A nanopore-based cucumber genome assembly reveals structural variations at two QTLs controlling hypocotyl elongation.

The Xishuangbanna (XIS) cucumber (Cucumis sativus var. xishuangbannanesis) is a semiwild variety that has many distinct agronomic traits. Here, long reads generated by Nanopore sequencing technology helped assembling a high-quality genome (contig N50 = 8.7 Mb) of landrace XIS49. A total of 10,036 structural/sequence variations (SVs) were identified when comparing with Chinese Long (CL), and known SVs controlling spines, tubercles, and carpel number were confirmed in XIS49 genome. Two QTLs of hypocotyl elongation under low light, SH3.1 and SH6.1, were fine-mapped using introgression lines (donor parent, XIS49; recurrent parent, CL). SH3.1 encodes a red-light receptor Phytochrome B (PhyB, CsaV3_3G015190). A ∼4 kb region with large deletion and highly divergent regions (HDRs) were identified in the promoter of the PhyB gene in XIS49. Loss of function of this PhyB caused a super-long hypocotyl phenotype. SH6.1 encodes a CCCH-type zinc finger protein FRIGIDA-ESSENTIAL LIKE (FEL, CsaV3_6G050300). FEL negatively regulated hypocotyl elongation but it was transcriptionally suppressed by long terminal repeats retrotransposon insertion in CL cucumber. Mechanistically, FEL physically binds to the promoter of CONSTITUTIVE PHOTOMORPHOGENIC 1a (COP1a), regulating the expression of COP1a and the downstream hypocotyl elongation. These above results demonstrate the genetic mechanism of cucumber hypocotyl elongation under low light.

Transcriptome analysis of frontal fibrosis alopecia revealed involvement of immune cells and ferroptosis.

BACKGROUND: Frontal fibrosis alopecia (FFA) is a primary cicatricial alopecia and has received increasing attention in recent years. However, the pathogenesis of FFA has not been fully elucidated. METHODS AND RESULTS: Herein, we collected the transcriptome data of scalp lesions of seven patients with FFA and seven healthy controls. The differential expression analysis and weighted gene co-expression network analysis were conducted and we identified 458 differentially expressed genes (DEGs) in two key modules. Later, we performed functional enrichment analysis and functional modules identification, revealing the participation of immune response and fatty acid metabolism. Based on the results, we processed further studies. On the one hand, we analyzed the infiltrating immune cells of FFA through CIBERSORT algorithm, indicating the activation of M1 macrophage and CD8+ T cell. On the other hand, considering lipid metabolism of FFA and oxidative stress of hair follicle cells in alopecia, we explored the potential ferroptosis of FFA. By intersection of DEGs and ferroptosis-related genes from FerrDb database, 19 genes were identified and their expression was validated in an external dataset containing 36 FFA cases and 12 controls. Then, we used LASSO algorithms to construct a four-gene diagnostic model, which achieved an AUC of 0.924 in validation dataset. Additionally, the immune cells were found to be related to ferroptosis in FFA. CONCLUSION: Taken together, this study contributed to reveal the molecular mechanisms of FFA and is expected to inspire future research on treatment.

Exploration of Single and Co-Toxic Effects of Polypropylene Micro-Plastics and Cadmium on Rice (Oryza sativa L.).

The widespread application of micro-plastics (MP) and their release in the open environment has become a matter of worldwide concern. When interacting with contaminants such as heavy metals in the soil ecosystem, MPs can result in detrimental effects on the soil environment and plant growth and development. However, information based on the interaction between MPs and heavy metals and their effects on terrestrial plants is still limited. Keeping this in mind, the present study was conducted to explore the single and combined toxicity of polypropylene (PP) MPs (13 and 6.5 µm) and cadmium (Cd) on germination indices; root and stem growth; fresh and dry weight; and anti-oxidative enzyme activities of rice (Oryza sativa L.) seedlings. Our results indicated that a single application of PP MP and Cd on rice seedlings inhibited most of the germination indicators, while their co-occurrence (PP + Cd) showed a reduction in the overall toxicity to some extent. A single application of both the contaminants significantly inhibited root length, stem length, fresh weight and the activities of catalase (CAT), peroxidase (POD) and superoxide dismutase (SOD) enzymes in rice seedling, while no significant effect on dry weight was observed. The combined toxicity of both PP and Cd revealed that 13 µm PP + Cd had an antagonistic effect on the growth of rice seedlings, while 6.5 µm PP + Cd showed a synergistic effect. The present study revealed that smaller PP MP particles (6.5 µm) prominently affected plant growth more as compared to larger particles (13 µm). Our work reported the combined effect of PP MP and Cd on the germination and growth of rice for the first time. This study can provide the basis for future research on the combined effects of different types and sizes of MPs and heavy metals on the terrestrial ecosystem.

PDP1 Promotes Cell Malignant Behavior and Is Associated with Worse Clinical Features in Ovarian Cancer Patients: Evidence from Bioinformatics and In Vitro Level.

PDP1 has been reported in multiple diseases. However, it has not been fully explored in ovarian cancer (OC). The public data was downloaded from The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) databases. Differentially expressed gene analysis was conducted out using the limma package. Prognosis analysis was performed using the survival package. Gene Set Enrichment Analysis (GSEA) was performed using the fgsea package. Immune infiltration analysis was performed based on the CIBERSORT algorithm. CCK8 assay was used to evaluate the cell proliferation ability of cancer cells. Transwell assay was used for the invasion and migration ability. Our result showed that PDP1 was overexpressed in OC tissue in RNA and protein level based on multiple databases (TCGA, GSE18520, GSE27651, and GSE54388). At the same time, we found PDP1 was correlated with poor prognosis and worse clinical parameters. In vitro experiment showed that PDP1 could significantly promote proliferation, invasion, and migration ability of OC cells. GSEA analysis showed that in the OC patients with high PDP1 expression, the pathway of IL6/JAK/STAT3 signaling, interferon-alpha response, apoptosis, adipogenesis, KRAS signaling, and IL2/STAT5 signaling was activated, which might be responsible for its oncogenic effect in OC. Immune infiltration analysis indicated that PDP1 was positively correlated with activated myeloid dendritic cells, resting CD4 memory T cells, neutrophil, and M1 and M2 macrophages, yet negatively correlated with M0 macrophages, plasma B cells, γδT cells, and activated CD4 memory T cells. Drug sensitivity analysis showed a negative correlation between PDP1 expression and the IC50 of bleomycin and gemcitabine, yet a positive correlation of cisplatin, indicating that the OC patients with high PDP1 expression might be more sensitive to bleomycin and gemcitabine and more resistant to cisplatin. PDP1 could facilitate OC progression and is associated with patient prognosis and chemosensitivity, making it an underlying biomarker of OC.

Application Potential of CTHRC1 as a Diagnostic and Prognostic Indicator for Colon Adenocarcinoma.

Colon adenocarcinoma (COAD), ranking third in incidence and second in mortality, is one of the most common cancer types in the world. The initial stages of COAD usually show no obvious clinical symptoms; moreover, effective screening or diagnostic indicators with high sensitivity and specificity are lacking, which often leads to missed treatment opportunities. Collagen triple helix repeat containing 1 (CTHRC1) is a glycosylated protein secreted during tissue repair, which reduces collagen matrix deposition and promotes cell migration. Under physiological conditions, the expression of CTHRC1 is conducive to wound healing; however, the pathological overexpression of CTHRC1 promotes tumour growth and proliferation. In this study, we evaluated the application potential of CTHRC1 as an early diagnosis and prognostic survival monitoring biomarker for COAD in addition to unravelling its molecular mechanism in the development of COAD and exploring new therapeutic targets. Therefore, various tumour databases were used to investigate the expression of CTHRC1 in COAD at the mRNA and protein levels. CTHRC1 expression was found to be significantly increased in COAD, regardless of clinical cancer stage, age, sex or race. Moreover, CTHRC1 expression was significantly correlated with poor prognosis and positively correlated with CD8+ T cell, CD4+ T cell, neutrophil, macrophage and dendritic cell infiltration. The relevant function pathways and neighbouring proteins to CTHRC1 in COAD were identified as ROR2, VAPA, LY6E and several collagen family proteins. Therefore, this study suggests that CTHRC1 is a potential diagnostic and prognostic biomarker for patients with COAD.

Local Treatment of Hydrogen-Rich Saline Promotes Wound Healing In Vivo by Inhibiting Oxidative Stress via Nrf-2/HO-1 Pathway.

Wound healing is a complex dynamic process involving a large number of biological events. Excessive oxidative stress is a key factor delaying wound healing. Hydrogen is an antioxidant, anti-inflammatory, and antiapoptotic medical gas with safety, effectiveness, and penetrability. However, the effects of local treatment of hydrogen on wound healing and its potential mechanisms remain unclear. In this study, Kunming (KM) mice were used to set up a wound model. All the mice were randomly divided into the control, the local treatment with saline group, the local treatment with the hydrogen-rich saline group, and the intraperitoneal injection of the hydrogen-rich saline group. To evaluate the impact of hydrogen-rich saline on wound healing, we assessed the wound healing rate, wound closure time, histomorphology, oxidative stress indicators, inflammatory cytokines, the apoptosis index, and the expression of the nuclear factor-erythroid-related factor 2(Nrf-2). Furthermore, the immortalized nontumorigenic human epidermal (HaCaT) cells were chosen to investigate the therapeutic effects of hydrogen-rich medium on oxidative stress and its underlying mechanisms. The results showed that local treatment of hydrogen-rich saline shortened wound closure time and reduced the level of proinflammatory cytokines and lipid peroxidation. Meanwhile, it decreased the cell apoptosis index and increased the Nrf-2 expression. Besides, hydrogen-rich medium relieved the oxidative stress via the activation of the Nrf-2/heme oxygenase-1 (HO-1) pathway. In conclusion, local treatment of hydrogen-rich saline exhibits the healing-promoting function through antioxidant, anti-inflammatory, and antiapoptotic effects. Hydrogen relieves the oxidative stress in the wound microenvironment via Nrf-2/HO-1 signaling pathway. This study may offer a new strategy to promote wound healing and a new perspective to illustrate the mechanism of wound healing.

c-Casitas b-Lineage Lymphoma Downregulation Improves the Ability of Long-term Cultured Mesenchymal Stem Cells for Promoting Angiogenesis and Diabetic Wound Healing.

The chronic wound induced by diabetes has poor efficacy and could lead to amputation. The repair function of mesenchymal stem cells (MSCs) impaired after long-term culture in vitro. Studies have shown that the proto-oncogene c-Casitas b-lineage lymphoma (c-Cbl) can regulate receptor- and non-receptor tyrosine kinase, which was also involved in the angiogenesis process. This study aimed to explore the regulative effect of c-Cbl on the proangiogenic functions of long-term cultured MSCs and evaluate its pro-healing effect on diabetic wounds. In this study, the c-Cbl level was downregulated by locked nucleic acid-modified antisense oligonucleotide gapmers (LNA Gapmers). We detected the effect of c-Cbl downregulation on long-term cultured MSCs in terms of phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt) signal, cellular proliferation, senescence, migration, and angiogenic factors paracrine activity in vitro. In vivo, we observed the pro-healing effect of long-term cultured MSCs, with or without c-Cbl downregulation, on the diabetic wound. We found that the phosphorylation level of c-Cbl increased and that of Akt decreased in passage 10 (P10) MSCs compared with passage 3 (P3) MSCs (P < 0.05). Additionally, the proliferation, paracrine, and migration capacity of P10 MSCs decreased significantly, accompanied by the increase of cellular senescence (P < 0.05). However, these functions, including PI3K/Akt activity of P10 MSCs, have been improved by c-Cbl downregulation (P < 0.05). Compared with P10 MSCs treatment, treatment with c-Cbl downregulated P10 MSCs accelerated diabetic wound healing, as defined by a more rapid wound closure (P < 0.05), more neovascularization (P < 0.05), and higher scores of wound histological assessment (P < 0.05) in a diabetic rat model. Our findings suggested that c-Cbl downregulation could attenuate the impairment of proangiogenic functions in MSCs induced by long-term culture in vitro and improve the effect of long-term cultured MSCs in promoting diabetic wound healing.

Topical effect of benzalkonium bromide on wound healing and potential cellular and molecular mechanisms.

Benzalkonium bromide (BB) has been widely used as a skin antiseptic for wound management. However, BB had proinflammation and reactive oxygen species (ROS) induction effect, making its role in wound healing complex and unclear. A rat full-thickness skin defect wound model was established. The effects of BB, povidone iodine (PVP-I), chlorhexidine gluconate (CHG), and normal saline (NS) on wound healing and infection control were then evaluated based on wound healing rate (WHr) and bacterial killing. The wound tissues were sectioned for histopathological evaluation and nuclear factor E2 related factor 2 (Nrf2) expression determination. The ROS production, Nrf2 activation, and heme oxygenase 1 (HO-1) expression of the HaCat cells and the cytotoxicity treated with BB were further explored. Compared with NS, PVP-I, and CHG, BB showed the best wound infection control efficiency while delayed wound healing with the WHr of 91.42 ± 5.12% at day 20. The wound tissue of the BB group showed many inflammatory cells but few granulation tissue and capillaries and no obvious collagen deposition, resulting in the lowest histopathological scores of 4.17 ± 0.75 for BB group. BB showed higher cytotoxicity on HaCat cells with the lowest IC25, IC50, and IC75 of 1.90, 4.16, and 9.09 g/mL compared with PVP-I and CHG. TUNEL staining evaluated the cytotoxicity of BB on wound tissue, which indicates the high apoptosis index BB group (5.05 ± 1.77). Compared with PVP-I and CHG, BB induced much more cell apoptosis. The results of flow cytometry and fluorescence staining showed that PVP-I, CHG, and BB induced ROS production in a concentration-dependent manner and cells treated with BB had the highest ROS production at the same inhibition concentration. The cells and the wound tissues treated with BB showed highest Nrf2 activation and HO-1 expression than PVP-I and CHG. BB was highly efficient in wound infection control while delayed wound healing. The prolonged and strengthened inflammation and the raised ROS production originating from BB administration may contribute to delayed wound healing.

A new species of glass sponge (Hexactinellida: Sceptrulophora: Uncinateridae) from the Weijia Seamount in the northwestern Pacific Ocean.

In this article we report on a hexactinellid sponge new to science, Tretopleura weijicus sp. nov., which was collected from the Weijia Seamount in the northwestern Pacific Ocean at a depth of 1995 m. Its blade-like and branching body form, the primary choanosomal framework consisting of multiaxial longitudinal strands without synapticula, and the presence of sceptrules suggest placement within the family Uncinateridae Reiswig, 2002, which is also supported by molecular phylogenetic evidence from COI and 16S sequences. The absence of swollen dermal spurs or regular pentactins and the presence of two types of discohexasters characterize it as a new species. More specimens should be collected to revise the generic characters.

TiO2 nanoparticles accelerate methanogenesis in mangrove wetlands sediment.

In this study, the response of methane (CH4) production to the addition of titanium dioxide nanoparticles (TiO2 NPs) with three types of short-chain fatty acids (sodium acetate, sodium propionate and sodium butyrate) as carbon sources in mangrove sediment was investigated. The results showed that the maximum CH4 formation rate increased by 45.2%, 32.7% and 48.6% and the maximum cumulative CH4 production increased by 25.2%, 7.7% and 6.3% with the addition of TiO2 NPs in the sodium acetate, sodium propionate and sodium butyrate systems, respectively. The microbial community analysis revealed that the electrogenic bacteria Proteiniclasticum and Pseudomonas, butyrate oxidizing bacteria Syntrophomonas and methanogens Methanobacterium and Methanosarcina were significantly enriched in the presence of TiO2 NPs, indicating that TiO2 NPs can enhance CH4 production by stimulating the growth of different species of methanogens and butyrate oxidizing bacteria. The enlarged distance between microbes, the enhanced conductivity of the sediment and the typical microorganisms for direct interspecies electron transfer (DIET) with the addition of TiO2 NPs suggest that the promoted DIET between distinct microorganisms could be another possible explanation for the improvement in CH4 production. It can be speculated that a weaker effect on methanogenesis increases under the natural concentration of TiO2 NPs compared with the experimental conditions; however, the amounts of TiO2 NPs are increasing enriched in wetland environments. Therefore, the findings of this study increase current knowledge about the effect of nanomaterials on global CH4 emissions and suggest that the discharge of wastewater containing TiO2 NPs from the synthesis and incorporation of TiO2 NPs in customer products needs to be monitored.

A new glass sponge genus (Hexactinellida: Euplectellidae) from abyssal depth of the Yap Trench, northwestern Pacific Ocean.

In this article we report on a hexactinellid sponge new to science, Rhizophyta yapensis gen. et sp. nov., which was collected from the Yap Trench in the northwestern Pacific Ocean at an abyssal depth of 4159-4779 m. Its fungus-like body form with long peduncle and absence of hypodermalia suggest placement within the euplectellid subfamily Bolosominae Tabachnick, 2002, whereas molecular phylogenetic evidence suggests that it is sister to all remaining Euplectellidae Gray, 1867. Its rhizophytous method of attachment to the substrate, hitherto unknown from bolosomine Euplectellidae, a veil of pentactins covering the peduncle, and the presence of toothed discohexasters as the only type of microscleres, clearly characterize it as a new genus. The intraspecific divergence between holotype and paratypes of the new species is examined with both morphological and molecular approaches. This report represents the first record of a hexactinellid sponge from the Yap Trench.

Comparison of lumbar plexus block using the short axis in-plane method at the plane of the transverse process and at the articular process: a randomized controlled trial.

BACKGROUND: Although the safety and effectiveness of the short-axis in-plane method has been confirmed for lumbar plexus block, the operation is difficult and has a high rate of epidural spread at the plane of the articular process. Therefore, we developed a new in-plane technique, called the beach chair method, which displays images from the transverse process. We compared the operative difficulty and incidence of epidural spread of the beach chair method with those of the control method (at the plane of the articular process) in this randomized controlled clinical trial. METHODS: Sixty patients, aged 18 to 75 years, scheduled for unilateral arthroscopic knee surgery were randomized to receive double-guided lumbar plexus block by the beach chair method (n = 30) or the control method (n = 30) with 30 ml 0.5% ropivacaine hydrochloride; all patients received a sciatic nerve block with 10 ml 1% lidocaine hydrochloride and 10 ml 0.5% ropivacaine hydrochloride. RESULTS: The incidence of epidural spread after lumbar plexus block was significantly lower in the beach chair group than that in the control group [1 case (3.3%) vs. 9 (30.0%), P = 0.006]. Moreover, the imaging time (34.2 ± 16.7 s vs. 48.9 ± 16.8 s, P = 0.001), needling time (85.0 ± 45.3 s vs. 131.4 ± 88.2 s, P = 0.013) and number of needle punctures (2.7 ± 1.3 vs. 4.5 ± 2.1, P = 0.000) were significantly lower in the beach chair group than those in the control group; the ultrasound visibility score of the beach chair group was better than that of the control group. There were no significant differences in the remaining indicators. CONCLUSIONS: The beach chair method was easier and was associated with a lower incidence of epidural spread than the control method. Therefore, the beach chair method (at the plane of the transverse process) provides another promising option for lumbar plexus block for the non-obese population. TRIAL REGISTRATION: Chinese Clinical Trial Registry (ChiCTR), Registration number:ChiCTR-INR-15007505, registered on November 06, 2015.

AMD3100 treatment attenuates pulmonary angiogenesis by reducing the c-kit (+) cells and its pro-angiogenic activity in CBDL rat lungs.

Recent studies have shown that pulmonary angiogenesis is an important pathological process in the development of hepatopulmonary syndrome (HPS), and growing evidence has indicated that Stromal cell-derived factor 1/C-X-C chemokine receptor type 4 (SDF-1/CXCR4) axis is involved in pulmonary vascular disease by mediating the accumulation of c-kit+ cells. This study aimed to test the effect of AMD3100, an antagonist of CXCR4, in HPS pulmonary angiogenesis. Common bile duct ligation (CBDL) rats were used as experimental HPS model and were treated with AMD3100 (1.25mg/kg/day, i.p.) or 0.9% saline for 3weeks. The sham rats underwent common bile duct exposure without ligation. The c-kit+ cells accounts and its angiogenic-related functions, prosurvival signals, pulmonary angiogenesis and arterial oxygenation were analysed in these groups. Our results showed that pulmonary SDF-1/CXCR4, Akt, Erk and VEGF/VEGFR2 were significantly activated in CBDL rats, and the numbers of circulating and pulmonary c-kit+ cells were increased in CBDL rats compared with control rats. Additionally, the angiogenic-related functions of c-kit+ cells and pulmonary microvessel counts were also elevated in CBDL rats. CXCR4 inhibition reduced pulmonary c-kit+ cells and microvessel counts and improved arterial oxygenation within 3weeks in CBDL rats. The pulmonary prosurvival signals and pro-angiogenic activity of c-kit+ cells were also down-regulated in AMD3100-treated rats. In conclusion, AMD3100 treatment attenuated pulmonary angiogenesis in CBDL rats and prevented the development of HPS via reductions in pulmonary c-kit+ cells and inhibition of the prosurvival signals. Our study provides new insights in HPS treatment.

The angiogenic related functions of bone marrow mesenchymal stem cells are promoted by CBDL rat serum via the Akt/Nrf2 pathway.

Hepatopulmonary syndrome (HPS) is a complication of severe liver disease. It is characterized by an arterial oxygenation defect. Recent studies have demonstrated that pulmonary angiogenesis contributes to the abnormal gas exchange found in HPS. Additionally, mesenchymal stem cells (MSCs) are considered the stable source of VEGF-producing cells and have the potential to differentiate into multiple cell types. However, it has not been determined whether bone marrow mesenchymal stem cells (BM-MSCs) are mobilized and involved in the pulmonary angiogenesis in HPS. In this study, a CFU-F assay showed that the number of peripheral blood MSCs was increased in common bile duct ligation (CBDL) rats; however, there was no significant difference found in the number of BM-MSCs. In vitro, CBDL rat serum induced the overexpression of CXCR4 and PCNA in BM-MSCs. Consistently, the directional migration as well as the proliferation ability of BM-MSCs were enhanced by CBDL rat serum, as determined by a transwell migration and MTT assays. Moreover, the secretion of VEGF by BM-MSCs increased after treatment with CBDL rat serum. We also found that the expression of phospho-Akt, phospho-ERK, and Nrf2 in BM-MSCs was significantly up-regulated by CBDL rat serum in a time dependent manner, and the blockage of the Akt/Nrf2 signalling pathway with an Akt Inhibitor or Nrf2 siRNA, instead of an ERK inhibitor, attenuated the migration, proliferation and paracrine capacity of BM-MSCs. In conclusion, these findings indicated that the number of MSCs increased in the peripheral blood of CBDL rats, and the Akt/Nrf2 pathway plays a vital role in promoting the angiogenic related functions of BM-MSCs, which could be a potent contributor to pulmonary angiogenesis in HPS.

Study on the cumulative impact of reclamation activities on ecosystem health in coastal waters.

The purpose of this study is to develop feasible tools to investigate the cumulative impact of reclamations on coastal ecosystem health, so that the strategies of ecosystem-based management can be applied in the coastal zone. An indicator system and model were proposed to assess the cumulative impact synthetically. Two coastal water bodies, namely Laizhou Bay (LZB) and Tianjin coastal waters (TCW), in the Bohai Sea of China were studied and compared, each in a different phase of reclamations. Case studies showed that the indicator scores of coastal ecosystem health in LZB and TCW were 0.75 and 0.68 out of 1.0, respectively. It can be concluded that coastal reclamations have a historically cumulative effect on benthic environment, whose degree is larger than that on aquatic environment. The ecosystem-based management of coastal reclamations should emphasize the spatially and industrially intensive layout.

Program specificity for Ptf1a in pancreas versus neural tube development correlates with distinct collaborating cofactors and chromatin accessibility.

The lineage-specific basic helix-loop-helix transcription factor Ptf1a is a critical driver for development of both the pancreas and nervous system. How one transcription factor controls diverse programs of gene expression is a fundamental question in developmental biology. To uncover molecular strategies for the program-specific functions of Ptf1a, we identified bound genomic regions in vivo during development of both tissues. Most regions bound by Ptf1a are specific to each tissue, lie near genes needed for proper formation of each tissue, and coincide with regions of open chromatin. The specificity of Ptf1a binding is encoded in the DNA surrounding the Ptf1a-bound sites, because these regions are sufficient to direct tissue-restricted reporter expression in transgenic mice. Fox and Sox factors were identified as potential lineage-specific modifiers of Ptf1a binding, since binding motifs for these factors are enriched in Ptf1a-bound regions in pancreas and neural tube, respectively. Of the Fox factors expressed during pancreatic development, Foxa2 plays a major role. Indeed, Ptf1a and Foxa2 colocalize in embryonic pancreatic chromatin and can act synergistically in cell transfection assays. Together, these findings indicate that lineage-specific chromatin landscapes likely constrain the DNA binding of Ptf1a, and they identify Fox and Sox gene families as part of this process.

A tripartite clustering analysis on microRNA, gene and disease model.

Alteration of gene expression in response to regulatory molecules or mutations could lead to different diseases. MicroRNAs (miRNAs) have been discovered to be involved in regulation of gene expression and a wide variety of diseases. In a tripartite biological network of human miRNAs, their predicted target genes and the diseases caused by altered expressions of these genes, valuable knowledge about the pathogenicity of miRNAs, involved genes and related disease classes can be revealed by co-clustering miRNAs, target genes and diseases simultaneously. Tripartite co-clustering can lead to more informative results than traditional co-clustering with only two kinds of members and pass the hidden relational information along the relation chain by considering multi-type members. Here we report a spectral co-clustering algorithm for k-partite graph to find clusters with heterogeneous members. We use the method to explore the potential relationships among miRNAs, genes and diseases. The clusters obtained from the algorithm have significantly higher density than randomly selected clusters, which means members in the same cluster are more likely to have common connections. Results also show that miRNAs in the same family based on the hairpin sequences tend to belong to the same cluster. We also validate the clustering results by checking the correlation of enriched gene functions and disease classes in the same cluster. Finally, widely studied miR-17-92 and its paralogs are analyzed as a case study to reveal that genes and diseases co-clustered with the miRNAs are in accordance with current research findings.

Activation of postnatal neural stem cells requires nuclear receptor TLX.

Neural stem cells (NSCs) continually produce new neurons in postnatal brains. However, the majority of these cells stay in a nondividing, inactive state. The molecular mechanism that is required for these cells to enter proliferation still remains largely unknown. Here, we show that nuclear receptor TLX (NR2E1) controls the activation status of postnatal NSCs in mice. Lineage tracing indicates that TLX-expressing cells give rise to both activated and inactive postnatal NSCs. Surprisingly, loss of TLX function does not result in spontaneous glial differentiation, but rather leads to a precipitous age-dependent increase of inactive cells with marker expression and radial morphology for NSCs. These inactive cells are mispositioned throughout the granular cell layer of the dentate gyrus during development and can proliferate again after reintroduction of ectopic TLX. RNA-seq analysis of sorted NSCs revealed a TLX-dependent global expression signature, which includes the p53 signaling pathway. TLX regulates p21 expression in a p53-dependent manner, and acute removal of p53 can rescue the proliferation defect of TLX-null NSCs in culture. Together, these findings suggest that TLX acts as an essential regulator that ensures the proliferative ability of postnatal NSCs by controlling their activation through genetic interaction with p53 and other signaling pathways.

Replacement of Rbpj with Rbpjl in the PTF1 complex controls the final maturation of pancreatic acinar cells.

BACKGROUND & AIMS: The mature pancreatic acinar cell is dedicated to the production of very large amounts of digestive enzymes. The early stages of pancreatic development require the Rbpj form of the trimeric Pancreas Transcription Factor 1 complex (PTF1-J). As acinar development commences, Rbpjl gradually replaces Rbpj; in the mature pancreas, PTF1 contains Rbpjl (PTF1-L). We investigated whether PTF1-L controls the expression of genes that complete the final stage of acinar differentiation. METHODS: We analyzed acinar development and transcription in mice with disrupted Rbpjl (Rbpjl(ko/ko) mice). We performed comprehensive analyses of the messenger RNA population and PTF1 target genes in pancreatic acinar cells from these and wild-type mice. RESULTS: In Rbpjl(ko/ko) mice, acinar differentiation was incomplete and characterized by decreased expression (as much as 99%) of genes that encode digestive enzymes or proteins of regulated exocytosis and mitochondrial metabolism. Whereas PTF1-L bound regulatory sites of genes in normal adult pancreatic cells, the embryonic form (PTF1-J) persisted in the absence of Rbpjl and replaced PTF1-L; the extent of replacement determined gene expression levels. Loss of PTF1-L reduced expression (>2-fold) of only about 50 genes, 90% of which were direct targets of PTF1-L. The magnitude of the effects on individual digestive enzyme genes correlated with the developmental timing of gene activation. Absence of Rbpjl increased pancreatic expression of liver-restricted messenger RNA. CONCLUSIONS: Replacement of Rbpj by Rbpjl in the PTF1 complex drives acinar differentiation by maximizing secretory protein synthesis, stimulating mitochondrial metabolism and cytoplasmic creatine-phosphate energy stores, completing the packaging and secretory apparatus, and maintaining acinar-cell homeostasis.

Massive Ca-induced membrane fusion and phospholipid changes triggered by reverse Na/Ca exchange in BHK fibroblasts.

Baby hamster kidney (BHK) fibroblasts increase their cell capacitance by 25-100% within 5 s upon activating maximal Ca influx via constitutively expressed cardiac Na/Ca exchangers (NCX1). Free Ca, measured with fluo-5N, transiently exceeds 0.2 mM with total Ca influx amounting to approximately 5 mmol/liter cell volume. Capacitance responses are half-maximal when NCX1 promotes a free cytoplasmic Ca of 0.12 mM (Hill coefficient approximately 2). Capacitance can return to baseline in 1-3 min, and responses can be repeated several times. The membrane tracer, FM 4-64, is taken up during recovery and can be released at a subsequent Ca influx episode. Given recent interest in signaling lipids in membrane fusion, we used green fluorescent protein (GFP) fusions with phosphatidylinositol 4,5-bisphosphate (PI(4,5)P(2)) and diacylglycerol (DAG) binding domains to analyze phospholipid changes in relation to these responses. PI(4,5)P(2) is rapidly cleaved upon activating Ca influx and recovers within 2 min. However, PI(4,5)P(2) depletion by activation of overexpressed hM1 muscarinic receptors causes only little membrane fusion, and subsequent fusion in response to Ca influx remains massive. Two results suggest that DAG may be generated from sources other than PI(4,5)P in these protocols. First, acylglycerols are generated in response to elevated Ca, even when PI(4,5)P(2) is metabolically depleted. Second, DAG-binding C1A-GFP domains, which are brought to the cell surface by exogenous ligands, translocate rapidly back to the cytoplasm in response to Ca influx. Nevertheless, inhibitors of PLCs and cPLA2, PI(4,5)P(2)-binding peptides, and PLD modification by butanol do not block membrane fusion. The cationic agents, FM 4-64 and heptalysine, bind profusely to the extracellular cell surface during membrane fusion. While this binding might reflect phosphatidylserine (PS) "scrambling" between monolayers, it is unaffected by a PS-binding protein, lactadherin, and by polylysine from the cytoplasmic side. Furthermore, the PS indicator, annexin-V, binds only slowly after fusion. Therefore, we suggest that the luminal surfaces of membrane vesicles that fuse to the plasmalemma may be rather anionic. In summary, our results provide no support for any regulatory or modulatory role of phospholipids in Ca-induced membrane fusion in fibroblasts.