage renal disease (ESRD) patients. Neuroimaging studies indicated depression could be a risk factor for cognitive control deficits, and amygdala-related circuitry may play a critical role in this abnormal interaction. To investigate the potential relationship between depressive symptoms and cognitive control reduction in ESRD patients, T1-weighted and resting fMRI images were obtained in 29 ESRD patients and 29 healthy controls. Voxel-based morphometry (VBM), structural covariance (SC) analysis based on grey matter volume (GMV), and functional connectivity (FC) analysis were adopted. All subjects performed the Beck Depression Inventory (BDI) assessment and Stroop test. The patients also underwent blood biochemistry tests (urea, creatinine, phosphate, Ca2+, hematocrit, cystatin, hemoglobin). Compared with controls, GMV reductions were found mainly in the anterior cingulate cortex (ACC) and bilateral amygdala, and decreased SC was found between the amygdala and ACC in ESRD patients. This indicated that structural changes in the amygdala may be related to the GMV alterations in the ACC. Additionally, decreased FC between the amygdala and ACC was revealed in ESRD patients. Negative correlation was found between the FC of the amygdala-ACC and reaction delay during the Stroop test, but this correlation disappeared after controlling BDI. Stepwise regression analysis showed that the low level of hemoglobin was contributed to the reduced FC of the amygdala-ACC in ESRD patients. Our results demonstrated the abnormal interaction between depressive mood and cognitive control deficits in ESRD patients.

Abnormal interaction between cognitive control network and affective network in patients with end-stage renal disease.

End-stage renal disease (ESRD) is a common complicated disorder that is generally associated with an altered central nervous system and cognitive impairment. Neuroimaging studies have recorded aberrant brain circuits in patients with ESRD that were closely associated with abnormal clinical manifestations. However, whether the altered interaction was within and/or between these circuits is largely unclear. We investigated brain topological organization and/or module interaction by employing resting-state functional magnetic resonance imaging (rs-fMRI) and modularity network analysis in 24 patients with ESRD and 20 age- and gender-matched healthy control (HC) subjects. Stroop task was used to evaluate the performance of cognitive control in all subjects. At the global level, ESRD patients exhibited significantly decreased global and local efficiency which were mainly related to abnormal functional connectivity of the amygdala and inferior frontal gyrus (IFG). Stepwise regression analysis was applied to estimate the relationships between network efficiency and blood biochemistry level (urea, creatine, phosphate, Ca2+, hematocrit, cystatin, hemoglobin levels, parathyroid hormone, K+ and Na+), and only the hematocrit level was significantly associated with global efficiency in patients with ESRD. At the modular level, we discovered an aberrant brain interaction between the amygdala- and IFG-related circuits in the ESRD group, and the regional efficiency of the amygdala was observably relative to the performance of cognitive control in patients with ESRD. Our results suggested that ESRD exhibited aberrant brain functional topological organization and module-level interaction between the affective and cognitive control circuits, providing crucial insights into the pathophysiological mechanism of ESRD patients.

Clb6-Cdc28 Promotes Ribonucleotide Reductase Subcellular Redistribution during S Phase.

A tightly controlled cellular deoxyribonucleotide (deoxynucleoside triphosphate [dNTP]) pool is critical for maintenance of genome integrity. One mode of dNTP pool regulation is through subcellular localization of ribonucleotide reductase (RNR), the enzyme that catalyzes the rate-limiting step of dNTP biosynthesis. In Saccharomyces cerevisiae, the RNR small subunit, Rnr2-Rnr4, is localized to the nucleus, whereas the large subunit, Rnr1, is cytoplasmic. As cells enter S phase or encounter DNA damage, Rnr2-Rnr4 relocalizes to the cytoplasm to form an active holoenzyme complex with Rnr1. Although the DNA damage-induced relocalization requires the checkpoint kinases Mec1-Rad53-Dun1, the S-phase-specific redistribution does not. Here, we report that the S-phase cyclin-cyclin-dependent kinase (CDK) complex Clb6-Cdc28 controls Rnr2-Rnr4 relocalization in S phase. Rnr2 contains a consensus CDK site and exhibits Clb6-dependent phosphorylation in S phase. Deletion of CLB6 or removal of the CDK site results in an increased association of Rnr2 with its nuclear anchor Wtm1, nuclear retention of Rnr2-Rnr4, and an enhanced sensitivity to the RNR inhibitor hydroxyurea. Thus, we propose that Rnr2-Rnr4 redistribution in S phase is triggered by Clb6-Cdc28-mediated phosphorylation of Rnr2, which disrupts the Rnr2-Wtm1 interaction and promotes the release of Rnr2-Rnr4 from the nucleus.

The Role of CtBP1 in Oncogenic Processes and Its Potential as a Therapeutic Target.

Transcriptional corepressor proteins have emerged as an important facet of cancer etiology. These corepressor proteins are often altered by loss- or gain-of-function mutations, leading to transcriptional imbalance. Thus, research directed at expanding our current understanding of transcriptional corepressors could impact the future development of new cancer diagnostics, prognostics, and therapies. In this review, our current understanding of the CtBP corepressors, and their role in both development and disease, is discussed in detail. Importantly, the role of CtBP1 overexpression in adult tissues in promoting the progression of multiple cancer types through their ability to modulate the transcription of developmental genes ectopically is explored. CtBP1 overexpression is known to be protumorigenic and affects the regulation of gene networks associated with "cancer hallmarks" and malignant behavior, including increased cell survival, proliferation, migration, invasion, and the epithelial-mesenchymal transition. As a transcriptional regulator of broad developmental processes capable of promoting malignant growth in adult tissues, therapeutically targeting the CtBP1 corepressor has the potential to be an effective method for the treatment of diverse tumor types. Although efforts to develop CtBP1 inhibitors are still in the early stages, the current progress and the future perspectives of therapeutically targeting this transcriptional corepressor are also discussed. Mol Cancer Ther; 16(6); 981-90. ©2017 AACR.

The diferric-tyrosyl radical cluster of ribonucleotide reductase and cytosolic iron-sulfur clusters have distinct and similar biogenesis requirements.

How each metalloprotein assembles the correct metal at the proper binding site presents challenges to the cell. The di-iron enzyme ribonucleotide reductase (RNR) uses a diferric-tyrosyl radical (FeIII2-Y•) cofactor to initiate nucleotide reduction. Assembly of this cofactor requires O2, FeII, and a reducing equivalent. Recent studies show that RNR cofactor biosynthesis shares the same source of iron, in the form of [2Fe-2S]-GSH2 from the monothiol glutaredoxin Grx3/4, and the same electron source, in the form of the Dre2-Tah18 electron transfer chain, with the cytosolic iron-sulfur protein assembly (CIA) machinery required for maturation of [4Fe-4S] clusters in cytosolic and nuclear proteins. Here, we further investigated the interplay between the formation of the FeIII2-Y• cofactor in RNR and the cellular iron-sulfur (Fe-S) protein biogenesis pathways by examining both the iron loading into the RNR ß subunit and the RNR catalytic activity in yeast mutants depleted of individual components of the mitochondrial iron-sulfur cluster assembly (ISC) and the CIA machineries. We found that both iron loading and cofactor assembly in RNR are dependent on the ISC machinery. We also found that Dre2 is required for RNR cofactor formation but appears to be dispensable for iron loading. None of the CIA components downstream of Dre2 was required for RNR cofactor formation. Thus, the pathways for RNR and Fe-S cluster biogenesis bifurcate after the Dre2-Tah18 step. We conclude that RNR cofactor biogenesis requires the ISC machinery to mature the Grx3/4 and Dre2 Fe-S proteins, which then function in iron and electron delivery to RNR, respectively.

Countdown timers, video surveillance and drivers' stop/go behavior: Winter versus summer.

This paper presents an analysis of drivers' stopping and traversing behaviors during inter-green periods. Eight intersections were observed in Changchun, China both with and without countdown timers and/or video surveillance during summer and winter. The impacts of the devices on the drivers' behavior were examined and compared between the two seasons from a safety perspective. During winter abrupt braking on icy road can be very dangerous. The analysis was performed by dividing the approach leg into 10m intervals up to 90m upstream from the stop line. Three impacts are studied, including the profile of approaching speeds, the stop/go decision, and the maximum acceleration and deceleration. The findings revealed that installing both a countdown timer and CCTV in summer, or either of the devices in winter can increase drivers' stopping tendency and hence reduce red-light violations. Especially on an icy road during winter, a countdown timer can help smooth decelerations, which tend to begin earlier than at the intersections without the device, reducing the incidence of sudden braking.

Yeast Dun1 Kinase Regulates Ribonucleotide Reductase Small Subunit Localization in Response to Iron Deficiency.

Ribonucleotide reductase (RNR) is an essential iron-dependent enzyme that catalyzes deoxyribonucleotide synthesis in eukaryotes. Living organisms have developed multiple strategies to tightly modulate RNR function to avoid inadequate or unbalanced deoxyribonucleotide pools that cause DNA damage and genome instability. Yeast cells activate RNR in response to genotoxic stress and iron deficiency by facilitating redistribution of its small heterodimeric subunit Rnr2-Rnr4 from the nucleus to the cytoplasm, where it forms an active holoenzyme with large Rnr1 subunit. Dif1 protein inhibits RNR by promoting nuclear import of Rnr2-Rnr4. Upon DNA damage, Dif1 phosphorylation by the Dun1 checkpoint kinase and its subsequent degradation enhances RNR function. In this report, we demonstrate that Dun1 kinase triggers Rnr2-Rnr4 redistribution to the cytoplasm in response to iron deficiency. We show that Rnr2-Rnr4 relocalization by low iron requires Dun1 kinase activity and phosphorylation site Thr-380 in the Dun1 activation loop, but not the Dun1 forkhead-associated domain. By using different Dif1 mutant proteins, we uncover that Dun1 phosphorylates Dif1 Ser-104 and Thr-105 residues upon iron scarcity. We observe that the Dif1 phosphorylation pattern differs depending on the stimuli, which suggests different Dun1 activating pathways. Importantly, the Dif1-S104A/T105A mutant exhibits defects in nucleus-to-cytoplasm redistribution of Rnr2-Rnr4 by iron limitation. Taken together, these results reveal that, in response to iron starvation, Dun1 kinase phosphorylates Dif1 to stimulate Rnr2-Rnr4 relocalization to the cytoplasm and promote RNR function.

The late-annotated small ORF LSO1 is a target gene of the iron regulon of Saccharomyces cerevisiae.

We have identified a new downstream target gene of the Aft1/2-regulated iron regulon in budding yeast Saccharomyces cerevisiae, the late-annotated small open reading frame LSO1. LSO1 transcript is among the most highly induced from a transcriptome analysis of a fet3-1 mutant grown in the presence of the iron chelator bathophenanthrolinedisulfonic acid. LSO1 has a paralog, LSO2, which is constitutively expressed and not affected by iron availability. In contrast, we find that the LSO1 promoter region contains three consensus binding sites for the Aft1/2 transcription factors and that an LSO1-lacZ reporter is highly induced under low-iron conditions in a Aft1-dependent manner. The expression patterns of the Lso1 and Lso2 proteins mirror those of their mRNAs. Both proteins are localized to the nucleus and cytoplasm, but become more cytoplasmic upon iron deprivation consistent with a role in iron transport. LSO1 and LSO2 appear to play overlapping roles in the cellular response to iron starvation since single lso1 and lso2 mutants are sensitive to iron deprivation and this sensitivity is exacerbated when both genes are deleted.

Children with congenital cystic adenomatoid malformation of the lung CT diagnosis.

In this study, we aim to investigate the imaging appearances of congenital cystic adenomatoid malformation (CCAM) of the lung, and to enhance the understanding of this disease. A total of 11 cases with CCAM of the lung were confirmed by surgery and pathology. Preoperative chest computed tomography (CT) scan was performed in all patients, and high resolution CT scan was performed in lesion areas of 7 cases. Our results showed that there were 3 cases involving left and right lung, 5 cases involving right lung and 3 cases involving left lung. CT scan showed 6 cases with single or multiple air-filled cavities (> 2 cm in diameter) and 5 cases with multiple honeycomb-like cysts (< 1 cm in diameter). The cysts of CCAM contained air in 9 cases and a small amount of liquid in 2 cases. The complications of CCAM included different degree of emphysema in 7 patients, mediastinal hernia in 5 cases and congenital pulmonary sequestration in 1 case. All lesions have certain space-occupying effect. In conclusion, CT manifestation of CCAM of lung has certain characteristics and can provide reliable information for diagnosis of the disease.

Essential role of the iron-sulfur cluster binding domain of the primase regulatory subunit Pri2 in DNA replication initiation.

DNA primase catalyzes de novo synthesis of a short RNA primer that is further extended by replicative DNA polymerases during initiation of DNA replication. The eukaryotic primase is a heterodimeric enzyme comprising a catalytic subunit Pri1 and a regulatory subunit Pri2. Pri2 is responsible for facilitating optimal RNA primer synthesis by Pri1 and mediating interaction between Pri1 and DNA polymerase α for transition from RNA synthesis to DNA elongation. All eukaryotic Pri2 proteins contain a conserved C-terminal iron-sulfur (Fe-S) cluster-binding domain that is critical for primase catalytic activity in vitro. Here we show that mutations at conserved cysteine ligands for the Pri2 Fe-S cluster markedly decrease the protein stability, thereby causing S phase arrest at the restrictive temperature. Furthermore, Pri2 cysteine mutants are defective in loading of the entire DNA pol α-primase complex onto early replication origins resulting in defective initiation. Importantly, assembly of the Fe-S cluster in Pri2 is impaired not only by mutations at the conserved cysteine ligands but also by increased oxidative stress in the sod1Δ mutant lacking the Cu/Zn superoxide dismutase. Together these findings highlight the critical role of Pri2's Fe-S cluster domain in replication initiation in vivo and suggest a molecular basis for how DNA replication can be influenced by changes in cellular redox state.

The expression profile and promoter analysis of ultraspiracle gene in the silkworm Bombyx mori.

The nuclear receptor, ultraspiracle protein (USP), is a transcription factor and an essential component of a heterodimeric receptor complex with ecdysone receptor. However, the mechanisms underlying the transcriptional regulation of USP in silkworm are unknown. In this study, using dual-spike-in qPCR method, we examined the expression of Bombyx ultraspiracle gene (BmUSP) in various tissues of silkworm as well as expression changes after stimulation with ecdysone. The results showed that the expression levels of BmUSP gene varied in different tissues and were increased 2 h after exposure to ecdysone. To identify the molecular mechanism underlying the regulation of USP gene expression in silkworm Bombyx mori, promoter truncation analyses were performed using the luciferase reporter assay and Bac-to-Bac expression system in several tissues of B. mori. BmUSP gene promoter with 5' end serial deletions showed different levels of activity in various tissues, higher in fat body and Malpighian tubule. Deletion of the region from -485 to -445 and -307 to -281 upstream of BmUSP gene abolished and increased its promoter activity, respectively. This region contains AP-1, Dfd transcription factor binding sites. These results indicate that BmUSP are expressed at different levels in different tissues of the silkworm, but all are subjected to the regulation by ecdysone. This study would provide an important foundation for investigating the mechanism underlying the transcriptional regulation of BmUSP in the silkworm.