Selenophosphate synthetase 1 is an essential protein with roles in regulation of redox homoeostasis in mammals.

Selenophosphate synthetase (SPS) was initially detected in bacteria and was shown to synthesize selenophosphate, the active selenium donor. However, mammals have two SPS paralogues, which are designated SPS1 and SPS2. Although it is known that SPS2 catalyses the synthesis of selenophosphate, the function of SPS1 remains largely unclear. To examine the role of SPS1 in mammals, we generated a Sps1-knockout mouse and found that systemic SPS1 deficiency led to embryos that were clearly underdeveloped by embryonic day (E)8.5 and virtually resorbed by E14.5. The knockout of Sps1 in the liver preserved viability, but significantly affected the expression of a large number of mRNAs involved in cancer, embryonic development and the glutathione system. Particularly notable was the extreme deficiency of glutaredoxin 1 (GLRX1) and glutathione transferase Omega 1 (GSTO1). To assess these phenotypes at the cellular level, we targeted the removal of SPS1 in F9 cells, a mouse embryonal carcinoma (EC) cell line, which affected the glutathione system proteins and accordingly led to the accumulation of hydrogen peroxide in the cell. Furthermore, we found that several malignant characteristics of SPS1-deficient F9 cells were reversed, suggesting that SPS1 played a role in supporting and/or sustaining cancer. In addition, the overexpression of mouse or human GLRX1 led to a reversal of observed increases in reactive oxygen species (ROS) in the F9 SPS1/GLRX1-deficient cells and resulted in levels that were similar to those in F9 SPS1-sufficient cells. The results suggested that SPS1 is an essential mammalian enzyme with roles in regulating redox homoeostasis and controlling cell growth.

Insights into starch-based gels: Selection, fabrication, and application.

Starch a natural polymer, has made significant advancements in recent decades, offering superior performance and versatility compared to synthetic materials. This review discusses up-to-date diverse applications of starch gels, their fabrication techniques, and their advantages over synthetic materials. Starch gels renewability, biocompatibility, biodegradability, scalability, and affordability make them attractive. Also, advanced theoretical foundations and emerging industrial technologies could further expand their scope and functions inspiring new applications.

Targeted insertion of cysteine by decoding UGA codons with mammalian selenocysteine machinery.

Cysteine (Cys) is inserted into proteins in response to UGC and UGU codons. Herein, we show that supplementation of mammalian cells with thiophosphate led to targeted insertion of Cys at the UGA codon of thioredoxin reductase 1 (TR1). This Cys was synthesized by selenocysteine (Sec) synthase on tRNA([Ser]Sec) and its insertion was dependent on the Sec insertion sequence element in the 3'UTR of TR1 mRNA. The substrate for this reaction, thiophosphate, was synthesized by selenophosphate synthetase 2 from ATP and sulfide and reacted with phosphoseryl-tRNA([Ser]Sec) to generate Cys-tRNA([Ser]Sec). Cys was inserted in vivo at UGA codons in natural mammalian TRs, and this process was regulated by dietary selenium and availability of thiophosphate. Cys occurred at 10% of the Sec levels in liver TR1 of mice maintained on a diet with normal amounts of selenium and at 50% in liver TR1 of mice maintained on a selenium deficient diet. These data reveal a novel Sec machinery-based mechanism for biosynthesis and insertion of Cys into protein at UGA codons and suggest new biological functions for thiophosphate and sulfide in mammals.

Inhibition of PPO-related browning in fresh noodles: A combination of chemical and heat treatment.

Enzymatic browning has been a significant factor affecting the sale of fresh noodles. This study used a combination of physical and chemical methods to achieve a long-lasting and effective anti-browning effect in fresh noodles. The results showed that the combinations of citric acid (CA), NaOH, and KOH with heat treatment blunted the polyphenol oxidase activity and improved the color of fresh noodles. Specifically, the L* value of fresh noodles stored at 6 °C treated by the combination of CA and 75 °C (CHFN-75) at 72 h (81.71) was significantly higher than that of the control at 72 h (74.42). Mixolab and confocal laser scanning microscopy showed that the combined treatment affected the protein and starch of the flour. However, the hardness and chewiness of the cooked noodles increased only slightly, and the adhesiveness decreased slightly. The innovative combination can be used as an effective way to delay the darkening of fresh noodles.

Correlation between the expressions of metastasis-associated factor-1 in colon cancer and vacuolar ATP synthase.

BACKGROUND: Clinical prognosis often worsens due to high recurrence rates following radical surgery for colon cancer. The examination of high-risk recurrence factors post-surgery provides critical insights for disease evaluation and treatment planning. AIM: To explore the relationship between metastasis-associated factor-1 in colon cancer (MACC1) and vacuolar ATP synthase (V-ATPase) expression in colon cancer tissues, and recurrence rate in patients undergoing radical colon cancer surgery. METHODS: We selected 104 patients treated with radical colon cancer surgery at our hospital from January 2018 to June 2021. Immunohistochemical staining was utilized to assess the expression levels of MACC1 and V-ATPase in these patients. RESULTS: The rates of MACC1 and V-ATPase positivity were 64.42% and 67.31%, respectively, in colon cancer tissues, which were significantly higher than in paracancerous tissues (P < 0.05). Among patients with TNM stage III, medium to low differentiation, and lymph node metastasis, the positive rates of MACC1 and V-ATPase were significantly elevated in comparison to patients with TNM stage I-II, high differentiation, and no lymph node metastasis (P < 0.05). The rate of MACC1 positivity was 76.67% in patients with tumor diameters > 5 cm, notably higher than in patients with tumor diameters ≤ 5 cm (P < 0.05). We observed a positive correlation between MACC1 and V-ATPase expression (rs = 0.797, P < 0.05). The positive rates of MACC1 and V-ATPase were significantly higher in patients with recurrence compared to those without (P < 0.05). Logistic regression analysis revealed TNM stage, lymph node metastasis, MACC1 expression, and V-ATPase expression as risk factors for postoperative colon cancer recurrence (OR = 6.322, 3.435, 2.683, and 2.421; P < 0.05). CONCLUSION: The upregulated expression of MACC1 and V-ATPase in colon cancer patients appears to correlate with clinicopathological features and post-radical surgery recurrence.

l(2)01810 is a novel type of glutamate transporter that is responsible for megamitochondrial formation.

l(2)01810 causes glutamine-dependent megamitochondrial formation when it is overexpressed in Drosophila cells. In the present study, we elucidated the function of l(2)01810 during megamitochondrial formation. The overexpression of l(2)01810 and the inhibition of glutamine synthesis showed that l(2)01810 is involved in the accumulation of glutamate. l(2)01810 was predicted to contain transmembrane domains and was found to be localized to the plasma membrane. By using (14)C-labelled glutamate, l(2)01810 was confirmed to uptake glutamate into Drosophila cells with high affinity (K(m)=69.4 µM). Also, l(2)01810 uptakes glutamate in a Na(+)-independent manner. Interestingly, however, this uptake was not inhibited by cystine, which is a competitive inhibitor of Na(+)-independent glutamate transporters, but by aspartate. A signal peptide consisting of 34 amino acid residues targeting to endoplasmic reticulum was predicted at the N-terminus of l(2)01810 and this signal peptide is essential for the protein's localization to the plasma membrane. In addition, l(2)01810 has a conserved functional domain of a vesicular-type glutamate transporter, and Arg(146) in this domain was found to play a key role in glutamate transport and megamitochondrial formation. These results indicate that l(2)01810 is a novel type of glutamate transporter and that glutamate uptake is a rate-limiting step for megamitochondrial formation.

Advances in neuroimaging studies of alcohol use disorder (AUD).

Alcohol use disorder (AUD) is a worldwide problem and the most common substance use disorder. Chronic alcohol consumption may have negative effects on the body, the mind, the family, and even society. With the progress of current neuroimaging methods, an increasing number of imaging techniques are being used to objectively detect brain impairment induced by alcoholism and serve a vital role in the diagnosis, prognosis, and treatment assessment of AUD. This article organizes and analyzes the research on alcohol dependence concerning the main noninvasive neuroimaging methods, structural magnetic resonance imaging, functional magnetic resonance imaging, and electroencephalography, as well as the most common noninvasive brain stimulation - transcranial magnetic stimulation, and intersperses the article with joint intra- and intergroup studies, providing an outlook on future research directions.

Drosophila selenophosphate synthetase 1 regulates vitamin B6 metabolism: prediction and confirmation.

BACKGROUND: There are two selenophosphate synthetases (SPSs) in higher eukaryotes, SPS1 and SPS2. Of these two isotypes, only SPS2 catalyzes selenophosphate synthesis. Although SPS1 does not contain selenophosphate synthesis activity, it was found to be essential for cell growth and embryogenesis in Drosophila. The function of SPS1, however, has not been elucidated. RESULTS: Differentially expressed genes in Drosophila SL2 cells were identified using two-way analysis of variance methods and clustered according to their temporal expression pattern. Gene ontology analysis was performed against differentially expressed genes and gene ontology terms related to vitamin B6 biosynthesis were found to be significantly affected at the early stage at which megamitochondria were not formed (day 3) after SPS1 knockdown. Interestingly, genes related to defense and amino acid metabolism were affected at a later stage (day 5) following knockdown. Levels of pyridoxal phosphate, an active form of vitamin B6, were decreased by SPS1 knockdown. Treatment of SL2 cells with an inhibitor of pyridoxal phosphate synthesis resulted in both a similar pattern of expression as that found by SPS1 knockdown and the formation of megamitochondria, the major phenotypic change observed by SPS1 knockdown. CONCLUSIONS: These results indicate that SPS1 regulates vitamin B6 synthesis, which in turn impacts various cellular systems such as amino acid metabolism, defense and other important metabolic activities.

Inhibition of selenocysteine tRNA[Ser]Sec aminoacylation provides evidence that aminoacylation is required for regulatory methylation of this tRNA.

There are two isoforms of selenocysteine (Sec) tRNA([Ser]Sec) that differ by a single methyl group, Um34. The non-Um34 isoform supports the synthesis of a subclass of selenoproteins, designated housekeeping, while the Um34 isoform supports the expression of another subclass, designated stress-related selenoproteins. Herein, we investigated the relationship between tRNA([Ser]Sec) aminoacylation and Um34 synthesis which is the last step in the maturation of this tRNA. Mutation of the discriminator base at position 73 in tRNA([Ser]Sec) dramatically reduced aminoacylation with serine, as did an inhibitor of seryl-tRNA synthetase, SB-217452. Although both the mutation and the inhibitor prevented Um34 synthesis, neither precluded the synthesis of any other of the known base modifications on tRNA([Ser]Sec) following microinjection and incubation of the mutant tRNA([Ser]Sec) transcript, or the wild type transcript along with inhibitor, in Xenopus oocytes. The data demonstrate that Sec tRNA([Ser]Sec) must be aminoacylated for Um34 addition. The fact that selenium is required for Um34 methylation suggests that Sec must be attached to its tRNA for Um34 methylation. This would explain why selenium is essential for the function of Um34 methylase and provides further insights into the hierarchy of selenoprotein expression.

[A study on allele frequencies and mismatching proportion of HLA-A, B, Cw, DRB1 and DQB1 on high-resolution donor-recipient typing in Chinese Han population].

OBJECTIVE: To analyze the allele frequencies and polymorphism of human leukocyte antigens (HLA) -A, B, Cw, DRB1 and DQB1 between donors-recipients on high-resolution typing; and to analyze the matching and mismatching proportion between donors and recipients. METHODS: HLA high-resolution types were determined by sequence based typing (SBT), sequence specific oligonucleotide probe (SSOP) and sequence specific primer (SSP) on 2540 unrelated Chinese Han individuals including 1168 recipients and 1372 donors, then statistical analyses were carried out. RESULTS: Forty-four HLA-A alleles were detected, and among them the frequencies of A*1101, A*2402, A*0201, A*0207, A*3303, A*0206 and A*3001 exceeded 0.05, and accounted for 80.4%. Eighty-one HLA-B alleles were detected, and the frequencies of B*4001, B*4601, B*5801, B*1302 and B*5101 exceeded 0.05, and accounted for 43.0% of total. There were 44 HLA-Cw alleles, among them the frequencies of Cw*0702, Cw*0102, Cw*0304, Cw*0801, Cw*0602, Cw*0303, Cw*0302 and Cw*0401 exceeded 0.05, and were 80.3% of total. There were 61 HLA-DRB1 alleles, the frequencies of DRB1*0901, DRB1*1501, DRB1*1202, DRB1*0803, DRB1*0701, DRB1*0405, DRB1*0301 and DRB1*1101 exceeded 0.05, and were 70.1% of total. Finally, 22 HLA-DQB1 alleles were detected, the frequencies of DQB1*0301, DQB1*0303, DQB1*0601, DQB1*0602, DQB1*0202, DQB1*0302, DQB1*0401, DQB1*0502 and DQB1*0201 exceeded 0.05, and they were 87.4% of total. All the five loci were of heterozygote deficiency. The HLA-A, B and DRB1 loci conformed to Hardy-Weinberg equilibrium (HWE) (P > 0.05); but HLA-Cw and HLA-DQB1 loci did not (P < 0.05). Except several particular genotypes, all the five loci conformed to HWE. After comparing data between donors and recipients, only 22.4% of recipients found HLA matched donors (10/10); 24.6% of recipients found single HLA allele mismatched donors (9/10); 26.3% of recipients had two HLA alleles mismatched donors (8/10). CONCLUSION: The characteristics of allele frequencies and polymorphism of HLA-A, B, Cw, DRB1 and DQB1 on high-resolution typing in Chinese Han population is valuable for donor searching in unrelated hematopoietic stem cell transplantation, and it provides genetic basis for donor registry and usage of donor resource for Chinese Marrow Donor Program.

Elevation of glutamine level by selenophosphate synthetase 1 knockdown induces megamitochondrial formation in Drosophila cells.

Although selenophosphate synthetase 1 (SPS1/SelD) is an essential gene in Drosophila, its function has not been determined. To elucidate its intracellular role, we targeted the removal of SPS1/SelD mRNA in Drosophila SL2 cells using RNA interference technology that led to the formation of vacuole-like globular structures. Surprisingly, these structures were identified as megamitochondria, and only depolarized mitochondria developed into megamitochondria. The mRNA levels of l(2)01810 and glutamine synthetase 1 (GS1) were increased by SPS1/SelD knockdown. Blocking the expression of GS1 and l(2)01810 completely inhibited the formation of megamitochondria induced by loss of SPS1/SelD activity and decreased the intracellular levels of glutamine to those of control cells suggesting that the elevated level of glutamine is responsible for megamitochondrial formation. Overexpression of GS1 and l(2)01810 had a synergistic effect on the induction of megamitochondrial formation and on the synthesis of glutamine suggesting that l(2)01810 is involved in glutamine synthesis presumably by activating GS1. Our results indicate that, in Drosophila, SPS1/SelD regulates the intracellular glutamine by inhibiting GS1 and l(2)01810 expression and that elevated levels of glutamine lead to a nutritional stress that provides a signal for megamitochondrial formation.

Human selenophosphate synthetase 1 has five splice variants with unique interactions, subcellular localizations and expression patterns.

Selenophosphate synthetase 1 (SPS1) is an essential cellular gene in higher eukaryotes. Five alternative splice variants of human SPS1 (major type, DeltaE2, DeltaE8, +E9, +E9a) were identified wherein +E9 and +E9a make the same protein. The major type was localized in both the nuclear and plasma membranes, and the others in the cytoplasm. All variants form homodimers, and in addition, the major type forms a heterodimer with DeltaE2, and DeltaE8 with +E9. The level of expression of each splice variant was different in various cell lines. The expression of each alternative splice variant was regulated during the cell cycle. The levels of the major type and DeltaE8 were gradually increased until G2/M phase and then gradually decreased. DeltaE2 expression peaked at mid-S phase and then gradually decreased. However, +E9/+E9a expression decreased gradually after cell cycle arrest. The possible involvement of SPS1 splice variants in cell cycle regulation is discussed.

Biosynthesis of selenocysteine on its tRNA in eukaryotes.

Selenocysteine (Sec) is cotranslationally inserted into protein in response to UGA codons and is the 21st amino acid in the genetic code. However, the means by which Sec is synthesized in eukaryotes is not known. Herein, comparative genomics and experimental analyses revealed that the mammalian Sec synthase (SecS) is the previously identified pyridoxal phosphate-containing protein known as the soluble liver antigen. SecS required selenophosphate and O-phosphoseryl-tRNA([Ser]Sec) as substrates to generate selenocysteyl-tRNA([Ser]Sec). Moreover, it was found that Sec was synthesized on the tRNA scaffold from selenide, ATP, and serine using tRNA([Ser]Sec), seryl-tRNA synthetase, O-phosphoseryl-tRNA([Ser]Sec) kinase, selenophosphate synthetase, and SecS. By identifying the pathway of Sec biosynthesis in mammals, this study not only functionally characterized SecS but also assigned the function of the O-phosphoseryl-tRNA([Ser]Sec) kinase. In addition, we found that selenophosphate synthetase 2 could synthesize monoselenophosphate in vitro but selenophosphate synthetase 1 could not. Conservation of the overall pathway of Sec biosynthesis suggests that this pathway is also active in other eukaryotes and archaea that synthesize selenoproteins.

Effect of re-acetylation on the acid hydrolysis of chitosan under an induced electric field.

This study explored the impact of re-acetylation on induced electric field (IEF)-assisted hydrolysis of chitosan. Chitosan with different degree of deacetylation (DD) exhibited different charge content but similar electrical conductivity and output voltage of the electrolyte system, thus theoretically resulting in equivalent electro-processing efficiency. However, molecular and rheological measurements suggested that the hydrolysis rate decreased with an increase of DD. Re-acetylation had no significant effect on the molecular weight and viscosity of chitosan but damaged its granular and crystal structure, making chitosan chains more susceptible to the acid. The IEF-assisted hydrolysis of dissolved chitosan confirmed that the structural compactness of chitosan played a more important role on the hydrolysis than the charge content. This study extended current knowledge regarding the effect of charge content and structural compactness on IEF-assisted hydrolysis process, which contributed to the applications of IEF in biopolymer modification.

Supramolecular complexes mediate selenocysteine incorporation in vivo.

Selenocysteine incorporation in eukaryotes occurs cotranslationally at UGA codons via the interactions of RNA-protein complexes, one comprised of selenocysteyl (Sec)-tRNA([Ser]Sec) and its specific elongation factor, EFsec, and another consisting of the SECIS element and SECIS binding protein, SBP2. Other factors implicated in this pathway include two selenophosphate synthetases, SPS1 and SPS2, ribosomal protein L30, and two factors identified as binding tRNA([Ser]Sec), termed soluble liver antigen/liver protein (SLA/LP) and SECp43. We report that SLA/LP and SPS1 interact in vitro and in vivo and that SECp43 cotransfection increases this interaction and redistributes all three proteins to a predominantly nuclear localization. We further show that SECp43 interacts with the selenocysteyl-tRNA([Ser]Sec)-EFsec complex in vitro, and SECp43 coexpression promotes interaction between EFsec and SBP2 in vivo. Additionally, SECp43 increases selenocysteine incorporation and selenoprotein mRNA levels, the latter presumably due to circumvention of nonsense-mediated decay. Thus, SECp43 emerges as a key player in orchestrating the interactions and localization of the other factors involved in selenoprotein biosynthesis. Finally, our studies delineating the multiple, coordinated protein-nucleic acid interactions between SECp43 and the previously described selenoprotein cotranslational factors resulted in a model of selenocysteine biosynthesis and incorporation dependent upon both cytoplasmic and nuclear supramolecular complexes.

The impact of sodium carbonate on physico-chemical properties and cooking qualities of starches isolated from alkaline yellow noodles.

The physicochemical and cooking properties of wheat starch isolated from alkaline yellow dough treated with sodium carbonate (Na2CO3; 0-3.2 g/100 g) were investigated. With increasing Na2CO3 addition, swelling power increased from 7.28 to 10.70 g/g. X-ray diffraction showed no changes in crystalline patterns while the relative crystallinity decreased from 30.11% to 23.13%. Differential scanning calorimetry results suggested that alkaline salt shifted the gelatinization peak of starch to higher temperatures. The values of pasting viscosity and pasting temperature in alkali-treated starch increased and decreased, respectively. Farinograph results revealed the strengthened structure of dough with alkali-treated starch that was manifested by an increase in the dough development time and dough stability time. Cooking loss and rehydration values of noodles prepared from alkali-treated starch increased by 42% and 36%, respectively. The results suggested that Na2CO3 affected starch crystalline structure, swelling power, gelatinization, pasting properties, starch-gluten interactions and cooking characteristics of noodle products.

Protective effect of isoflavones from Trifolium pratense on dopaminergic neurons.

In the present study, protective effect of five isoflavones (formononetin, daidzein, pratensein, calycosin and irilone) from Trifolium pratense on lipopolysaccharide-induced dopaminergic neurodegeneration was studied for the first time. The results showed that all five isoflavones attenuated LPS-induced decrease in dopamine uptake and the number of dopaminergic neurons in a dose-dependent manner in rat mesencephalic neuron-glia cultures. Moreover, they also significantly inhibited LPS-induced activation of microglia and production of tumor necrosis factor-alpha, nitric oxide and superoxide in mesencephalic neuron-glia cultures and microglia-enriched cultures. In addition, the rank order of protective potency of five isoflavones was: pratensein>daidzein>calycosin>formononetin>irilone. This study suggested that all five isoflavones protected dopaminergic neurons against LPS-induced injury through inhibition of microglia activation and proinflammatory factors generation.

Luteolin protects dopaminergic neurons from inflammation-induced injury through inhibition of microglial activation.

Parkinson's disease is a neurodegenerative disorder characterized by progressive degeneration of dopaminergic neurons in the substantia nigra. Accumulating evidence has suggested that inflammation in the brain participates in the pathogenesis of Parkinson's disease. Luteolin, a polyphenolic compound found in foods of plant origin, belongs to the flavone subclass of flavonoids, and has been shown to possess antimutagenic, antitumorigenic, antioxidant and antiinflammatory properties. In this study, we found that luteolin concentration-dependently attenuated the lipopolysaccharide (LPS)-induced decrease in [(3)H]dopamine uptake and loss of tyrosine hydroxylase-immunoreactive neurons in primary mesencephalic neuron-glia cultures. Moreover, luteolin also significantly inhibited LPS-induced activation of microglia and excessive production of tumor necrosis factor-alpha, nitric oxide and superoxide in mesencephalic neuron-glia cultures and microglia-enriched cultures. Our results demonstrate that luteolin may protect dopaminergic neurons from LPS-induced injury and its efficiency in inhibiting microglia activation may underlie the mechanism.

Selenophosphate synthetase 2 is essential for selenoprotein biosynthesis.

Selenophosphate synthetase (SelD) generates the selenium donor for selenocysteine biosynthesis in eubacteria. One homologue of SelD in eukaryotes is SPS1 (selenophosphate synthetase 1) and a second one, SPS2, was identified as a selenoprotein in mammals. Earlier in vitro studies showed SPS2, but not SPS1, synthesized selenophosphate from selenide, whereas SPS1 may utilize a different substrate. The roles of these enzymes in selenoprotein synthesis in vivo remain unknown. To address their function in vivo, we knocked down SPS2 in NIH3T3 cells using small interfering RNA and found that selenoprotein biosynthesis was severely impaired, whereas knockdown of SPS1 had no effect. Transfection of SPS2 into SPS2 knockdown cells restored selenoprotein biosynthesis, but SPS1 did not, indicating that SPS1 cannot complement SPS2 function. These in vivo studies indicate that SPS2 is essential for generating the selenium donor for selenocysteine biosynthesis in mammals, whereas SPS1 probably has a more specialized, non-essential role in selenoprotein metabolism.

HPTLC-FLD-SERS as a facile and reliable screening tool: Exemplarily shown with tyramine in cheese.

The serious cytotoxicity of tyramine attracted marked attention as it induced necrosis of human intestinal cells. This paper presented a novel and facile high performance thin-layer chromatography (HPTLC) method tailored for screening tyramine in cheese. Separation was performed on glass backed silica gel plates, using methanol/ethyl acetate/ammonia (6/4/1 v/v/v) as the mobile phase. Special efforts were focused on optimizing conditions (substrate preparation, laser wavelength, salt types and concentrations) of surface enhanced Raman spectroscopy (SERS) measurements directly on plates after derivatization, which enabled molecule-specific identification of targeted bands. In parallel, fluorescent densitometry (FLD) scanning at 380