Loss of O-GlcNAcylation on MeCP2 at Threonine 203 Leads to Neurodevelopmental Disorders / 神经科学通报·英文版

Mutations of the X-linked methyl-CpG-binding protein 2 (MECP2) gene in humans are responsible for most cases of Rett syndrome (RTT), an X-linked progressive neurological disorder. While genome-wide screens in clinical trials have revealed several putative RTT-associated mutations in MECP2, their causal relevance regarding the functional regulation of MeCP2 at the etiologic sites at the protein level requires more evidence. In this study, we demonstrated that MeCP2 was dynamically modified by O-linked-β-N-acetylglucosamine (O-GlcNAc) at threonine 203 (T203), an etiologic site in RTT patients. Disruption of the O-GlcNAcylation of MeCP2 specifically at T203 impaired dendrite development and spine maturation in cultured hippocampal neurons, and disrupted neuronal migration, dendritic spine morphogenesis, and caused dysfunction of synaptic transmission in the developing and juvenile mouse cerebral cortex. Mechanistically, genetic disruption of O-GlcNAcylation at T203 on MeCP2 decreased the neuronal activity-induced induction of Bdnf transcription. Our study highlights the critical role of MeCP2 T203 O-GlcNAcylation in neural development and synaptic transmission potentially via brain-derived neurotrophic factor.

Preliminary exploration of the metabolic profile and metabolic pathways in newly diagnosed multiple myeloma / 中华肿瘤杂志

Objective: To explore the metabolite profile and metabolic pathways of newly diagnosed multiple myeloma (MM). Methods: Gas chromatography-mass spectrometry (GC-MS) was employed for the high-throughput detection and identification of serum samples from 55 patients with MM and 37 healthy controls matched for age and sex from 2016 to 2017 collected at the First Affiliated Hospital of Soochow University. The relative standard deviation (RSD) of quality control (QC) samples was employed to validate the reproducibility of GC-MS approach. The differential metabolites between patients with MM and healthy controls were detected by partial least squares discrimination analysis (PLS-DA), and t-test with false discovery rate (FDR) correction. Metabolomics pathway analysis (MetPA) was employed to construct metabolic pathways. Results: There were 55 MM patients, including 34 males and 21 females. The median age was 60 years old (42-73 years old). There were 30 cases of IgG type, 9 cases of IgA type, 1 case of IgM type, 2 cases of non-secreted type, 1 case of double clone type and 12 cases of light chain type, including 3 cases of kappa light chain type and 9 cases of lambda light chain type. The result of QC sample test showed that the proportion of compounds with the RSD of the relative content of metabolites < 15% was 70.21% obtained by the reproducibility of GC-MS experimental data, which implied that the experimental data were reliable. A total of 17 metabolites were screened differently with the healthy control group, including myristic acid, hydroxyproline, cysteine, palmitic acid, L-leucine, stearic acid, methionine, phenylalanine, glycerin, serine, isoleucine, tyrosine, valine, citric acid, inositol, threonine, and oxalic acid (VIP>1, P<0.05). Metabolic pathway analysis suggested that metabolic disorders in MM patients comprised mainly phenylalanine metabolism, glyoxylic acid and dicarboxylic acid metabolism, phosphoinositide metabolism, cysteine and methionine metabolism, glycerolipid metabolism, glycine, serine, and threonine metabolism. Conclusion: Compared with normal people, patients with newly diagnosed MM have obvious differences in metabolic profiles and metabolic pathways.

Biosynthesis of 2,5-dimethylpyrazine from L-threonine by whole-cell biocatalyst of recombinant Escherichia coli / 生物工程学报

2,5-dimethylpyrazine (2,5-DMP) is of important economic value in food industry and pharmaceutical industry, and is now commonly produced by chemical synthesis. In this study, a recombinant Escherichia coli high-efficiently converting L-threonine to 2,5-DMP was constructed by combination of metabolic engineering and cofactor engineering. To do this, the effect of different threonine dehydrogenase (TDH) on 2,5-DMP production was investigated, and the results indicate that overexpression of EcTDH in E. coli BL21(DE3) was beneficial to construct a 2,5-DMP producer with highest 2,5-DMP production. The recombinant strain E. coli pRSFDuet-tdh(Ec) produced (438.3±23.7) mg/L of 2,5-DMP. Furthermore, the expression mode of NADH oxidase (NoxE) from Lactococcus cremoris was optimized, and fusion expression of EcTDH and LcNoxE led to balance the intracellular NADH/NAD⁺ level and to maintain the high survival rate of cells, thus further increasing 2,5-DMP production. Finally, the accumulation of by-products was significantly decreased because of disruption of shunt metabolic pathway, thereby increasing 2,5-DMP production and the conversion ratio of L-threonine. Combination of these genetic modifications resulted in an engineered E. coli Δkbl ΔtynA ΔtdcB ΔilvA pRSFDuet-tdhEcnoxELc-PsstT (EcΔkΔAΔBΔA/TDH(Ec)NoxE(Lc)-PSstT) capable of producing (1 095.7±81.3) mg/L 2,5-DMP with conversion ratio of L-threonine of 76% and a yield of 2,5-DMP of 28.8% in 50 mL transformation system with 5 g/L L-threonine at 37 °C and 200 r/min for 24 h. Therefore, this study provides a recombinant E. coli with high-efficiently catalyzing L-threonine to biosynthesize 2,5-DMP, which can be potentially used in biosynthesis of 2,5-DMP in industry.

Catalytic mechanism, molecular engineering and applications of threonine aldolases / 生物工程学报

Threonine aldolases catalyze the aldol condensation of aldehydes with glycine to furnish β-hydroxy-α-amino acid with two stereogenic centers in a single reaction. This is one of the most promising green methods for the synthesis of optically pure β-hydroxy-α-amino acid with high atomic economy and less negative environmental impact. Several threonine aldolases from different origins have been identified and characterized. The insufficient -carbon stereoselectivity and the challenges of balancing kinetic versus thermodynamic control to achieve the optimal optical purity and yield hampered the application of threonine aldolases. This review summarizes the recent advances in discovery, catalytic mechanism, high-throughput screening, molecular engineering and applications of threonine aldolases, with the aim to provide some insights for further research in this field.

Production of L-2-aminobutyric acid from L-threonine using a trienzyme cascade / 生物工程学报

L-2-aminobutyric acid (L-ABA) is an important chemical raw material and chiral pharmaceutical intermediate. The aim of this study was to develop an efficient method for L-ABA production from L-threonine using a trienzyme cascade route with Threonine deaminase (TD) from Escherichia. coli, Leucine dehydrogenase (LDH) from Bacillus thuringiensis and Formate dehydrogenase (FDH) from Candida boidinii. In order to simplify the production process, the activity ratio of TD, LDH and FDH was 1:1:0.2 after combining different activity ratios in the system in vitro. The above ratio was achieved in the recombinant strain E. coli 3FT+L. Moreover, the transformation conditions were optimized. Finally, we achieved L-ABA production of 68.5 g/L with a conversion rate of 99.0% for 12 h in a 30-L bioreactor by whole-cell catalyst. The environmentally safe and efficient process route represents a promising strategy for large-scale L-ABA production in the future.

Betaine supplementation improved L-threonine fermentation of Escherichia coli THRD by upregulating zwf (glucose-6-phosphate dehydrogenase) expression

BACKGROUND: The supplementation of betaine, an osmoprotective compatible solute, in the cultivation media has been widely used to protect bacterial cells. To explore the effects of betaine addition on industrial fermentation, Escherichia coli THRD, an L-threonine producer, was used to examine the production of L-threonine with betaine supplementation and the underlying mechanism through which betaine functions was investigated. RESULTS: Betaine supplementation in the medium of E. coli THRD significantly improved L-threonine fermentation parameters. The transcription of zwf and corresponding enzyme activity of glucose-6-phosphate dehydrogenase were significantly promoted by betaine addition, which contributed to an enhanced expression of zwf that provided more nicotinamide adenine dinucleotide phosphate (NADPH) for L-threonine synthesis. In addition, as a result of the betaine addition, the betaine-stimulated expression of enhanced green fluorescent protein (eGFP) under the zwf promoter within a plasmid-based cassette proved to be a transcription-level response of zwf. Finally, the promoter of the phosphoenolpyruvate carboxylase gene ppc in THRD was replaced with that of zwf, while L-threonine fermentation of the new strain was promoted by betaine addition. Conclusions: We reveal a novel mode of betaine that facilitates the microbial production of useful compounds. Betaine supplementation upregulates the expression of zwf and increases the NADPH synthesis, which may be beneficial for the cell growth and thereby promote the production of L-threonine. This finding might be useful for the production of NADPH-dependent amino acids and derivatives in E. coli THRD or other E. coli strains.

Anti-Helicobacter pylori, Anti-apoptotic, and Cytoprotective Effects of Threonine Synthesized from Corynebacterium glutamicum in Gastric Epithelial Cells

BACKGROUND/AIMS: Among irritants causing gastric ulcer, Helicobacter pylori (H. pylori) might be pivotal, after which eradication became essential way in either inhibiting ulcerogenesis or preventing ulcer recurrence. Since threonine is essential in either mucus synthesis or cytoprotection, we hypothesized that the dietary threonine from Corynebacterium glutamicum (C. glutamicum) can mitigate the cytotoxicity of H. pylori infection.MATERIALS AND METHODS: RGM-1 cells were challenged with 100 multiplicity of infection H. pylori for 6 hours, during which threonine alone or combination with Corynebacterium sp. was administered and compared for anti-Helicobacter, anti-inflammation, anti-oxidative, and cytoprotective actions.RESULTS: Threonine alone or combination of threonine and C. glutamicum yielded significant bacteriostatic outcomes. The increased expressions of interleukin (IL)-1β, IL-8, Cox-2, and iNOS mRNA after H. pylori infection were significantly decreased with either threonine alone or the combination of threonine and C. glutamicum. The elevated expressions of NF-kB, HIF-1a, and c-jun after H. pylori infection were all significantly decreased with the combination of threonine and broth from C. glutamicum (P < 0.05), leading to significant decreases in 2′,7′-dichlorofluorescein-diacetate (P < 0.01). Tracing further host antioxidative response, the attenuated expression of heme oxygenase-1, Nrf2, and dehydrogenase quinone-1 after H. pylori infection was significantly preserved with combination of threonine and C. glutamicum. H. pylori infection led to significant increases in apoptosis accompanied with Bcl-2 decreases and Bax increases, while the combination of threonine and C. glutamicum significantly attenuated apoptosis, in which attenuated EGF, TGF-β, and VEGF were significantly regulated, while β-catenin did not change.CONCLUSIONS: Threonine synthesized from C. glutamicum significantly alleviated the cytotoxicity of H. pylori in gastric epithelial cells.

MicroRNA-206 Reduces Osteosarcoma Cell Malignancy In Vitro by Targeting the PAX3-MET Axis

PURPOSE: This study was undertaken to explore how miR-206 represses osteosarcoma (OS) development. MATERIALS AND METHODS: Expression levels of miR-206, PAX3, and MET mRNA were explored in paired OS and adjacent tissue specimens. A patient-derived OS cell line was established. miR-206 overexpression and knockdown were achieved by lentiviral transduction. PAX3 and MET overexpression were achieved by plasmid transfection. Treatment with hepatocyte growth factor (HGF) was utilized to activate c-Met receptor. Associations between miR-206 and PAX3 or MET mRNA in OS cells were verified by AGO2-RNA immunoprecipitation assay and miRNA pulldown assay. OS cell malignancy was evaluated in vitro by cell proliferation, metastasis, and apoptosis assays. PAX3 and MET gene expression in OS cells was assayed by RT-qPCR and Western blot. Activation of PI3K-AKT and MAPK-ERK in OS cells were assayed by evaluating Akt1 Ser473 phosphorylation and total threonine phosphorylation of Erk1/2, respectively. RESULTS: Expression levels of miR-206 were significantly decreased in OS tissue specimens, compared to adjacent counterparts, and were inversely correlated with expression of PAX3 and MET mRNA. miR-206 directly interacted with PAX3 and MET mRNA in OS cells. miR-206 overexpression significantly reduced PAX3 and MET gene expression in OS cells in vitro, resulting in significant decreases in Akt1 and Erk1/2 activation, cell proliferation, and metastasis, as well as increases in cell apoptosis, while miR-206 knockdown showed the opposite effects. The effects of miR-206 overexpression on OS cells were reversed by PAX3 or MET overexpression, but only partially attenuated by HGF treatment. CONCLUSION: miR-206 reduces OS cell malignancy in vitro by targeting PAX3 and MET gene expression.

Therapeutic Effects of Amino Acids in Liver Diseases: Current Studies and Future Perspectives

Hepatocellular carcinoma (HCC) is the most common primary malignant tumor of the liver and the third most common cause of cancer-related death worldwide. HCC is caused by infection of hepatitis B/C virus and liver dysfunctions, such as alcoholic liver disease, nonalcoholic fatty liver disease, and cirrhosis. Amino acids are organic substances containing amine and carboxylic acid functional groups. There are over 700 kinds of amino acids in nature, but only about 20 of them are used to synthesize proteins in cells. Liver is an important organ for protein synthesis, degradation and detoxification as well as amino acid metabolism. In the liver, there are abundant non-essential amino acids, such as alanine, aspartate, glutamate, glycine, and serine and essential amino acids, such as histidine and threonine. These amino acids are involved in various cellular metabolisms, the synthesis of lipids and nucleotides as well as detoxification reactions. Understanding the role of amino acids in the pathogenesis of liver and the effects of amino acid intake on liver disease can be a promising strategy for the prevention and treatment of liver disease. In this review, we describe the biochemical properties and functions of amino acids and to review how they have been applied to treatment of liver diseases.

Effects of serine/threonine-protein kinase B-Raf-activated long-chain non-coding RNA on apoptosis and autophagy in thyroid carcinoma cells / 中南大学学报(医学版)

To investigate the effect of serine/threonine-protein kinase B-Raf (BRAF)-activated long-chain non-coding RNA (lncRNA-BANCR) on apoptosis and autophagy in thyroid carcinoma cells and the underlying mechanisms.
 Methods: RT-PCR was used to detect the expression of lncRNA-BANCR in thyroid carcinoma and normal thyroid tissues. The association between lncRNA-BANCR and clinicopathological data was analyzed in patients with thyroid cancer. Cell counting kit-8 (CCK-8) was used to detect the effect of lncRNA-BANCR on the proliferation of thyroid cancer cells. The effect of lncRNA-BANCR on the apoptosis of thyroid carcinoma cells was detected by flow cytometry. Transwell invasion assay was used to detect the effect of lncRNA-BANCR on the invasive ability of thyroid cancer cells. Western blot was used to detect the changes of autophagy proteins LC3-I and LC3-II after the lncRNA-BANCR expression was suppressed.
 Results: Compared with normal thyroid tissues, the expression level of lncRNA-BANCR in thyroid carcinoma tissues was elevated (P<0.05). The expression of lncRNA-BANCR was positively related to the pathological stage of thyroid carcinoma and the lymph node metastasis. Inhibition of lncRNA-BANCR expression attenuated the proliferation and invasion ability of thyroid cancer cells (both P<0.05); but the apoptosis was enhanced (P<0.05); the expression levels of autophagy protein LC3-I and LC3-II were also increased (P<0.05).
 Conclusion: The expression level of lncRNA-BANCR affects the proliferation, invasion and apoptosis of thyroid cancer cells through modulation of autophagy behavior.

Estimação de valores genéticos para codornas europeias em função dos níveis da relação treonina: lisina da dieta: do nascimento aos 21 dias de idade / Estimation of breeding values for European quails in function of threonine: lysine ratios of diet: from birth to 21 days of age

O presente trabalho foi realizado com o objetivo de avaliar a sensibilidade dos valores genéticos dos pesos corporais e as características de carcaças de codornas europeias às mudanças do gradiente ambiental (níveis da relação treonina com a lisina das dietas), do nascimento aos 21 dias de idade, por meio de modelos de regressão aleatória com diferentes classes de variância residual. Os dados utilizados neste estudo são provenientes de 915 codornas de corte da linhagem LF1 e 839 da linhagem LF2, pertencentes ao Programa de Melhoramento Genético da Universidade Federal dos Vales do Jequitinhonha e Mucuri. Foram avaliados os pesos corporais e os rendimentos da carcaça das aves. As sensibilidades dos valores genéticos às mudanças nos níveis da relação treonina:lisina (interação genótipo x ambiente) foram obtidas por modelos de regressão aleatória (utilizando normas de reação) por meio do programa Wombat, que utiliza o princípio da máxima verossimilhança restrita (REML). O modelo de regressão aleatória que considerou duas classes de variância residual foi o mais indicado para a maioria das análises realizadas. Verificaram-se alterações na classificação dos valores genéticos para as duas linhagens de codornas de corte estudadas. Esse comportamento indica sensibilidade de valores genéticos aditivos às mudanças nutricionais, o que caracteriza a existência de interação genótipo x ambiente. A predição dos valores genéticos deve ser feita com o mesmo nível da relação treonina:lisina da dieta com a qual as codornas serão alimentadas no sistema de produção.(AU)

This research was carried out to evaluate the sensitivity of breeding values of body weight and carcass traits in two lines of European quails (LF1 and LF2) to changes in the environment gradient (levels of threonine: lysine ratio of diets) from hatch to 21 days of age in two lines LF1 and LF2 using Random Regression Models with different classes of residual variance. Records are from 915 quails of line LF1 and 839 of line LF2 belonging to the Breeding Improvement Program of Universidade Federal dos Vales do Jequitinhonha e Mucuri. Live body weight and weights and yields of carcass, breast, and thigh and drumstick were measured. The sensitivities of breeding values to changes in threonine: lysine ratios (genotype x environment interaction) of diets were obtained by random regression models (reaction model) using the WOMBAT program using the Restricted Maximum Likelihood principle. Model considering two classes of residual variance showed the best goodness of fit. The Reaction Norms analyses indicated changes in the ranking of breeding values for both lines suggesting quails selected in one level of threonine: lysine ratio will not express all their genetic potential if fed different threonine: lysine ratio diets. This behavior indicates sensitivity of breeding values to changes in the nutrition characterizing the genotype by environment interaction. The prediction of breeding values must be performed using the same level of threonine: lysine ratio in diet the quails will be fed in the production system.(AU)

Reserpine treatment activates AMP activated protein kinase (AMPK)

Reserpine is a well-known medicine for the treatment of hypertension, however the role of reserpine in cell signaling is not fully understood. Here, we report that reserpine treatment induces the phosphorylation of AMP activated protein kinase (AMPK) at threonine 172 (T172) in PC12 cells. Phosphorylation of AMPK T172 is regulated by upstream signaling molecules, and the increase of phospho-T172 indicates that AMPK is activated. When we examined the FOXO3a dependent transcription by using the FHRE-Luc reporter assay, reserpine treatment repressed the FHRE-Luc reporter activity in a dose dependent manner. Finally, we showed that reserpine treatment induced the phosphorylation of AMPK as well as cell death in MCF-7 cells. These results suggest that AMPK is a potential cellular target of reserpine.

Morfometria intestinal em codornas de corte alimentadas com treonina digestível / Intestinal morphometry in meat quails fed with digestible threonine

O objetivo deste estudo foi avaliar a morfometria intestinal por meio da inclusão de níveis de treonina digestível na dieta de codornas para corte em crescimento. O delineamento experimental adotado foi o inteiramente ao acaso, em esquema fatorial: quatro níveis nutricionais e três segmentos do intestino com dez repetições. Foram coletadas amostras dos três segmentos do intestino delgado. As variáveis estudadas foram área e altura das vilosidades intestinais, profundidade de cripta e relação vilo e cripta. No estudo da altura da vilosidade, foi obtida interação significativa para todas as idades estudadas. A profundidade da cripta em codornas de corte aos 21 dias de idade mostrou-se significativa para as variáveis inclusão de treonina na dieta e para a porção do intestino estudada. Na profundidade da cripta aos 42 dias, ficou evidenciada uma interação significativa entre níveis de inclusão de treonina na dieta e porção do intestino delgado. No estudo da relação altura de vilosidade e profundidade de cripta em codornas aos 21 dias de idade, foram encontrados resultados significativos para a interação tratamento e porção do intestino. Aos 21 e aos 42 dias de idade, a suplementação de 1,04 e 0,78%, respectivamente, de treonina digestível na dieta promove melhores resultados para morfometria intestinal.(AU)

The aim of the study was to evaluate intestinal morphology data by including different levels of threonine in the diet of meat quails in the growing phase. The experimental design was completely randomized in a factorial arrangement: four nutritional levels and three intestinal segments with ten repetitions. Samples were collected to obtain morphometry data (villi and crypt) of the three segments of the small intestine (duodenum, jejunum, and ileum). In the study of the intestinal villus height a highly significant interaction was obtained for all ages studied. A crypt depth was evidenced at 42 days with a significant interaction between levels of inclusion of threonine in the diet and small intestine. In the study of the relationship villus height and depth crypt quails at 21 days of age, the results were significant for the interaction treatment (T) and part of the intestine. At 21 and 42 days of age, supplementation of 1.04 and 0.78% respectively of threonine in the diet promotes better outcomes for intestinal morphology.(AU)

Suppression of β-catenin Signaling Pathway in Human Prostate Cancer PC3 Cells by Delphinidin

Delphinidin possesses strong anti-oxidant, anti-inflammatory, and anti-cancer properties. Suppression of the Wnt/β-catenin signaling pathway is a potential strategy for chemoprevention and therapy. As aberrant activation of the β-catenin signaling pathway contributes to prostate cancer progression, we evaluated the effect of delphinidin on this pathway in human PC3 prostate cancer cells. An MTT assay showed that treatment with delphinidin (15-180 μM, 72 hours) resulted in a dose-dependent growth inhibition of cells. Treatment with delphinidin increased the phosphorylation of serine or threonine residues on β-catenin and decreased the levels of cytoplasmic β-catenin. Moreover, treatment with delphinidin inhibited the nuclear translocation of β-catenin and the expression of β-catenin target genes such as cyclin D1, c-myc, Axin-2, and T cell factor-1. Delphinidin also induced the phosphorylation of glycogen synthase kinase 3β and the expression of adenomatous polyposis coli and Axin proteins. Our results indicate that inhibition of cell growth by delphinidin is mediated, at least in part, through modulation of the β-catenin signaling pathway. We suggest that delphinidin is a potent inhibitor of Wnt/β-catenin signaling in prostate cancer cells.

LRRK2 Inhibits FAK Activity by Promoting FERM-mediated Autoinhibition of FAK and Recruiting the Tyrosine Phosphatase, SHP-2

Mutation of leucine-rich repeat kinase 2 (LRRK2) causes an autosomal dominant and late-onset familial Parkinson's disease (PD). Recently, we reported that LRRK2 directly binds to and phosphorylates the threonine 474 (T474)-containing Thr-X-Arg(Lys) (TXR) motif of focal adhesion kinase (FAK), thereby inhibiting the phosphorylation of FAK at tyrosine (Y) 397 residue (pY397-FAK), which is a marker of its activation. Mechanistically, however, it remained unclear how T474-FAK phosphorylation suppressed FAK activation. Here, we report that T474-FAK phosphorylation could inhibit FAK activation via at least two different mechanisms. First, T474 phosphorylation appears to induce a conformational change of FAK, enabling its N-terminal FERM domain to autoinhibit Y397 phosphorylation. This is supported by the observation that the levels of pY397-FAK were increased by deletion of the FERM domain and/or mutation of the FERM domain to prevent its interaction with the kinase domain of FAK. Second, pT474-FAK appears to recruit SHP-2, which is a phosphatase responsible for dephosphorylating pY397-FAK. We found that mutation of T474 into glutamate (T474E-FAK) to mimic phosphorylation induced more strong interaction with SHP-2 than WT-FAK, and that pharmacological inhibition of SHP-2 with NSC-87877 rescued the level of pY397 in HEK293T cells. These results collectively show that LRRK2 suppresses FAK activation through diverse mechanisms that include the promotion of autoinhibition and/or the recruitment of phosphatases, such as SHP-2.

A Pedigree with c.179 Cytosine to Threonine Missense Mutation of SLC12A3 Gene Presenting Gitelman's Syndrome

A 42-year-old man came to the hospital presenting chest discomfort and general weakness. He had come to the hospital with the same symptoms 3 months ago and 12 years prior. His laboratory test showed hypokalemia, hypomagnesemia and hypocalciuria. The arterial blood gas analysis showed hypochloremic metabolic alkalosis. He had an ultrasonography guided renal biopsy, the result was normal at light microscopy and immunofluorescence microscopy. However, a special stain for Na-Cl cotransporter was weakly expressed compared with the control. The patient and his family underwent genetic sequencing about the SLC12A3 gene. He had a homozygous mutation in the 179th nucleotide of Exon 1 on the SLC12A3 gene (p.Thr60Met) and his parents and sisters were diagnosed as carrier state of Gitelman's syndrome (GS). GS is an inherited tubular disorder which presents mild hypokalemia, hypomagnesemia and hypocalciuria. Since the symptoms and laboratory results are not severe, it can go unnoticed by physicians. Herein we present a family with GS, diagnosed by genetic sequencing.

Desempenho e rendimento de carcaça em frangos de corte de um a 46 dias alimentados com dietas contendo diferentes níveis de valina e reduzido teor proteico / Performance and carcass yield of broilers from 1 to 46 days fed diets containing different levels of valine and reduced protein content

Este estudo teve como objetivo avaliar as relações de valina:lisina digestíveis em dietas com teor reduzido de proteína bruta (PB) e os efeitos dessa redução sobre desempenho e rendimento de carcaça em frangos de corte. Foram utilizados 1200 pintos machos seguindo modelo inteiramente ao acaso, com seis tratamentos de seis repetições (exceto controle, com 10 repetições), compostos por 30 aves cada. O tratamento controle (T1) foi formulado conforme os níveis de proteína bruta e aminoácidos (AAs) recomendados por Rostagno et al . (2011), e os demais tratamentos (T2 a T6) tiveram seus níveis de PB reduzidos (4% em relação ao controle) e variaram em função da relação valina:lisina digestíveis, com cinco níveis equidistantes em intervalos de 0,07:1, variando de 0,63:1 e 0,91:1 (dietas até 21 dias) e de 0,64:1 e 0,92:1 (dietas após 21 dias). As seguintes características de desempenho foram avaliadas: ganho de peso, consumo de ração, conversão alimentar, viabilidade criatória e índice de eficiência produtiva. Aos 46 dias de idade, seis animais por repetição foram abatidos para determinação de rendimento de carcaça e de cortes comerciais. As diferentes relações valina:lisina digestíveis não influenciaram o desempenho dos animais (P>0,05) para nenhuma característica avaliada. A redução proteica piorou a conversão alimentar dos animais (P≤0,05) até os 21 dias. Os resultados sugerem que os níveis de valina utilizados não afetam o desempenho dos animais, apenas o rendimento de peito e que, portanto, a redução proteica não é recomendada durante as três primeiras semanas de criação.

This study aimed to evaluate valine:lysine ratios in diets with reduced content of crude protein and the effects of this reduction on the performance of broiler chickens. 1200 male chicks were used following a complete randomized design with six replicates of six treatments (except control, with 10 replicates), each one with 30 chicks. The control treatment (T1) was formulated following levels of crude protein (CP) and the amino acids (AAs) recommended by Rostagno et al. (2011), and the other treatments (T2 to T6) had reduced levels of CP (4 % compared to control) and varied in proportion valine:lysine, with 5 levels at equidistant intervals 0.07:1 ranging from 0.63:1 to 0.91:1 (up to 21 days) and from 0.64:1 to 0.92:1 (after 21 days). The performance characteristics measured were: weight gain, feed intake, feed conversion, viability and productive efficiency index. At 46 days six animals per replicate were slaughtered for evaluation of carcass and commercial cuts. The different valine:lysine ratios did not affect animal performance (P>0.05). Reducing protein impaired feed conversion (P≤.05) up to 21 days. The results suggest that levels of valine used did not affect the broilers' performance, however, breast meat yield and reduced protein are not recommended during the first three weeks.

Threonine 32 (Thr32) of FoxO3 is critical for TGF-β-induced apoptosis via Bim in hepatocarcinoma cells

Transforming growth factor-β (TGF-β) exerts apoptotic effects on various types of malignant cells, including liver cancer cells. However, the precise mechanisms by which TGF-β induces apoptosis remain poorly known. In the present study, we have showed that threonine 32 (Thr32) residue of FoxO3 is critical for TGF-β to induce apoptosis via Bim in hepatocarcinoma Hep3B cells. Our data demonstrated that TGF-β induced FoxO3 activation through specific de-phosphorylation at Thr32. TGF-β-activated FoxO3 cooperated with Smad2/3 to mediate Bim up-regulation and apoptosis. FoxO3 (de)phosphorylation at Thr32 was regulated by casein kinase I-ε (CKI-ε). CKI inhibition by small molecule D4476 could abrogate TGF-β-induced FoxO/Smad activation, reverse Bim up-regulation, and block the sequential apoptosis. More importantly, the deregulated levels of CKI-ε and p32FoxO3 were found in human malignant liver tissues. Taken together, our findings suggest that there might be a CKI-FoxO/Smad-Bim engine in which Thr32 of FoxO3 is pivotal for TGF-β-induced apoptosis, making it a potential therapeutic target for liver cancer treatment.

Transcriptional Profiling of an Attenuated Salmonella Typhimurium ptsI Mutant Strain Under Low-oxygen Conditions using Microarray Analysis

Salmonella causes a wide variety of diseases ranging from mild diarrhea to severe systemic infections, such as like typhoid fever, in multiple organisms, ranging from mice to humans. A lack of ptsI, which encodes the first component of phosphoenolpyruvate (PEP) : carbohydrate phosphotransferase system (PTS), is known to cause Salmonella Typhimurium attenuation; however, the mechanisms behind this have not yet been elucidated. In this study, a DNA microarray was performed to determine why the virulence of ptsI mutants is attenuated under low-oxygen conditions in which the ptsI expression is enhanced. Of 106 down-regulated genes, the most repressed were pdu and tdc genes, which are required for propanediol utilization and threonine and serine metabolism, respectively. In addition, half the flagellar genes were down-regulated in the ptsI mutant strain. Because pdu genes are induced during infection and Tdc products and flagella-mediated motility are necessary for the invasion of S. Typhimurium, the invasive ability of ptsI mutants was examined. We found that ptsI mutation reduced the ability of S. Typhimurium to invade into host cells, suggesting that reduced expression of the pdu, tdc, and flagellar genes is involved in the attenuation of ptsI mutants.

Genes Involved in the Biosynthesis and Transport of Acinetobactin in Acinetobacter baumannii

Pathogenic bacteria survive in iron-limited host environments by using several iron acquisition mechanisms. Acinetobacter baumannii, causing serious infections in compromised patients, produces an iron-chelating molecule, called acinetobactin, which is composed of equimolar quantities of 2,3-dihydroxybenzoic acid (DHBA), L-threonine, and N-hydroxyhistamine, to compete with host cells for iron. Genes that are involved in the production and transport of acinetobactin are clustered within the genome of A. baumannii. A recent study showed that entA, located outside of the acinetobactin gene cluster, plays important roles in the biosynthesis of the acinetobactin precursor DHBA and in bacterial pathogenesis. Therefore, understanding the genes that are associated with the biosynthesis and transport of acinetobactin in the bacterial genome is required. This review is intended to provide a general overview of the genes in the genome of A. baumannii that are required for acinetobactin biosynthesis and transport.