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Mutations in any one of the four subunits (ɛ4, β4, μ4 and σ4) comprising the adaptor protein Complex 4 results in a complex form of hereditary spastic paraplegia, often termed adaptor protein Complex 4 deficiency syndrome. Deficits in adaptor protein Complex 4 complex function have been shown to disrupt intracellular trafficking, resulting in a broad phenotypic spectrum encompassing severe intellectual disability and progressive spastic paraplegia of the lower limbs in patients. Here we report the presence of neuropathological hallmarks of adaptor protein Complex 4 deficiency syndrome in a clustered regularly interspaced short palindromic repeats-mediated Ap4b1 -knockout mouse model. Mice lacking the β4 subunit, and therefore lacking functional adaptor protein Complex 4, have a thin corpus callosum, enlarged lateral ventricles, motor co-ordination deficits, hyperactivity, a hindlimb clasping phenotype associated with neurodegeneration, and an abnormal gait. Analysis of autophagy-related protein 9A (a known cargo of the adaptor protein Complex 4 in these mice shows both upregulation of autophagy-related protein 9A protein levels across multiple tissues, as well as a striking mislocalization of autophagy-related protein 9A from a generalized cytoplasmic distribution to a marked accumulation in the trans -Golgi network within cells. This mislocalization is present in mature animals but is also in E15.5 embryonic cortical neurons. Histological examination of brain regions also shows an accumulation of calbindin-positive spheroid aggregates in the deep cerebellar nuclei of adaptor protein Complex 4-deficient mice, at the site of Purkinje cell axonal projections. Taken together, these findings show a definitive link between loss-of-function mutations in murine Ap4b1 and the development of symptoms consistent with adaptor protein Complex 4 deficiency disease in humans. Furthermore, this study provides strong evidence for the use of this model for further research into the aetiology of adaptor protein Complex 4 deficiency in humans, as well as its use for the development and testing of new therapeutic modalities., (© The Author(s) 2023. Published by Oxford University Press on behalf of the Guarantors of Brain.)

Context: A few papillary thyroid microcarcinomas (PTMCs) may have skip metastasis (SLNM), but the risk factors remain controversial and the prognosis is unclear., Objectives: To investigate the incidence, lymph node metastasis (LNM) patterns, risk factors, and prognosis of SLNM in PTMCs., Methods: We reviewed the medical records of PTMC patients who underwent thyroid surgery in our institution. Analyses of risk factors were performed for SLNM. Recurrence-free survival (RFS) of SLNM, central lymph node metastasis (CLNM), and continuous metastasis (CLNM and lateral lymph node metastasis [CLNM + LLNM]) were compared after propensity score matching (PSM)., Results: SLNM was detected in 1.7% (50/3923) and frequently involved level III (66.7%). Compared with CLNM + LLNM, SLNM had more LNM at a single level (P < 0.01) and less LNM at 2 levels (P < 0.05). A tumor size of 0.5 to 1 cm (odds ratio [OR], 2.26; 95% CI, 1.27-4.00) and location in the upper pole (OR, 3.30; 95% CI, 2.02-5.40) were independent risk factors for SLNM. A total of 910 (23.2%) PTMCs with LNM were included in the prognostic analysis. At a median follow-up of 60 months, the RFS of SLNM did not differ from that of CLNM (P = 0.10) but was significantly higher than that of CLNM + LLNM (P < 0.01) after using PSM., Conclusions: When the tumor size is 0.5 to 1 cm or its location is in the upper pole, we must remain vigilant to SLNM in PTMC. Because its prognosis is comparable to that of only CLNM and better than that of CLNM + LLNM, less intensive treatment should be considered., (© The Author(s) 2022. Published by Oxford University Press on behalf of the Endocrine Society. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Background: The National Comprehensive Cancer Network's Rectal Cancer Guideline Panel recommends American Joint Committee of Cancer and College of American Pathologists (AJCC/CAP) tumor regression grading (TRG) system to evaluate pathologic response to neoadjuvant chemoradiotherapy for locally advanced rectal cancer (LARC). Yet, the clinical significance of the AJCC/CAP TRG system has not been fully defined., Materials and Methods: This was a multicenter, retrospectively recruited, and prospectively maintained cohort study. Patients with LARC from one institution formed the discovery set, and cases from external independent institutions formed a validation set to verify the findings from discovery set. Overall survival (OS), disease-free survival (DFS), local recurrence-free survival (LRFS), and distant metastasis-free survival (DMFS) were assessed by Kaplan-Meier analysis, log-rank test, and Cox regression model., Results: The discovery set (940 cases) found, and the validation set (2,156 cases) further confirmed, that inferior AJCC/CAP TRG categories were closely /ccorrelated with unfavorable survival (OS, DFS, LRFS, and DMFS) and higher risk of disease progression (death, accumulative relapse, local recurrence, and distant metastasis) (all p < .05). Significantly, pairwise comparison revealed that any two of four TRG categories had the distinguished survival and risk of disease progression. After propensity score matching, AJCC/CAP TRG0 category (pathological complete response) patients treated with or without adjuvant chemotherapy displayed similar survival of OS, DFS, LRFS, and DMFS (all p > .05). For AJCC/CAP TRG1-3 cases, adjuvant chemotherapy treatment significantly improved 3-year OS (90.2% vs. 84.6%, p < .001). Multivariate analysis demonstrated the AJCC/CAP TRG system was an independent prognostic surrogate., Conclusion: AJCC/CAP TRG system, an accurate prognostic surrogate, appears ideal for further strategizing adjuvant chemotherapy for LARC., Implications for Practice: The National Comprehensive Cancer Network recommends the American Joint Committee of Cancer and College of American Pathologists (AJCC/CAP) tumor regression grading (TRG) four-category system to evaluate the pathologic response to neoadjuvant treatment for patients with locally advanced rectal cancer; however, the clinical significance of the AJCC/CAP TRG system has not yet been clearly addressed. This study found, for the first time, that any two of four AJCC/CAP TRG categories had the distinguished long-term survival outcome. Importantly, adjuvant chemotherapy may improve the 3-year overall survival for AJCC/CAP TRG1-3 category patients but not for AJCC/CAP TRG0 category patients. Thus, AJCC/CAP TRG system, an accurate surrogate of long-term survival outcome, is useful in guiding adjuvant chemotherapy management for rectal cancer., (© 2021 AlphaMed Press.)

The spikelet is an inflorescence structure unique to grasses. The molecular mechanisms underlying spikelet development and evolution are unclear. In this study, we characterized three allelic recessive mutants in rice ( Oryza sativa ): nonstop glumes 1-1 ( nsg1-1 ), nsg1-2 , and nsg1-3 In these mutants, organs such as the rudimentary glume, sterile lemma, palea, lodicule, and filament were elongated and/or widened, or transformed into lemma- and/or marginal region of the palea-like organs. NSG1 encoded a member of the C2H2 zinc finger protein family and was expressed mainly in the organ primordia of the spikelet. In the nsg1-1 mutant spikelet, LHS1 DL , and MFO1 were ectopically expressed in two or more organs, including the rudimentary glume, sterile lemma, palea, lodicule, and stamen, whereas G1 was downregulated in the rudimentary glume and sterile lemma. Furthermore, the NSG1 protein was able to bind to regulatory regions of LHS1 and then recruit the corepressor TOPLESS-RELATED PROTEIN to repress expression by downregulating histone acetylation levels of the chromatin. The results suggest that NSG1 plays a pivotal role in maintaining organ identities in the spikelet by repressing the expression of LHS1 , DL , and MFO1., (© 2020 American Society of Plant Biologists. All rights reserved.)

Phosphate (Pi), as the main form of phosphorus that can be absorbed by plants, is one of the most limiting macro-nutrients for plants. However, the mechanism for maintaining Pi homeostasis in rice (Oryza sativa) is still not well understood. We identified a Pi-starvation-induced E3 ligase (OsPIE1) in rice. Using an in vitro self-ubiquitination assay, we determined the E3 ligase activities of OsPIE1. Using GUS staining and GFP detection, we analyzed tissue expression patterns of OsPIE1 and the subcellular localization of its encoded protein. The function of OsPIE1 in Pi homeostasis was analyzed using OsPIE1 overexpressors and ospie1 mutants. OsPIE1 was localized to the nucleus, and expressed in epidermis, exodermis and sclerenchyma layers of primary root. Under Pi-sufficient condition, overexpression of OsPIE1 upregulated the expression of OsPT2, OsPT3, OsPT10 and OsPAP21b, resulting in Pi accumulation and acid phosphatases (APases) induction in roots. OsSPX2 was strongly suppressed in OsPIE1 overexpressors. Further comparative transcriptome analysis, tissue expression patterns and genetic interaction analysis indicated that the enhancing of Pi accumulation and APase activities upon overexpression of OsPIE1 was (at least in part) caused by repression of OsSPX2. These results indicate that OsPIE1 plays an important role in maintaining Pi homeostasis in rice.

Evolutionary mechanisms of substrate specificities of enzyme families remain poorly understood. Plant SABATH methyltransferases catalyze methylation of the carboxyl group of various low molecular weight metabolites. Investigation of the functional diversification of the SABATH family in plants could shed light on the evolution of substrate specificities in this enzyme family. Previous studies identified 28 SABATH genes from the Populus trichocarpa genome. In this study, we re-annotated the Populus SABATH gene family, and performed molecular evolution, gene expression and biochemical analyses of this large gene family. Twenty-eight Populus SABATH genes were divided into three classes with distinct divergences in their gene structure, expression responses to abiotic stressors and enzymatic properties of encoded proteins. Populus class I SABATH proteins converted IAA to methyl-IAA, class II SABATH proteins converted benzoic acid (BA) and salicylic acid (SA) to methyl-BA and methyl-SA, while class III SABATH proteins converted farnesoic acid (FA) to methyl-FA. For Populus class II SABATH proteins, both forward and reverse mutagenesis studies showed that a single amino acid switch between PtSABATH4 and PtSABATH24 resulted in substrate switch. Our findings provide new insights into the evolution of substrate specificities of enzyme families.

Global climate change could have profound effects on belowground microbial communities and subsequently affect soil biogeochemical processes. The interactive effects of multiple co-occurring climate change factors on microbially mediated processes are not well understood. A four-factorial field experiment with elevated CO2, watering, nitrogen (N) addition and night warming was conducted in a temperate steppe of northern China. Real-time polymerase chain reaction and terminal-restriction fragment length polymorphism, combined with clone library techniques, were applied to examine the effects of those climate change factors on N-related microbial abundance and community composition. Only the abundance of ammonia-oxidizing bacteria significantly increased by nitrogen addition and decreased by watering. The interactions of watering × warming on the bacterial amoA community and warming × nitrogen addition on the nosZ community were found. Redundancy analysis indicated that the ammonia-oxidizing archaeal community was affected by total N and total carbon, while the community of bacterial amoA and nosZ were significantly affected by soil pH. According to a structural equation modeling analysis, climate change influenced net primary production indirectly by altering microbial abundance and activities. These results indicated that microbial responses to the combination of chronic global change tend to be smaller than expected from single-factor global change manipulations., (© FEMS 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Whole-genome duplication (WGD), or polyploidy, is a major force in plant genome evolution. A duplicate of all genes is present in the genome immediately following a WGD event. However, the evolutionary mechanisms responsible for the loss of, or retention and subsequent functional divergence of polyploidy-derived duplicates remain largely unknown. In this study we reconstructed the evolutionary history of the glutathione S-transferase (GST) gene family from the soybean genome, and identified 72 GST duplicated gene pairs formed by a recent Glycine-specific WGD event occurring approximately 13 Ma. We found that 72% of duplicated GST gene pairs experienced gene losses or pseudogenization, whereas 28% of GST gene pairs have been retained in the soybean genome. The GST pseudogenes were under relaxed selective constraints, whereas functional GSTs were subject to strong purifying selection. Plant GST genes play important roles in stress tolerance and detoxification metabolism. By examining the gene expression responses to abiotic stresses and enzymatic properties of the ancestral and current proteins, we found that polyploidy-derived GST duplicates show the divergence in enzymatic activities. Through site-directed mutagenesis of ancestral proteins, this study revealed that nonsynonymous substitutions of key amino acid sites play an important role in the divergence of enzymatic functions of polyploidy-derived GST duplicates. These findings provide new insights into the evolutionary and functional dynamics of polyploidy-derived duplicate genes., (© The Author 2015. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.)

A simple method based on liquid chromatography-full-scan high-resolution accurate mass spectrometry (LC-HRMS) using a quadrupole time-of-flight (TOF) mass spectrometry was developed and optimized for corticosterone quantification in mouse plasma. Mouse plasma (100 μL) was extracted with methyl tert-butyl ether using prednisone as internal standard. Separation was performed on a short C18 column using a methanol-water gradient. Full-scan data were acquired in the TOF only mode, and extracted ion chromatograms were generated post-acquisition with the extract masses of the analytes. Enhanced sensitivity and reproducibility were acquired with optimized mass parameters. The calibration range was 8.24-412 ng/mL, and the limit of quantitation was 5.088 g/mL. Accuracy was between -5.9 and 8.6%. The precision of between-run (interday) and within-run (intraday) was within 5.6 and 6.9%, respectively. The LC-HRMS method was applied for plasma samples analysis from the stressed mice with and without ginseng treatment for the stress state estimation., (© The Author 2014. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Objectives: To determine the genome sequence of Acinetobacter baumannii strain MDR-TJ and characterize the mechanisms of multidrug resistance in this strain., Methods: The whole-genome sequence was determined using Roche 454 GS FLX Titanium. Subsequently, the gaps were closed by sequencing PCR products. The genome of strain MDR-TJ was annotated using IMG ER, the RAST annotation server and the BASys bacterial annotation system. The comM gene of MDR-TJ was examined to identify a possible antibiotic resistance island. Based on the results of multilocus sequence typing, we investigated seven multidrug-resistant A. baumannii strains belonging to global clone 2 (GC2) isolated from Asia, Australia and Europe to determine the backbone shared by resistance islands of GC2 isolates., Results: The A. baumannii strain MDR-TJ genome consists of a circular chromosome and a plasmid, pABTJ1. Strain MDR-TJ was assigned to sequence type ST2. Strain MDR-TJ harbours a 41.6 kb resistance island designated RI(MDR-TJ), which can be derived from the backbone of Tn6167 through the insertion of a Tn6022 into the 3'-end of the tetA(B) gene. Comparative analysis showed that transposon Tn6022 and its truncated forms prevailed in the antibiotic resistance islands of GC2 isolates. The carbapenem resistance gene bla(OXA-23) carried by transposon Tn2009 is located on a putatively conjugative plasmid, pABTJ1., Conclusions: A. baumannii strain MDR-TJ belongs to GC2 and is resistant to multiple antibiotics. A. baumannii MDR-TJ harbours a genomic resistance island that interrupts the comM gene. The carbapenem resistance of MDR-TJ is mediated by a putatively conjugative plasmid, pABTJ1.

Identifying how genes and their functions evolve after duplication is central to understanding gene family radiation. In this study, we systematically examined the functional diversification of the glutathione S-transferase (GST) gene family in Populus trichocarpa by integrating phylogeny, expression, substrate specificity, and enzyme kinetic data. GSTs are ubiquitous proteins in plants that play important roles in stress tolerance and detoxification metabolism. Genome annotation identified 81 GST genes in Populus that were divided into eight classes with distinct divergence in their evolutionary rate, gene structure, expression responses to abiotic stressors, and enzymatic properties of encoded proteins. In addition, when all the functional parameters were examined, clear divergence was observed within tandem clusters and between paralogous gene pairs, suggesting that subfunctionalization has taken place among duplicate genes. The two domains of GST proteins appear to have evolved under differential selective pressures. The C-terminal domain seems to have been subject to more relaxed functional constraints or divergent directional selection, which may have allowed rapid changes in substrate specificity, affinity, and activity, while maintaining the primary function of the enzyme. Our findings shed light on mechanisms that facilitate the retention of duplicate genes, which can result in a large gene family with a broad substrate spectrum and a wide range of reactivity toward different substrates.

Phylogenetic relationships of commercial Chinese truffles and their allies were investigated mainly by morphological studies and analyses of the sequences of ITS regions of nuclear ribosomal DNA. Two species, Tuber indicum and T. himalayense, closely related to the European T. melanosporum (the Perigord Truffle), are recognized among commercial Chinese black truffles. Both T. pseudohimalayense and T. sinense should be regarded as synonyms of T. indicum. Tuber species producing excavated ascomata are not monophyletic, suggesting that excavation of ascomata may have evolved more than once, or evolved once during the evolution of truffle species and then was lost once during the evolution of Tuber species.

Polymorphism for deletions was investigated in 1027 lines of tetraploid and hexaploid wheat and 420 lines of wheat diploid ancestors. A total of 26 deletions originating during the evolution of polyploid wheat were discovered among 155 investigated loci. Wheat chromosomes were divided into a proximal, low-recombination interval containing 69 loci and a distal, high-recombination interval containing 86 loci. A total of 23 deletions involved loci in the distal, high-recombination interval and only 3 involved loci in the proximal, low-recombination interval. The rates of DNA loss differed by several orders of magnitude in the two intervals. The rate of diploidization of polyploid wheat by deletions was estimated and was shown to have proceeded faster in the distal, high-recombination interval than in the proximal, low-recombination interval.

Recombination of chromosomes 3A(m) and 5A(m) of Triticum monococcum with closely homeologous chromosomes 3A and 5A of T. aestivum was compared with recombination across corresponding homologous chromosome pairs. Differentiation between the homeologues impacted recombination in the proximal regions of the long arms the most and in the distal regions of the long arms the least. It is concluded that this variation principally reflects allocation of multiple crossovers across an arm and positive crossover interference across chromosome arms. Recombination rates between homeologous chromosomes 5A(m) and 5A differed in the opposite sexes.

Homeologous wheat chromosome arms that differ by the presence or absence of a Nor locus or greatly differ in the numbers of copies of rRNA genes per Nor locus show conspicuous differences in the distribution of recombination. To assess directly the position effects of Nor loci on recombination across chromosome arms, a Triticum monococcum Nor9 haplotype was substituted for Triticum aestivum Nor9 haplotypes on two T. aestivum 1A chromosomes in the isogenic background of cv Chinese Spring. The numbers of rRNA genes in the 1A Nor9 haplotypes are greatly reduced relative to the T. monococcum haplotype. The substitution resulted in reduced recombination rate in the vicinity of the Nor9 locus. An intra-arm compensatory increase was observed in the proximal region of the arm so that the genetic length of the chromosome arm was unchanged. These findings suggest that Nor loci suppress recombination in their vicinity and change recombination patterns in Nor-bearing chromosome arms.

RFLP was investigated at 52 single-copy gene loci among six species of Aegilops, including both cross-fertilizing and self-fertilizing species. Average gene diversity (H) was found to correlate with the level of outcrossing. No relationship was found between H and the phylogenetic status of a species. In all six species, the level of RFLP at a locus was a function of the position of the locus on the chromosome and the recombination rate in the neighborhood of the locus. Loci in the proximal chromosome regions, which show greatly reduced recombination rates relative to the distal regions, were significantly less variable than loci in the distal chromosome regions in all six species. Variation in recombination rates was also reflected in the haplotype divergence between closely related species; loci in the chromosome regions with low recombination rates were found to be diverged less than those in the chromosome regions with high recombination rates. This relationship was not found among the more distantly related species.