Identification of Genes Directly Involved in Shell Formation and Their Functions in Pearl Oyster, Pinctada fucata

Mollusk shell formation is a fascinating aspect of biomineralization research. Shell matrix proteins play crucial roles in the control of calcium carbonate crystallization during shell formation in the pearl oyster, Pinctada fucata. Characterization of biomineralization-related genes during larval development could enhance our understanding of shell formation. Genes involved in shell biomineralization were isolated by constructing three suppression subtractive hybridization (SSH) libraries that represented genes expressed at key points during larval shell formation. A total of 2,923 ESTs from these libraries were sequenced and gave 990 unigenes. Unigenes coding for secreted proteins and proteins with tandem-arranged repeat units were screened in the three SSH libraries. A set of sequences coding for genes involved in shell formation was obtained. RT-PCR and in situ hybridization assays were carried out on five genes to investigate their spatial expression in several tissues, especially the mantle tissue. They all showed a different expression pattern from known biomineralization-related genes. Inhibition of the five genes by RNA interference resulted in different defects of the nacreous layer, indicating that they all were involved in aragonite crystallization. Intriguingly, one gene (UD_Cluster94.seq.Singlet1) was restricted to the ‘aragonitic line’. The current data has yielded for the first time, to our knowledge, a suite of biomineralization-related genes active during the developmental stages of P.fucata, five of which were responsible for nacreous layer formation. This provides a useful starting point for isolating new genes involved in shell formation. The effects of genes on the formation of the ‘aragonitic line’, and other areas of the nacreous layer, suggests a different control mechanism for aragonite crystallization initiation from that of mature aragonite growth

Molecular mechanisms of cadmium-induced cytotoxicity in human ovarian granulosa cells identified using integrated omics

Epidemiological and clinical data have demonstrated that exposure to cadmium (Cd), a toxic heavy metal, is associated with an increased risk of female infertility. Granulosa cells, the main somatic cells comprising ovarian follicles, are one of the main targets of Cd in the ovaries. However, the mechanism by which Cd induces cytotoxicity in granulosa cells has not been fully elucidated. In this study, we exposed human ovarian granulosa cells (KGN cells) to Cd and conducted in vitro cell experiments and multi-omics (metabolomics and transcriptomics) methods to elucidate these mechanisms. Cd exposure was found to not only induce the apoptosis of the KGN cells but also further reduced mitochondrial function by decreasing mitochondrial membrane potential, ATP production, and respiratory chain complex activity as well as increasing mitochondrial reactive oxygen species (ROS) production. A total of 443 differentially expressed metabolites (160 upregulated and 283 downregulated) and 5200 differentially expressed genes (4634 upregulated and 566 downregulated) were observed in the Cd exposed-cells. The multi-omics data showed that Cd interfered with citric acid cycle (TCA cycle), amino acid (including alanine, glycine, serine, threonine, arginine, and proline) metabolism, and calcium signaling. These findings help to better elucidate the potential toxicity mechanisms of Cd on granulosa cells and the ovary

A review of the cascade refrigeration system

This paper provides a literature review of the cascade refrigeration system (CRS). It is an important system that can achieve an evaporating temperature as low as −170 °C and broadens the refrigeration temperature range of conventional systems. In this paper, several research options such as various designs of CRS, studies on refrigerants, and optimization works on the systems are discussed. Moreover, the influence of parameters on system performance, the economic analysis, and applications are defined, followed by conclusions and suggestions for future studies

Zinc finger and SCAN domain-containing protein 18 is a potential DNA methylation-modified tumor suppressor and biomarker in breast cancer

IntroductionZinc finger and SCAN domain-containing protein 18 (ZSCAN18) has been investigated as a putative biomarker of multiple human cancers. However, the expression profile, epigenetic modification, prognostic value, transcription regulation, and molecular mechanism of ZSCAN18 in breast cancer (BC) remain unknown.MethodsIn the study, we present an integrated analysis of ZSCAN18 in BC based on public omics datasets with the use of multiple bioinformatics tools. Genes potentially regulated through restoration of ZSCAN18 expression in MDA-MB-231 cells were investigated to identify pathways associated with BC.ResultsWe observed that ZSCAN18 was downregulated in BC and mRNA expression was significantly correlated with clinicopathological parameters. Low expression of ZSCAN18 was found in the HER2-positive and TNBC subtypes. High expression of ZSCAN18 was associated with good prognosis. As compared to normal tissues, the extent of ZSCAN18 DNA methylation was greater with fewer genetic alterations in BC tissues. ZSCAN18 was identified as a transcription factor that might be involved in intracellular molecular and metabolic processes. Low ZSCAN18 expression was associated with the cell cycle and glycolysis signaling pathway. Overexpression of ZSCAN18 inhibited mRNA expression of genes associated with the Wnt/β-catenin and glycolysis signaling pathways, including CTNNB1, BCL9, TSC1, and PFKP. ZSCAN18 expression was negatively correlated with infiltrating B cells and dendritic cells (DCs), as determined by the TIMER web server and reference to the TISIDB. ZSCAN18 DNA methylation was positively correlated with activated B cells, activated CD8+ and CD4+ T cells, macrophages, neutrophils, and activated DCs. Moreover, five ZSCAN18-related hub genes (KDM6B, KAT6A, KMT2D, KDM1A, and HSPBP1) were identified. ZSCAN18, ZNF396, and PGBD1 were identified as components of a physical complex.ConclusionZSCAN18 is a potential tumor suppressor in BC, as expression is modified by DNA methylation and associated with patient survival. In addition, ZSCAN18 plays important roles in transcription regulation, the glycolysis signaling pathway, and the tumor immune microenvironment

Overexpression of UGT74E2, an Arabidopsis IBA Glycosyltransferase, Enhances Seed Germination and Modulates Stress Tolerance via ABA Signaling in Rice

UDP-glycosyltransferases (UGTs) play key roles in modulating plant development and responses to environmental challenges. Previous research reported that the Arabidopsis UDP-glucosyltransferase 74E2 (AtUGT74E2), which transfers glucose to indole-3-butyric acid (IBA), is involved in regulating plant architecture and stress responses. Here, we show novel and distinct roles of UGT74E2 in rice. We found that overexpression of AtUGT74E2 in rice could enhance seed germination. This effect was also observed in the presence of IBA and abscisic acid (ABA), as well as salt and drought stresses. Further investigation indicated that the overexpression lines had lower levels of free IBA and ABA compared to wild-type plants. Auxin signaling pathway gene expression such as for OsARF and OsGH3 genes, as well as ABA signaling pathway genes OsABI3 and OsABI5, was substantially downregulated in germinating seeds of UGT74E2 overexpression lines. Consistently, due to reduced IBA and ABA levels, the established seedlings were less tolerant to drought and salt stresses. The regulation of rice seed germination and stress tolerance could be attributed to IBA and ABA level alterations, as well as modulation of the auxin/ABA signaling pathways by UGT74E2. The distinct roles of UGT74E2 in rice implied that complex and different molecular regulation networks exist between Arabidopsis and rice

Financial indicators of security of the banking system as indicators of competitiveness

На сучасному етапі розвитку економіки особливо гостро постає проблема досягнення конкурентоспроможного стану банківської сис- теми України, що неможливо без забезпечення її фінансової безпеки

Ubiquitylation Functions in the Calcium Carbonate Biomineralization in the Extracellular Matrix

<div><p>Mollusks shell formation is mediated by matrix proteins and many of these proteins have been identified and characterized. However, the mechanisms of protein control remain unknown. Here, we report the ubiquitylation of matrix proteins in the prismatic layer of the pearl oyster, <em>Pinctada fucata</em>. The presence of ubiquitylated proteins in the prismatic layer of the shell was detected with a combination of western blot and immunogold assays. The coupled ubiquitins were separated and identified by Edman degradation and liquid chromatography/mass spectrometry (LC/MS). Antibody injection <em>in vivo</em> resulted in large amounts of calcium carbonate randomly accumulating on the surface of the nacreous layer. These ubiquitylated proteins could bind to specific faces of calcite and aragonite, which are the two main mineral components of the shell. In the <em>in vitro</em> calcium carbonate crystallization assay, they could reduce the rate of calcium carbonate precipitation and induce the calcite formation. Furthermore, when the attached ubiquitins were removed, the functions of the EDTA-soluble matrix of the prismatic layer were changed. Their potency to inhibit precipitation of calcium carbonate was decreased and their influence on the morphology of calcium carbonate crystals was changed. Taken together, ubiquitylation is involved in shell formation. Although the ubiquitylation is supposed to be involved in every aspect of biophysical processes, our work connected the biomineralization-related proteins and the ubiquitylation mechanism in the extracellular matrix for the first time. This would promote our understanding of the shell biomineralization and the ubiquitylation processes.</p> </div

The physiological functions of the ubiquitylated proteins were inhibited by antibody injection.

<p>(A) SEM image of the inner surface of the low dosage antibody-injected group. The stair-like growth pattern was disturbed. (B) Enlargement of the box shown in (A), illustrating the crystals deposited on the inner nacreous layer surface. (C) SEM image of the inner surface of the high dosage antibody-injected group. More crystals were randomly accumulated. (D) Enlargement of the box shown in (C), illustrating that the crystals were linked together to form a new layer. (E) SEM image of the inner surface of preimmune rabbit serum injected <i>P. fucata</i> shell showing the stair-like growth pattern. (F) Enlargement of the box shown in (E), illustrating the flat tablets. (G) Energy-dispersive x-ray spectroscopy analysis of the deposition (black asterisk) shown in (D). Scale bars, 200 µm in (A) and (C); 50 µm in (E); 25 µm in (B) and (D); 10 µm in (F).</p

Ubiquitylation of <i>P. fucata</i> matrix proteins.

<p>(A) The ubiquitylated proteins were characterized by western blotting of EDTA extracts of nacre and prisms separated from the shell. The ubiquitylated proteins were mainly present in the EDTA-soluble matrix of calcitic prisms. P-ESM, EDTA-soluble matrix of the prismatic layer; P-EISM, the EDTA-insoluble matrix of the prismatic layer; N-ESM, EDTA-soluble matrix of the nacreous layer; N-EISM, the denatured fraction of the EDTA-insoluble matrix of the nacreous layer. (B) Time-course reaction of isopeptidase with the EDTA-soluble matrix fraction of the prismatic layer. Reaction products were analyzed by western blotting. The reaction was performed at 37°C with a volume of 15 µL containing 0.1 µM of isopeptidase, 2 µg of substrate, for the indicated times. Mono Ubi, mono-ubiquitin. (C) Amino acid sequence of ubiquitin showing the residues identified by Edman degradation (underlined) and the peptide sequences identified by LC-MS analysis (red highlights).</p