Glia Accumulate Evidence that Actions Are Futile and Suppress Unsuccessful Behavior.

When a behavior repeatedly fails to achieve its goal, animals often give up and become passive, which can be strategic for preserving energy or regrouping between attempts. It is unknown how the brain identifies behavioral failures and mediates this behavioral-state switch. In larval zebrafish swimming in virtual reality, visual feedback can be withheld so that swim attempts fail to trigger expected visual flow. After tens of seconds of such motor futility, animals became passive for similar durations. Whole-brain calcium imaging revealed noradrenergic neurons that responded specifically to failed swim attempts and radial astrocytes whose calcium levels accumulated with increasing numbers of failed attempts. Using cell ablation and optogenetic or chemogenetic activation, we found that noradrenergic neurons progressively activated brainstem radial astrocytes, which then suppressed swimming. Thus, radial astrocytes perform a computation critical for behavior: they accumulate evidence that current actions are ineffective and consequently drive changes in behavioral states. VIDEO ABSTRACT.

Neurophysiology of Human Perceptual Decision-Making.

The discovery of neural signals that reflect the dynamics of perceptual decision formation has had a considerable impact. Not only do such signals enable detailed investigations of the neural implementation of the decision-making process but they also can expose key elements of the brain's decision algorithms. For a long time, such signals were only accessible through direct animal brain recordings, and progress in human neuroscience was hampered by the limitations of noninvasive recording techniques. However, recent methodological advances are increasingly enabling the study of human brain signals that finely trace the dynamics of the unfolding decision process. In this review, we highlight how human neurophysiological data are now being leveraged to furnish new insights into the multiple processing levels involved in forming decisions, to inform the construction and evaluation of mathematical models that can explain intra- and interindividual differences, and to examine how key ancillary processes interact with core decision circuits.

Lipid metabolism in sickness and in health: Emerging regulators of lipotoxicity.

Lipids play crucial roles in signal transduction, contribute to the structural integrity of cellular membranes, and regulate energy metabolism. Questions remain as to which lipid species maintain metabolic homeostasis and which disrupt essential cellular functions, leading to metabolic disorders. Here, we discuss recent advances in understanding lipid metabolism with a focus on catabolism, synthesis, and signaling. Technical advances, including functional genomics, metabolomics, lipidomics, lipid-protein interaction maps, and advances in mass spectrometry, have uncovered new ways to prioritize molecular mechanisms mediating lipid function. By reviewing what is known about the distinct effects of specific lipid species in physiological pathways, we provide a framework for understanding newly identified targets regulating lipid homeostasis with implications for ameliorating metabolic diseases.

Biochemical evidence accumulates across neurons to drive a network-level eruption.

Neural network computations are usually assumed to emerge from patterns of fast electrical activity. Challenging this view, we show that a male fly's decision to persist in mating hinges on a biochemical computation that enables processing over minutes to hours. Each neuron in a recurrent network contains slightly different internal molecular estimates of mating progress. Protein kinase A (PKA) activity contrasts this internal measurement with input from the other neurons to represent accumulated evidence that the goal of the network has been achieved. When consensus is reached, PKA pushes the network toward a large-scale and synchronized burst of calcium influx that we call an eruption. Eruptions transform continuous deliberation within the network into an all-or-nothing output, after which the male will no longer sacrifice his life to continue mating. Here, biochemical activity, invisible to most large-scale recording techniques, is the key computational currency directing behavior and motivational state.

Accelerated plasma-cell differentiation in Bach2-deficient mouse B cells is caused by altered IRF4 functions.

Transcription factors BACH2 and IRF4 are both essential for antibody class-switch recombination (CSR) in activated B lymphocytes, while they oppositely regulate the differentiation of plasma cells (PCs). Here, we investigated how BACH2 and IRF4 interact during CSR and plasma-cell differentiation. We found that BACH2 organizes heterochromatin formation of target gene loci in mouse splenic B cells, including targets of IRF4 activation such as Aicda, an inducer of CSR, and Prdm1, a master plasma-cell regulator. Release of these gene loci from heterochromatin in response to B-cell receptor stimulation was coupled to AKT-mTOR pathway activation. In Bach2-deficient B cells, PC genes' activation depended on IRF4 protein accumulation, without an increase in Irf4 mRNA. Mechanistically, a PU.1-IRF4 heterodimer in activated B cells promoted BACH2 function by inducing gene expression of Bach2 and Pten, a negative regulator of AKT signaling. Elevated AKT activity in Bach2-deficient B cells resulted in IRF4 protein accumulation. Thus, BACH2 and IRF4 mutually modulate the activity of each other, and BACH2 inhibits PC differentiation by both the repression of PC genes and the restriction of IRF4 protein accumulation.

Tropical trees inherit low-frequency somatic mutations.

A new study by Schmitt et al. revealed that somatic mutations in tropical trees are passed on to their offspring. Furthermore, the study noted that the majority of inherited mutations were present at low allelic frequencies within the tree.

CHP1 Regulates Compartmentalized Glycerolipid Synthesis by Activating GPAT4.

Cells require a constant supply of fatty acids to survive and proliferate. Fatty acids incorporate into membrane and storage glycerolipids through a series of endoplasmic reticulum (ER) enzymes, but how these enzymes are regulated is not well understood. Here, using a combination of CRISPR-based genetic screens and unbiased lipidomics, we identified calcineurin B homologous protein 1 (CHP1) as a major regulator of ER glycerolipid synthesis. Loss of CHP1 severely reduces fatty acid incorporation and storage in mammalian cells and invertebrates. Mechanistically, CHP1 binds and activates GPAT4, which catalyzes the initial rate-limiting step in glycerolipid synthesis. GPAT4 activity requires CHP1 to be N-myristoylated, forming a key molecular interface between the two proteins. Interestingly, upon CHP1 loss, the peroxisomal enzyme, GNPAT, partially compensates for the loss of ER lipid synthesis, enabling cell proliferation. Thus, our work identifies a conserved regulator of glycerolipid metabolism and reveals plasticity in lipid synthesis of proliferating cells.

Balance between asymmetric allocation and repair of somatic damage in unicellular life forms as an ancient form of r/K selection.

Over the course of multiple divisions, cells accumulate diverse nongenetic, somatic damage including misfolded and aggregated proteins and cell wall defects. If the rate of damage accumulation exceeds the rate of dilution through cell growth, a dedicated mitigation strategy is required to prevent eventual population collapse. Strategies for somatic damage control can be divided into two categories, asymmetric allocation and repair, which are not, in principle, mutually exclusive. We explore a mathematical model to identify the optimal strategy, maximizing the total cell number, over a wide range of environmental and physiological conditions. The optimal strategy is primarily determined by extrinsic, damage-independent mortality and the physiological model for damage accumulation that can be either independent (linear) or increasing (exponential) with respect to the prior accumulated damage. Under the linear regime, the optimal strategy is either exclusively repair or asymmetric allocation, whereas under the exponential regime, the optimal strategy is a combination of asymmetry and repair. Repair is preferred when extrinsic mortality is low, whereas at high extrinsic mortality, asymmetric damage allocation becomes the strategy of choice. We hypothesize that at an early stage of life evolution, optimization over repair and asymmetric allocation of somatic damage gave rise to r and K selection strategists.

Broad variation in response of individual introns to splicing inhibitors in a humanized yeast strain.

Intron branchpoint (BP) recognition by the U2 snRNP is a critical step of splicing, vulnerable to recurrent cancer mutations and bacterial natural product inhibitors. The BP binds a conserved pocket in the SF3B1 (human) or Hsh155 (yeast) U2 snRNP protein. Amino acids that line this pocket affect the binding of splicing inhibitors like Pladienolide-B (Plad-B), such that organisms differ in their sensitivity. To study the mechanism of splicing inhibitor action in a simplified system, we modified the naturally Plad-B resistant yeast Saccharomyces cerevisiae by changing 14 amino acids in the Hsh155 BP pocket to those from human. This humanized yeast grows normally, and splicing is largely unaffected by the mutation. Splicing is inhibited within minutes after the addition of Plad-B, and different introns appear inhibited to different extents. Intron-specific inhibition differences are also observed during cotranscriptional splicing in Plad-B using single-molecule intron tracking to minimize gene-specific transcription and decay rates that cloud estimates of inhibition by standard RNA-seq. Comparison of Plad-B intron sensitivities to those of the structurally distinct inhibitor Thailanstatin-A reveals intron-specific differences in sensitivity to different compounds. This work exposes a complex relationship between the binding of different members of this class of inhibitors to the spliceosome and intron-specific rates of BP recognition and catalysis. Introns with variant BP sequences seem particularly sensitive, echoing observations from mammalian cells, where monitoring individual introns is complicated by multi-intron gene architecture and alternative splicing. The compact yeast system may hasten the characterization of splicing inhibitors, accelerating improvements in selectivity and therapeutic efficacy.

NME4 mediates metabolic reprogramming and promotes nonalcoholic fatty liver disease progression.

Nonalcoholic fatty liver disease (NAFLD) is mainly characterized by excessive fat accumulation in the liver, and it is associated with liver-related complications and adverse systemic diseases. NAFLD has become the most prevalent liver disease; however, effective therapeutic agents for NAFLD are still lacking. We combined clinical data with proteomics and metabolomics data, and found that the mitochondrial nucleoside diphosphate kinase NME4 plays a central role in mitochondrial lipid metabolism. Nme4 is markedly upregulated in mice fed with high-fat diet, and its expression is positively correlated with the level of steatosis. Hepatic deletion of Nme4 suppresses the progression of hepatic steatosis. Further studies demonstrated that NME4 interacts with several key enzymes in coenzyme A (CoA) metabolism and increases the level of acetyl-CoA and malonyl-CoA, which are the major lipid components of the liver in NAFLD. Increased level of acetyl-CoA and malonyl-CoA  lead to increased triglyceride levels and lipid accumulation in the liver. Taken together, these findings reveal that NME4 is a critical regulator of NAFLD progression and a potential therapeutic target for NAFLD.

Metformin Promotes Antitumor Immunity via Endoplasmic-Reticulum-Associated Degradation of PD-L1.

Metformin has been reported to possess antitumor activity and maintain high cytotoxic T lymphocyte (CTL) immune surveillance. However, the functions and detailed mechanisms of metformin's role in cancer immunity are not fully understood. Here, we show that metformin increases CTL activity by reducing the stability and membrane localization of programmed death ligand-1 (PD-L1). Furthermore, we discover that AMP-activated protein kinase (AMPK) activated by metformin directly phosphorylates S195 of PD-L1. S195 phosphorylation induces abnormal PD-L1 glycosylation, resulting in its ER accumulation and ER-associated protein degradation (ERAD). Consistently, tumor tissues from metformin-treated breast cancer patients exhibit reduced PD-L1 levels with AMPK activation. Blocking the inhibitory signal of PD-L1 by metformin enhances CTL activity against cancer cells. Our findings identify a new regulatory mechanism of PD-L1 expression through the ERAD pathway and suggest that the metformin-CTLA4 blockade combination has the potential to increase the efficacy of immunotherapy.

Mixed Selectivity Coding of Content-Temporal Detail by Dorsomedial Posterior Parietal Neurons.

The dorsomedial posterior parietal cortex (dmPPC) is part of a higher-cognition network implicated in elaborate processes underpinning memory formation, recollection, episode reconstruction, and temporal information processing. Neural coding for complex episodic processing is however under-documented. Here, we recorded extracellular neural activities from three male rhesus macaques (Macaca mulatta) and revealed a set of neural codes of "neuroethogram" in the primate parietal cortex. Analyzing neural responses in macaque dmPPC to naturalistic videos, we discovered several groups of neurons that are sensitive to different categories of ethogram items, low-level sensory features, and saccadic eye movement. We also discovered that the processing of category and feature information by these neurons is sustained by the accumulation of temporal information over a long timescale of up to 30 s, corroborating its reported long temporal receptive windows. We performed an additional behavioral experiment with additional two male rhesus macaques and found that saccade-related activities could not account for the mixed neuronal responses elicited by the video stimuli. We further observed monkeys' scan paths and gaze consistency are modulated by video content. Taken altogether, these neural findings explain how dmPPC weaves fabrics of ongoing experiences together in real time. The high dimensionality of neural representations should motivate us to shift the focus of attention from pure selectivity neurons to mixed selectivity neurons, especially in increasingly complex naturalistic task designs.

The rearrangement of co-cultured cellular model systems via collective cell migration.

Cancer invasion through the surrounding epithelium and extracellular matrix (ECM) is the one of the main characteristics of cancer progression. While significant effort has been made to predict cancer cells response under various drug therapies, much less attention has been paid to understand the physical interactions between cancer cells and their microenvironment, which are essential for cancer invasion. Considering these physical interactions on various co-cultured in vitro model systems by emphasizing the role of viscoelasticity, the tissue surface tension, solid stress, and their inter-relations is a prerequisite for establishing the main factors that influence cancer cell spread and develop an efficient strategy to suppress it. This review focuses on the role of viscoelasticity caused by collective cell migration (CCM) in the context of mono-cultured and co-cultured cancer systems, and on the modeling approaches aimed at reproducing and understanding these biological systems. In this context, we do not only review previously-published biophysics models for collective cell migration, but also propose new extensions of those models to include solid stress accumulated within the spheroid core region and cell residual stress accumulation caused by CCM.

The dynamics along the biointerface between the epithelial and cancer mesenchymal cells: Modeling consideration.

Epithelial cancer is the one of most lethal cancer type worldwide. Targeting the early stage of disease would allow dramatic improvements in the survival of cancer patients. The early stage of the disease is related to cancer cell spreading across surrounding healthy epithelium. Consequently, deeper insight into cell dynamics along the biointerface between epithelial and cancer (mesenchymal) cells is necessary in order to control the disease as soon as possible. Cell dynamics along this epithelial-cancer biointerface is the result of the interplay between various biological and physical mechanisms. Despite extensive research devoted to study cancer cell spreading across the epithelium, we still do not understand the physical mechanisms which influences the dynamics along the biointerface. These physical mechanisms are related to the interplay between physical parameters such as: (1) interfacial tension between cancer and epithelial subpopulations, (2) established interfacial tension gradients, (3) the bending rigidity of the biointerface and its impact on the interfacial tension, (4) surface tension of the subpopulations, (5) viscoelasticity caused by collective cell migration, and (6) cell residual stress accumulation. The main goal of this study is to review some of these physical parameters in the context of the epithelial/cancer biointerface elaborated on the model system such as the biointerface between breast epithelial MCF-10A cells and cancer MDA-MB-231 cells and then to incorporate these parameters into a new biophysical model that could describe the dynamics of the biointerface. We conclude by discussing three biophysical scenarios for cell dynamics along the biointerface, which can occur depending on the magnitude of the generated shear stress: a smooth biointerface, a slightly-perturbed biointerface and an intensively-perturbed biointerface in the context of the Kelvin-Helmholtz instability. These scenarios are related to the probability of cancer invasion.

The acyl-acyl carrier protein thioesterases GmFATA1 and GmFATA2 are essential for fatty acid accumulation and growth in soybean.

Acyl-acyl carrier protein (ACP) thioesterases (FAT) hydrolyze acyl-ACP complexes to release FA in plastids, which ultimately affects FA biosynthesis and profiles. Soybean GmFATA1 and GmFATA2 are homoeologous genes encoding oleoyl-ACP thioesterases whose role in seed oil accumulation and plant growth has not been defined. Using CRISPR/Cas9 gene editing mutation of Gmfata1 or 2 led to reduced leaf FA content and growth defect at the early seedling stage. In contrast, no homozygous double mutants were obtained. Combined this indicates that GmFATA1 and GmFATA2 display overlapping, but not complete functional redundancy. Combined transcriptomic and lipidomic analysis revealed a large number of genes involved in FA synthesis and FA chain elongation are expressed at reduced level in the Gmfata1 mutant, accompanied by a lower triacylglycerol abundance at the early seedling stage. Further analysis showed that the Gmfata1 or 2 mutants had increased composition of the beneficial FA, oleic acid. The growth defect of Gmfata1 could be at least partially attributed to reduced acetyl-CoA carboxylase activity, reduced abundance of five unsaturated monogalactosyldiacylglycerol lipids, and altered chloroplast morphology. On the other hand, overexpression of GmFATA in soybean led to significant increases in leaf FA content by 5.7%, vegetative growth, and seed yield by 26.9%, and seed FA content by 23.2%. Thus, overexpression of GmFATA is an effective strategy to enhance soybean oil content and yield.

The R2R3 MYB Ruby1 is activated by two cold responsive ethylene response factors, via the retrotransposon in its promoter, to positively regulate anthocyanin biosynthesis in citrus.

Anthocyanins are natural pigments and dietary antioxidants that play multiple biological roles in plants and are important in animal and human nutrition. Low temperature (LT) promotes anthocyanin biosynthesis in many species including blood orange. A retrotransposon in the promoter of Ruby1, which encodes an R2R3 MYB transcription factor, controls cold-induced anthocyanin accumulation in blood orange flesh. However, the specific mechanism remains unclear. In this study, we characterized two LT-induced ETHYLENE RESPONSE FACTORS (CsERF054 and CsERF061). Both CsERF054 and CsERF061 can activate the expression of CsRuby1 by directly binding to a DRE/CRT cis-element within the retrotransposon in the promoter of CsRuby1, thereby positively regulating anthocyanin biosynthesis. Further investigation indicated that CsERF061 also forms a protein complex with CsRuby1 to co-activate the expression of anthocyanin biosynthetic genes, providing a dual mechanism for the upregulation of the anthocyanin pathway. These results provide insights into how LT mediates anthocyanin biosynthesis and increase the understanding of the regulatory network of anthocyanin biosynthesis in blood orange.

Utilizing high-resolution ribosome profiling for the global investigation of gene expression in Chlamydomonas.

Ribosome profiling (Ribo-seq) is a powerful method for the deep analysis of translation mechanisms and regulatory circuits during gene expression. Extraction and sequencing of ribosome-protected fragments (RPFs) and parallel RNA-seq yields genome-wide insight into translational dynamics and post-transcriptional control of gene expression. Here, we provide details on the Ribo-seq method and the subsequent analysis with the unicellular model alga Chlamydomonas reinhardtii (Chlamydomonas) for generating high-resolution data covering more than 10 000 different transcripts. Detailed analysis of the ribosomal offsets on transcripts uncovers presumable transition states during translocation of elongating ribosomes within the 5' and 3' sections of transcripts and characteristics of eukaryotic translation termination, which are fundamentally distinct for chloroplast translation. In chloroplasts, a heterogeneous RPF size distribution along the coding sequence indicates specific regulatory phases during protein synthesis. For example, local accumulation of small RPFs correlates with local slowdown of psbA translation, possibly uncovering an uncharacterized regulatory step during PsbA/D1 synthesis. Further analyses of RPF distribution along specific cytosolic transcripts revealed characteristic patterns of translation elongation exemplified for the major light-harvesting complex proteins, LHCs. By providing high-quality datasets for all subcellular genomes and attaching our data to the Chlamydomonas reference genome, we aim to make ribosome profiles easily accessible for the broad research community. The data can be browsed without advanced bioinformatic background knowledge for translation output levels of specific genes and their splice variants and for monitoring genome annotation.

PbrbZIP15 promotes sugar accumulation in pear via activating the transcription of the glucose isomerase gene PbrXylA1.

The composition and abundance of soluble sugars in mature pear (Pyrus) fruit are important for its acceptance by consumers. However, our understanding of the genes responsible for soluble sugar accumulation remains limited. In this study, a S1-group member of bZIP gene family, PbrbZIP15, was characterized from pear genome through the combined analyses of metabolite and transcriptome data followed by experimental validation. PbrbZIP15, located in nucleus, was found to function in fructose, sucrose, and total soluble sugar accumulation in pear fruit and calli. After analyzing the expression profiles of sugar-metabolism-related genes and the distribution of cis-acting elements in their promoters, the glucose isomerase 1 gene (PbrXylA1), whose corresponding protein catalyzed the isomerization of glucose and fructose in vitro, was identified as a downstream target gene of PbrbZIP15. PbrbZIP15 could directly bind to the G-box element in PbrXylA1 promoter and activate its transcription, as evidenced by chromatin immunoprecipitation-quantitative PCR, yeast one-hybrid, electrophoretic mobility shift assay, and dual-luciferase assay. PbrXylA1, featuring a leucine-rich signal peptide in its N-terminal, was localized to the endoplasmic reticulum. It was validated to play a significant role in fructose, sucrose, and total soluble sugar accumulation in pear fruit and calli, which was associated with the upregulated fructose/glucose ratio. Further studies revealed a positive correlation between the sucrose content and the expression levels of several sucrose-biosynthesis-related genes (PbrFRK3/8, PbrSPS1/3/4/8, and PbrSPP1) in PbrbZIP15-/PbrXylA1-transgenic fruit/calli. In conclusion, our results suggest that PbrbZIP15-induced soluble sugar accumulation during pear development is at least partly attributed to the activation of PbrXylA1 transcription.

Genetic factors of grain cadmium concentration in Polish wheat (Triticum polonicum L.).

Wheat (Triticum aestivum L.) is one of the most important crops worldwide and a major source of human Cd intake. Limiting grain Cd concentration (Gr_Cd_Conc) in wheat is necessary to ensure food safety. However, the genetic factors associated with Cd uptake, translocation, distribution, and Gr_Cd_Conc in wheat are poorly understood. Here, we mapped quantitative trait loci (QTL) for Gr_Cd_Conc and its related transport pathway using a recombinant inbred line (RIL_DT) population derived from two Polish wheat varieties (dwarf Polish wheat [DPW] and tall Polish wheat [TPW]). We identified 29 novel major QTLs for grain and tissue Cd concentration; 14 novel major QTLs for Cd uptake, translocation, and distribution; and 27 major QTLs for agronomic traits. We also analyzed the pleiotropy of these QTLs. Six novel QTLs (QGr_Cd_Conc-1A, QGr_Cd_Conc-3A, QGr_Cd_Conc-4B, QGr_Cd_Conc-5B, QGr_Cd_Conc-6A and QGr_Cd_Conc-7A) for Gr_Cd_Conc explained 8.16-17.02% of the phenotypic variation. QGr_Cd_Conc-3A, QGr_Cd_Conc-6A and QGr_Cd_Conc-7A pleiotropically regulated Cd transport; three other QTLs were organ-specific for Gr_Cd_Conc. We fine-mapped the locus of QGr_Cd_Conc-4B and identified the candidate gene as Cation/Ca exchanger 2 (TpCCX2-4B), which was differentially expressed in DPW and TPW. It encodes an endoplasmic reticulum membrane/plasma membrane-localized Cd efflux transporter in yeast. Overexpression of TpCCX2-4B reduced Gr_Cd_Conc in rice. The average Gr_Cd_Conc was significantly lower in TpCCX2-4BDPW genotypes than in TpCCX2-4BTPWgenotypes of the RIL_DT population and two other natural populations, based on a KASP marker derived from the different promoter sequences between TpCCX2-4BDPW and TpCCX2-4BTPW. Our study reveals the genetic mechanism of Cd accumulation in wheat and provides valuable resources for genetic improvement of low-Cd-accumulating wheat cultivars.

Mathematical modeling of fluid dynamics in in vitro gut fermentation systems: A new tool to improve the interpretation of microbial metabolism.

In vitro systems are widely employed to assess the impact of dietary compounds on the gut microbiota and their conversion into beneficial bacterial metabolites. However, the complex fluid dynamics and multi-segmented nature of these systems can complicate the comprehensive analysis of dietary compound fate, potentially confounding physical dilution or washout with microbial catabolism. In this study, we developed fluid dynamics models based on sets of ordinary differential equations to simulate the behavior of an inert compound within two commonly used in vitro systems: the continuous two-stage PolyFermS system and the semi-continuous multi-segmented SHIME® system as well as into various declinations of those systems. The models were validated by investigating the fate of blue dextran, demonstrating excellent agreement between experimental and modeling data (with r2 values ranging from 0.996 to 0.86 for different approaches). As a proof of concept for the utility of fluid dynamics models in in vitro system, we applied generated models to interpret metabolomic data of procyanidin A2 (ProA2) generated from the addition of proanthocyanidin (PAC)-rich cranberry extract to both the PolyFermS and SHIME® systems. The results suggested ProA2 degradation by the gut microbiota when compared to the modeling of an inert compound. Models of fluid dynamics developed in this study provide a foundation for comprehensive analysis of gut metabolic data in commonly utilized in vitro PolyFermS and SHIME® bioreactor systems and can enable a more accurate understanding of the contribution of bacterial metabolism to the variability in the concentration of target metabolites.