Nitrate is the main form of inorganic nitrogen that crop absorbs, and nitrate transporter 2 (NRT2) is a high affinity transporter using nitrate as a specific substrate. When the available nitrate is limited, the high affinity transport systems are activated and play an important role in the process of nitrate absorption and transport. Most NRT2 cannot transport nitrates alone and require the assistance of a helper protein belonging to nitrate assimilation related family (NAR2) to complete the absorption or transport of nitrates. Crop nitrogen utilization efficiency is affected by environmental conditions, and there are differences between varieties, so it is of great significance to develop varieties with high nitrogen utilization efficiency. Sorghum bicolor has high stress tolerance and is more efficient in soil nitrogen uptake and utilization. The S. bicolor genome database was scanned to systematically analyze the gene structure, chromosomal localization, physicochemical properties, secondary structure and transmembrane domain, signal peptide and subcellular localization, promoter region cis-acting elements, phylogenetic evolution, single nucleotide polymorphism (SNP) recognition and annotation, and selection pressure of the gene family members. Through bioinformatics analysis, 5 NRT2 gene members (designated as SbNRT2-1a, SbNRT2-1b, SbNRT2-2, SbNRT2-3, and SbNRT2-4) and 2 NAR2 gene members (designated as SbNRT3-1 and SbNRT3-2) were identified, the number of which was less than that of foxtail millet. SbNRT2/3 were distributed on 3 chromosomes, and could be divided into four subfamilies. The genetic structure of the same subfamilies was highly similar. The average value of SbNRT2/3 hydrophilicity was positive, indicating that they were all hydrophobic proteins, whereas α-helix and random coil accounted for more than 70% of the total secondary structure. Subcellular localization occurred on plasma membrane, where SbNRT2 proteins did not contain signal peptides, but SbNRT3 proteins contained signal peptides. Further analysis revealed that the number of transmembrane domains of the SbNRT2s family members was greater than 10, while that of the SbNRT3s were 2. There was a close collinearity between NRT2/3s of S. bicolor and Zea mays. Protein domains analysis showed the presence of MFS_1 and NAR2 protein domains, which supported executing high affinity nitrate transport. Phylogenetic tree analysis showed that SbNRT2/3 were more closely related to those of Z. mays and Setaria italic. Analysis of gene promoter cis-acting elements indicated that the promoter region of SbNRT2/3 had several plant hormones and stress response elements, which might respond to growth and environmental cues. Gene expression heat map showed that SbNRT2-3 and SbNRT3-1 were induced by nitrate in the root and stem, respectively, and SbNRT2-4 and SbNRT2-3 were induced by low nitrogen in the root and stem. Non-synonymous SNP variants were found in SbNRT2-4 and SbNRT2-1a. Selection pressure analysis showed that the SbNRT2/3 were subject to purification and selection during evolution. The expression of SbNRT2/3 gene and the effect of aphid infection were consistent with the expression analysis results of genes in different tissues, and SbNRT2-1b and SbNRT3-1 were significantly expressed in the roots of aphid lines 5-27sug, and the expression levels of SbNRT2-3, SbNRT2-4 and SbNRT3-2 were significantly reduced in sorghum aphid infested leaves. Overall, genome-wide identification, expression and DNA variation analysis of NRT2/3 gene family of Sorghum bicolor provided a basis for elucidating the high efficiency of sorghum in nitrogen utilization.
Nitrate Transporters ; Nitrates/metabolism* ; Sorghum/metabolism* ; Anion Transport Proteins/metabolism* ; Phylogeny ; Protein Sorting Signals/genetics* ; Nitrogen/metabolism* ; DNA ; Gene Expression Regulation, Plant ; Plant Proteins/metabolism*

Sweet potato is an important food crop that can also be used as an industrial raw material. Sucrose is the main form of long-distance carbohydrate transport in plants, and sucrose transporter (SUT) regulates the transmembrane transport and distribution of sucrose during plant growth and metabolism. Moreover, SUT plays a key role in phloem mediated source-to-sink sucrose transport and physiological activities, supplying sucrose for the sink tissues. In this study, the full-length cDNA sequences of IbSUT₆₂₇₈₈ and IbSUT₈₁₆₁₆ were obtained by rapid amplification of cDNA ends (RACE) cloning according to the transcripts of the two SUT coding genes which were differentially expressed in sweet potato storage roots with different starch properties. Phylogenetic analysis was performed to clarify the classification of IbSUT₆₂₇₈₈ and IbSUT₈₁₆₁₆. The subcellular localization of IbSUT₆₂₇₈₈ and IbSUT₈₁₆₁₆ was determined by transient expression in Nicotiana benthamiana. The function of IbSUT₆₂₇₈₈ and IbSUT₈₁₆₁₆ in sucrose and hexose absorption and transport was identified using yeast functional complementarity system. The expression pattern of IbSUT₆₂₇₈₈ and IbSUT₈₁₆₁₆ in sweet potato organs were analyzed by real-time fluorescence quantitative PCR (RT-qPCR). Arabidopsis plants heterologous expressing IbSUT₆₂₇₈₈ and IbSUT₈₁₆₁₆ genes were obtained using floral dip method. The differences in starch and sugar contents between transgenic and wild-type Arabidopsis were compared. The results showed IbSUT₆₂₇₈₈ and IbSUT₈₁₆₁₆ encoded SUT proteins with a length of 505 and 521 amino acids, respectively, and both proteins belonged to the SUT1 subfamily. IbSUT₆₂₇₈₈ and IbSUT₈₁₆₁₆ were located in the cell membrane and were able to transport sucrose, glucose and fructose in the yeast system. In addition, IbSUT₆₂₇₈₈ was also able to transport mannose. The expression of IbSUT₆₂₇₈₈ was higher in leaves, lateral branches and main stems, and the expression of IbSUT₈₁₆₁₆ was higher in lateral branches, stems and storage roots. After IbSUT₆₂₇₈₈ and IbSUT₈₁₆₁₆ were heterologously expressed in Arabidopsis, the plants grew normally, but the biomass increased. The heterologous expression of IbSUT₆₂₇₈₈ increased the soluble sugar content, leaf size and 1 000-seed weight of Arabidopsis plants. Heterologous expression of IbSUT₈₁₆₁₆ increased starch accumulation in leaves and root tips and 1 000-seed weight of seeds, but decreased soluble sugar content. The results obtained in this study showed that IbSUT₆₂₇₈₈ and IbSUT₈₁₆₁₆ might be important genes regulating sucrose and sugar content traits in sweet potato. They might carry out physiological functions on cell membrane, such as transmembrane transport of sucrose, sucrose into and out of sink tissue, as well as transport and unloading of sucrose into phloem. The changes in traits result from their heterologous expression in Arabidopsis indicates their potential in improving the yield of other plants or crops. The results obtained in this study provide important information for revealing the functions of IbSUT₆₂₇₈₈ and IbSUT₈₁₆₁₆ in starch and glucose metabolism and formation mechanism of important quality traits in sweet potato.
Ipomoea batatas/metabolism* ; Arabidopsis/metabolism* ; Sucrose/metabolism* ; Saccharomyces cerevisiae/metabolism* ; DNA, Complementary ; Phylogeny ; Plants, Genetically Modified/genetics* ; Membrane Transport Proteins/metabolism* ; Starch/metabolism* ; Plant Proteins/metabolism* ; Gene Expression Regulation, Plant

Objective To study the pathological types,expression of mismatch repair protein,human epidermal growth factor receptor 2(HER2),and Pan-TRK,and Epstein-Barr virus(EBV)infection in patients with colorectal cancer resected in Tibet. Methods A total of 79 patients with colorectal cancer resected in Tibet Autonomous Region People's Hospital from December 2013 to July 2021 were enrolled in this study.The clinical and pathological data of the patients were collected.The expression of mismatch repair protein,HER2,and Pan-TRK was detected by immunohistochemical(IHC)staining,and detection of HER2 gene by fluorescence in situ hybridization(FISH)in the patients with HER2 IHC results of 2+ or above.EBV was detected by in situ hybridization with EBV-encoded small RNA. Results A total of 79 colorectal cancer patients were included in this study,with the male-to-female ratio of 1.26:1 and the mean age of(57.06±12.74)years(24-83 years).Among them,4 patients received preoperative neoadjuvant therapy.Colonic cancer and rectal cancer occurred in 57(57/79,72.15%,including 31 and 26 in the right colon and left colon,respectively)and 22(22/79,27.85%)patients,respectively.The maximum diameter of tumor varied within the range of 1-20 cm,with the mean of(6.61±3.33)cm.Among the 79 colorectal cancer patients,75(75/79,94.94%)patients showed adenocarcinoma.Lymph node metastasis occurred in 12(12/21,57.14%)out of the 21 patients with severe tumor budding,13(13/23,56.52%)out of the 23 patients with moderate tumor budding,and 2(2/31,6.45%)out of the 31 patients with mild tumor budding,respectively.The lymph node metastasis rate showed differences between the patients with severe/moderate tumor budding and the patients with mild tumor budding(all P<0.001).The IHC staining showed that mismatch repair protein was negative in 10(10/65,15.38%)patients,including 5 patients with both MSH2 and MSH6 negative,4 patients with both MLH1 and PMS2 negative,and 1 patient with MSH6 negative.Pan-TRK was negative in 65 patients.The IHC results of HER2 showed 0 or 1+ in 60 patients and 2+ in 5 patients.FISH showed no positive signal in the 5 patients with HER2 IHC results of 2+.The detection with EBV-encoded small RNA showed positive result in 1(1/65,1.54%)patient. Conclusions Non-specific adenocarcinoma of the right colon is the most common in the patients with colorectal cancer resected in Tibet,and 15% of the patients showed mismatch repair protein defects.EBV-associated colorectal carcer is rare,Pan-TRK expression and HER2 gene amplification are seldom.The colorectal cancer patients with moderate and severe tumor budding are more likely to have lymph node metastasis.
Adult ; Aged ; Female ; Humans ; Male ; Middle Aged ; Adenocarcinoma ; Biomarkers, Tumor/genetics* ; Colorectal Neoplasms/pathology* ; DNA Mismatch Repair ; DNA-Binding Proteins/genetics* ; Epstein-Barr Virus Infections/diagnosis* ; Herpesvirus 4, Human/metabolism* ; In Situ Hybridization, Fluorescence ; Lymphatic Metastasis ; Tibet ; Young Adult ; Aged, 80 and over

OBJECTIVE: To carry out preimplantation genetic testing for monogenic/single gene disorders (PGT-M) for a Chinese family affected with Molybdenum co-factor deficiency due to pathogenic variant of MOCS2 gene. METHODS: A family with molybdenum co-factor deficiency who attended to the Maternal and Child Health Care Hospital of Guangxi Zhuang Autonomous Region in April 2020 was selected as the research subject. Trophoblast cells were biopsied from blastocysts fertilized by intracytoplasmic sperm injection. Embryos carrying the MOCS2 gene variant and chromosome copy number variation (CNV) of more than 4 Mb were detected by single-cell whole genome amplification, high-throughput sequencing and single nucleotide polymorphism typing. Embryos without or carrying the heterozygous variant and without abnormal chromosome CNV were transplanted. During mid-pregnancy, amniotic fluid sample was collected for prenatal diagnosis to verify the results of PGT-M. RESULTS: Eleven oocytes were obtained, among which three blastocysts were formed through culturing. Results of genetic testing suggested that one embryo was heterozygous for the maternally derived MOCS2 gene variant and without chromosomal CNV. Following embryo transfer, intrauterine singleton pregnancy was attained. Prenatal diagnosis by amniocentesis at 18 weeks of gestation revealed that the MOCS2 gene variant and chromosomal analysis results were both consistent with that of PGT-M, and a healthy male infant was born at 37⁺⁵ weeks of gestation. CONCLUSION PGT-M has helped the couple carrying the MOCS2 gene variant to have a healthy offspring, and may become an important method for couples carrying other pathogenic genetic variants.
Female ; Humans ; Pregnancy ; Aneuploidy ; China ; DNA Copy Number Variations ; Genetic Testing/methods* ; Preimplantation Diagnosis/methods* ; Metal Metabolism, Inborn Errors/genetics*

MAGED4B belongs to the melanoma-associated antigen family; originally found in melanoma, it is expressed in various types of cancer, and is especially enriched in glioblastoma. However, the functional role and molecular mechanisms of MAGED4B in glioma are still unclear. In this study, we found that the MAGED4B level was higher in glioma tissue than that in non-cancer tissue, and the level was positively correlated with glioma grade, tumor diameter, Ki-67 level, and patient age. The patients with higher levels had a worse prognosis than those with lower MAGED4B levels. In glioma cells, MAGED4B overexpression promoted proliferation, invasion, and migration, as well as decreasing apoptosis and the chemosensitivity to cisplatin and temozolomide. On the contrary, MAGED4B knockdown in glioma cells inhibited proliferation, invasion, and migration, as well as increasing apoptosis and the chemosensitivity to cisplatin and temozolomide. MAGED4B knockdown also inhibited the growth of gliomas implanted into the rat brain. The interaction between MAGED4B and tripartite motif-containing 27 (TRIM27) in glioma cells was detected by co-immunoprecipitation assay, which showed that MAGED4B was co-localized with TRIM27. In addition, MAGED4B overexpression down-regulated the TRIM27 protein level, and this was blocked by carbobenzoxyl-L-leucyl-L-leucyl-L-leucine (MG132), an inhibitor of the proteasome. On the contrary, MAGED4B knockdown up-regulated the TRIM27 level. Furthermore, MAGED4B overexpression increased TRIM27 ubiquitination in the presence of MG132. Accordingly, MAGED4B down-regulated the protein levels of genes downstream of ubiquitin-specific protease 7 (USP7) involved in the tumor necrosis factor-alpha (TNF-α)-induced apoptotic pathway. These findings indicate that MAGED4B promotes glioma growth via a TRIM27/USP7/receptor-interacting serine/threonine-protein kinase 1 (RIP1)-dependent TNF-α-induced apoptotic pathway, which suggests that MAGED4B is a potential target for glioma diagnosis and treatment.
Humans ; Tumor Necrosis Factor-alpha ; DNA-Binding Proteins/metabolism* ; Ubiquitin-Specific Peptidase 7 ; Cisplatin ; Temozolomide ; Transcription Factors ; Glioma ; Cell Proliferation ; Melanoma ; Cell Line, Tumor ; Apoptosis ; Nuclear Proteins/genetics*

Pregnancy ; Female ; Humans ; Fetal Blood/metabolism* ; DNA Methylation/genetics* ; Epigenesis, Genetic ; Placenta/metabolism* ; Exercise Therapy

Autism spectrum disorder (ASD) is a highly heritable neurodevelopmental disorder characterized by deficits in social interactions and repetitive behaviors. Although hundreds of ASD risk genes, implicated in synaptic formation and transcriptional regulation, have been identified through human genetic studies, the East Asian ASD cohorts are still under-represented in genome-wide genetic studies. Here, we applied whole-exome sequencing to 369 ASD trios including probands and unaffected parents of Chinese origin. Using a joint-calling analytical pipeline based on GATK toolkits, we identified numerous de novo mutations including 55 high-impact variants and 165 moderate-impact variants, as well as de novo copy number variations containing known ASD-related genes. Importantly, combined with single-cell sequencing data from the developing human brain, we found that the expression of genes with de novo mutations was specifically enriched in the pre-, post-central gyrus (PRC, PC) and banks of the superior temporal (BST) regions in the human brain. By further analyzing the brain imaging data with ASD and healthy controls, we found that the gray volume of the right BST in ASD patients was significantly decreased compared to healthy controls, suggesting the potential structural deficits associated with ASD. Finally, we found a decrease in the seed-based functional connectivity between BST/PC/PRC and sensory areas, the insula, as well as the frontal lobes in ASD patients. This work indicated that combinatorial analysis with genome-wide screening, single-cell sequencing, and brain imaging data reveal the brain regions contributing to the etiology of ASD.
Humans ; Autism Spectrum Disorder/metabolism* ; Autistic Disorder ; Exome Sequencing ; DNA Copy Number Variations ; East Asian People ; Brain/metabolism* ; Mutation/genetics* ; Genetic Predisposition to Disease/genetics*

Objective: To explore the concordance and causes of different mismatch repair (MMR) and microsatellite instability (MSI) detection results in endometrial carcinoma (EC) molecular typing. Methods: A total of 214 EC patients diagnosed from January 2021 to April 2023 were selected at the Department of Pathology, Peking University Third Hospital. The immunohistochemistry (IHC) results of MMR protein were reviewed. Tumor specific somatic mutations, MMR germline mutations, microsatellite scores and tumor mutation burden (TMB) were detected by next-generation sequencing (NGS) with multi-gene panel. Methylation-specific PCR was used to detect the methylation status of MLH1 gene promoter in cases with deficient MLH1 protein expression. In cases with discrepant results between MMR-IHC and MSI-NGS, the MSI status was detected again by PCR (MSI-PCR), and the molecular typing was determined by combining the results of TMB and MLH1 gene promoter methylation. Results: (1) In this study, there were 22 cases of POLE gene mutation subtype, 55 cases of mismatch repair deficient (MMR-d) subtype, 29 cases of p53 abnormal subtype, and 108 cases of no specific molecular profile (NSMP). The median age at diagnosis of MMR-d subtype (54 years old) and the proportion of aggressive histological types (40.0%, 22/55) were higher than those of NSMP subtype [50 years old and 12.0% (13/108) respectively; all P<0.05]. (2) Among 214 patients, MMR-IHC test showed that 153 patients were mismatch repair proficient (MMR-p), 49 patients were MMR-d, and 12 patients were difficult to evaluate directly. MSI-NGS showed that 164 patients were microsatellite stable (MSS; equal to MMR-p), 48 patients were high microsatellite instability (MSI-H; equal to MMR-d), and 2 patients had no MSI-NGS results because the effective sequencing depth did not meet the quality control. The overall concordance between MMR-IHC and MSI-NGS was 94.3% (200/212). All the 12 discrepant cases were MMR-d or subclonal loss of MMR protein by IHC, but MSS by NGS. Among them, 10 cases were loss or subclonal loss of MLH1 and (or) PMS2 protein. Three discrepant cases were classified as POLE gene mutation subtype. In the remaining 9 cases, 5 cases and 3 cases were confirmed as MSI-H and low microsatellite instability (MSI-L) respectively by MSI-PCR, 6 cases were detected as MLH1 gene promoter methylation and 7 cases demonstrated high TMB (>10 mutations/Mb). These 9 cases were classified as MMR-d EC. (3) Lynch syndrome was diagnosed in 27.3% (15/55) of all 55 MMR-d EC cases, and the TMB of EC with MSH2 and (or) MSH6 protein loss or associated with Lynch syndrome [(71.0±26.2) and (71.5±20.1) mutations/Mb respectively] were significantly higher than those of EC with MLH1 and (or) PMS2 loss or sporadic MMR-d EC [(38.2±19.1) and (41.9±24.3) mutations/Mb respectively, all P<0.01]. The top 10 most frequently mutated genes in MMR-d EC were PTEN (85.5%, 47/55), ARID1A (80.0%, 44/55), PIK3CA (69.1%, 38/55), KMT2B (60.0%, 33/55), CTCF (45.5%, 25/55), RNF43 (40.0%, 22/55), KRAS (36.4%, 20/55), CREBBP (34.5%, 19/55), LRP1B (32.7%, 18/55) and BRCA2 (32.7%, 18/55). Concurrent PTEN, ARID1A and PIK3CA gene mutations were found in 50.9% (28/55) of MMR-d EC patients. Conclusions: The concordance of MMR-IHC and MSI-NGS in EC is relatively high.The discordance in a few MMR-d EC are mostly found in cases with MLH1 and (or) PMS2 protein loss or MMR protein subclonal staining caused by MLH1 gene promoter hypermethylation. In order to provide accurate molecular typing for EC patients, MLH1 gene methylation, MSI-PCR, MMR gene germline mutation and TMB should be combined to comprehensively evaluate MMR and MSI status.
Female ; Humans ; Middle Aged ; Class I Phosphatidylinositol 3-Kinases/metabolism* ; Colorectal Neoplasms, Hereditary Nonpolyposis/diagnosis* ; DNA Mismatch Repair/genetics* ; Endometrial Neoplasms/pathology* ; Microsatellite Instability ; Mismatch Repair Endonuclease PMS2/genetics* ; Molecular Typing

OBJECTIVE: To explore the clinical features and genetic basis for a child with 3-methylglutaconic aciduria type VII. METHODS: A child who was diagnosed at the Gansu Provincial Maternity and Child Health Care Hospital on August 9, 2019 was selected as the study subject. Clinical data of the child, including urine gas chromatography and mass spectrometry, were collected. The child and her parents were subjected to whole exome sequencing. RESULTS: The child, a female neonate, had presented mainly with intermittent skin cyanosis, convulsions, hypomagnesemia, apnea, neutropenia after birth. Her urine 3-methylpentenedioic acid has increased to 17.53 μmol/L. DNA sequencing revealed that she has harbored compound heterozygous variants of the CLPB gene, namely c.1016delT (p.L339Rfs*5) and c.1087A>G (p.R363G), which were respectively inherited from her mother and father. Both variants were unreported previously. Based on the standards from the American College of Medical Genetics and Genomics (ACMG), the variants were respectively predicted to be pathogenic and likely pathogenic. CONCLUSION The child was diagnosed with 3-methylglutenedioic aciduria type VII. Discovery of the c.1016delT and c.1087A>G variants has enriched the mutational spectrum of the CLPB gene.
Female ; Humans ; Infant, Newborn ; Pregnancy ; Base Sequence ; Metabolism, Inborn Errors/diagnosis* ; Mutation ; Neutropenia/genetics* ; Sequence Analysis, DNA

DNA sensor, a kind of pattern recognition receptor (PRR), is widely expressed in innate immune cells. It activates the inflammatory signaling pathways and triggers an innate immune response by recognizing the pathogens or DNA in abnormal host cells. DNA-dependent activator of IFN-regulatory factors (DAI) is the first cytoplasmic DNA receptor discovered, which plays an important role in regulating the innate immune responses characterized by induction of interferon and programmed cell death. The article summarizes the molecular characteristics of DAI, its downstream signaling pathways, and its role and mechanism in anti-infective immunity, tumor immunity and inflammatory diseases. It also makes a preliminary exploration of the correlation between DAI and transplantation immunology, and provides a new target for the therapy of various immune diseases.
DNA/metabolism* ; Receptors, Pattern Recognition ; Immunity, Innate ; Signal Transduction/genetics* ; DNA-Binding Proteins/genetics*