Every data-rich community research effort requires a clear plan for ensuring the quality of the data interpretation and comparability of analyses. To address this need within the Human Proteome ...Project (HPP) of the Human Proteome Organization (HUPO), we have developed through broad consultation a set of mass spectrometry data interpretation guidelines that should be applied to all HPP data contributions. For submission of manuscripts reporting HPP protein identification results, the guidelines are presented as a one-page checklist containing 15 essential points followed by two pages of expanded description of each. Here we present an overview of the guidelines and provide an in-depth description of each of the 15 elements to facilitate understanding of the intentions and rationale behind the guidelines, for both authors and reviewers. Broadly, these guidelines provide specific directions regarding how HPP data are to be submitted to mass spectrometry data repositories, how error analysis should be presented, and how detection of novel proteins should be supported with additional confirmatory evidence. These guidelines, developed by the HPP community, are presented to the broader scientific community for further discussion.
WNT signaling activation in colorectal cancers (CRCs) occurs through APC inactivation or β-catenin mutations. Both processes promote β-catenin nuclear accumulation, which up-regulates ...epithelial-to-mesenchymal transition (EMT). We investigated β-catenin localization, transcriptome, and phenotypic differences of HCT116 cells containing a wild-type (HCT116-WT) or mutant β-catenin allele (HCT116-MT), or parental cells with both WT and mutant alleles (HCT116-P). We then analyzed β-catenin expression and associated phenotypes in CRC tissues. Wild-type β-catenin showed membranous localization, whereas mutant showed nuclear localization; both nuclear and non-nuclear localization were observed in HCT116-P. Microarray analysis revealed down-regulation of Claudin-7 and E-cadherin in HCT116-MT vs. HCT116-WT. Claudin-7 was also down-regulated in HCT116-P vs. HCT116-WT without E-cadherin dysregulation. We found that ZEB1 is a critical EMT factor for mutant β-catenin-mediated loss of E-cadherin and Claudin-7 in HCT116-P and HCT116-MT cells. We also demonstrated that E-cadherin binds to both WT and mutant β-catenin, and loss of E-cadherin releases β-catenin from the cell membrane and leads to its degradation. Alteration of Claudin-7, as well as both Claudin-7 and E-cadherin respectively caused tight junction (TJ) impairment in HCT116-P, and dual loss of TJs and adherens junctions (AJs) in HCT116-MT. TJ loss increased cell motility, and subsequent AJ loss further up-regulated that. Immunohistochemistry analysis of 101 CRCs revealed high (14.9%), low (52.5%), and undetectable (32.6%) β-catenin nuclear expression, and high β-catenin nuclear expression was significantly correlated with overall survival of CRC patients (P = 0.009). Our findings suggest that β-catenin activation induces EMT progression by modifying cell-cell junctions, and thereby contributes to CRC aggressiveness.
Reproductive plasticity is the ability of an animal to modulate its reproductive functions in response to environmental changes. For example, Caenorhabditis elegans, a free-living nematode, can ...adjust the onsets of oogenesis and embryogenesis under harsh environmental conditions, including starvation. However, the molecular mechanisms used to perceive and translate environmental signals into reproductive functional adjustments remain largely uncharacterized. We discovered that in C. elegans, the glutamate receptor homolog MGL-1 initiated reproductive plasticity in response to starvation. A genetic analysis of the mutant gene, mgl-1(tm1811), suggested that starvation delayed the onsets of oogenesis and embryogenesis via MGL-1. Cell-specific rescues of mgl-1 deletion mutants, which used transgenic lines designed to express MGL-1 in different neurons (e.g., RMD, AIA, AIY, and NSM), suggested that only AIY-rescued animals exhibited normal delays in oogenesis and embryogenesis equivalent to those of wild-type animals, suggesting recovery. Furthermore, in AIY neurons, MGL-1 appears to use neuropeptide signaling, rather than glutamate, to translate starvation stimuli into delayed oogenesis and embryogenesis. Our findings, which reveal molecular linkages between starvation signals and reproductive alterations, may provide a basis for understanding energy reallocation mechanisms, as the mgl-1 deletion mutant exhibited more severe reductions in lifespan and fat accumulation than did wild-type animals under starvation conditions. Taken together, MGL-1 is the molecular driver underlying the translation of starvation signals to reproduction plasticity in an AIY neuron-specific manner.
•MGL-1 delays oogenesis and embryogenesis in response to starvation.•MGL-1 exerts effects on reproductive plasticity via AIY neurons.•MMGL-1 use neuropeptides in AIY neurons to adjust reproduction under starvation.•mgl-1(-) plays a trade-off role between less fat deposit and lifespan under starvation.
The Human Proteome Organization’s (HUPO) Human Proteome Project (HPP) developed Mass Spectrometry (MS) Data Interpretation Guidelines that have been applied since 2016. These guidelines have helped ...ensure that the emerging draft of the complete human proteome is highly accurate and with low numbers of false-positive protein identifications. Here, we describe an update to these guidelines based on consensus-reaching discussions with the wider HPP community over the past year. The revised 3.0 guidelines address several major and minor identified gaps. We have added guidelines for emerging data independent acquisition (DIA) MS workflows and for use of the new Universal Spectrum Identifier (USI) system being developed by the HUPO Proteomics Standards Initiative (PSI). In addition, we discuss updates to the standard HPP pipeline for collecting MS evidence for all proteins in the HPP, including refinements to minimum evidence. We present a new plan for incorporating MassIVE-KB into the HPP pipeline for the next (HPP 2020) cycle in order to obtain more comprehensive coverage of public MS data sets. The main checklist has been reorganized under headings and subitems, and related guidelines have been grouped. In sum, Version 2.1 of the HPP MS Data Interpretation Guidelines has served well, and this timely update to version 3.0 will aid the HPP as it approaches its goal of collecting and curating MS evidence of translation and expression for all predicted ∼20 000 human proteins encoded by the human genome.
Background
Carbohydrate antigen (CA) 19–9 is a known pancreatic cancer (PC) biomarker, but is not commonly used for general screening due to its low sensitivity and specificity. This study aimed to ...develop a serum metabolites‐based diagnostic calculator for detecting PC with high accuracy.
Methods
A targeted quantitative approach of direct flow injection‐tandem mass spectrometry combined with liquid chromatography–tandem mass spectrometry was employed for metabolomic analysis of serum samples using an Absolute IDQ™ p180 kit. Integrated metabolomic analysis was performed on 241 pooled or individual serum samples collected from healthy donors and patients from nine disease groups, including chronic pancreatitis, PC, other cancers, and benign diseases. Orthogonal partial least squares discriminant analysis (OPLS‐DA) based on characteristics of 116 serum metabolites distinguished patients with PC from those with other diseases. Sparse partial least squares discriminant analysis (SPLS‐DA) was also performed, incorporating simultaneous dimension reduction and variable selection. Predictive performance between discrimination models was compared using a 2‐by‐2 contingency table of predicted probabilities obtained from the models and actual diagnoses.
Results
Predictive values obtained through OPLS‐DA for accuracy, sensitivity, specificity, balanced accuracy, and area under the receiver operating characteristic curve (AUC) were 0.9825, 0.9916, 0.9870, 0.9866, and 0.9870, respectively. The number of metabolite candidates was narrowed to 76 for SPLS‐DA. The SPLS‐DA‐obtained predictive values for accuracy, sensitivity, specificity, balanced accuracy, and AUC were 0.9773, 0.9649, 0.9832, 0.9741, and 0.9741, respectively.
Conclusions
We successfully developed a 76 metabolome‐based diagnostic panel for detecting PC that demonstrated high diagnostic performance in differentiating PC from other diseases.
Pheromones are neuronal signals that stimulate conspecific individuals to react to environmental stressors or stimuli. Research on the ascaroside (ascr) pheromones in
and other nematodes has made ...great progress since ascr#1 was first isolated and biochemically defined in 2005. In this review, we highlight the current research on the structural diversity, biosynthesis, and pleiotropic neuronal functions of ascr pheromones and their implications in animal physiology. Experimental evidence suggests that ascr biosynthesis starts with conjugation of ascarylose to very long-chain fatty acids that are then processed via peroxisomal β-oxidation to yield diverse ascr pheromones. We also discuss the concentration and stage-dependent pleiotropic neuronal functions of ascr pheromones. These functions include dauer induction, lifespan extension, repulsion, aggregation, mating, foraging and detoxification, among others. These roles are carried out in coordination with three G protein-coupled receptors that function as putative pheromone receptors: SRBC-64/66, SRG-36/37, and DAF-37/38. Pheromone sensing is transmitted in sensory neurons via DAF-16-regulated glutamatergic neurotransmitters. Neuronal peroxisomal fatty acid β-oxidation has important cell-autonomous functions in the regulation of neuroendocrine signaling, including neuroprotection. In the future, translation of our knowledge of nematode ascr pheromones to higher animals might be beneficial, as ascr#1 has some anti-inflammatory effects in mice. To this end, we propose the establishment of
(
omone
) as a new subset of integrated disciplinary research area within chemical ecology for system-wide investigation of animal pheromones.
Although peroxisomal fatty acid (FA) β-oxidation is known to be critical for animal development, the cellular mechanisms that control the manner in which its neuronal deficiency causes developmental ...defects remain unclear. To elucidate the potential cellular consequences of neuronal FA metabolic disorder for dauer development, an alternative developmental process in Caenorhabditis elegans that occurs during stress, we investigated the sequential effects of its corresponding genetic deficiency. Here, we show that the daf-22 gene in peroxisomal FA β-oxidation plays a distinct role in ASK neurons, and its deficiency interrupts dauer development even in the presence of the exogenous ascaroside pheromones that induce such development. Un-metabolized FAs accumulated in ASK neurons of daf-22 mutants stimulate the endoplasmic reticulum (ER) stress response, which may enhance the XBP-1 activity that promotes the transcription of neuronal insulin-like peptides. These sequential cell-autonomous reactions in ASK neurons then activate insulin/IGF-1 signaling, which culminates in the suppression of DAF-16/FOXO activity. This suppression results in the interruption of dauer development, independently of pheromone presence. These findings suggest that neuronal peroxisomal FA β-oxidation is indispensable for animal development by regulating the ER stress response and neuroendocrine signaling.
Under stressful conditions, the early larvae of
enter dauer diapause, a non-aging period, driven by the seemingly opposite influence of ascaroside pheromones (ASCRs) and steroid hormone dafachronic ...acids (DAs). However, the molecular basis of how these small molecules engage in competitive crosstalk in coordination with insulin/IGF-1 signaling (IIS) remains elusive. Here we report a novel transcriptional regulatory pathway that seems to operate between the ASCR and DA biosynthesis under
(AL) feeding conditions or bacterial deprivation (BD). Although expression of the ASCR and DA biosynthetic genes reciprocally inhibit each other, ironically and interestingly, such dietary cue-mediated modulation requires the presence of the competitors. Under BD, induction of ASCR biosynthetic gene expression required DA, while ASCR suppresses the expression of the DA biosynthetic gene
. The negative regulation of DA by ASCR was IIS-dependent, whereas
regulation appeared to be independent of IIS. These observations suggest that the presence of ASCR determines the IIS-dependency of DA gene expression regardless of dietary conditions. Thus, our work defines a molecular basis for a novel reciprocal gene regulation of pheromones and hormones to cope with stressful conditions during development and aging.
The Human Proteome Project (HPP) annually reports on progress throughout the field in credibly identifying and characterizing the human protein parts list and making proteomics an integral part of ...multiomics studies in medicine and the life sciences. NeXtProt release 2018–01–17, the baseline for this sixth annual HPP special issue of the Journal of Proteome Research, contains 17 470 PE1 proteins, 89% of all neXtProt predicted PE1–4 proteins, up from 17 008 in release 2017–01–23 and 13 975 in release 2012–02–24. Conversely, the number of neXtProt PE2,3,4 missing proteins has been reduced from 2949 to 2579 to 2186 over the past two years. Of the PE1 proteins, 16 092 are based on mass spectrometry results, and 1378 on other kinds of protein studies, notably protein–protein interaction findings. PeptideAtlas has 15 798 canonical proteins, up 625 over the past year, including 269 from SUMOylation studies. The largest reason for missing proteins is low abundance. Meanwhile, the Human Protein Atlas has released its Cell Atlas, Pathology Atlas, and updated Tissue Atlas, and is applying recommendations from the International Working Group on Antibody Validation. Finally, there is progress using the quantitative multiplex organ-specific popular proteins targeted proteomics approach in various disease categories.
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