Candidate Signals Research Articles

Role of α2δ3 in Cellular Synchronization of the Suprachiasmatic Nucleus Under Constant Light Conditions

By the effort to identify candidate signaling molecules important for the formation of robust circadian rhythms in the suprachiasmatic nucleus (SCN), the mammalian circadian center, here we characterize the role of α2δ proteins, synaptic molecules initially identified as an auxiliary subunit of the voltage dependent calcium channel, in circadian rhythm formation. In situ hybridization study demonstrated that type 3 α2δ gene (α2δ3) was strongly expressed in the SCN. Mice without this isoform (Cacna2d3−/−) did not maintain proper circadian locomotor activity rhythms under a constant light (LL) condition, whereas under a constant dark (DD) condition, these mice showed a similar period length and similar light-responsiveness as compared to wild type mice. Reflecting this behavioral phenotype, Cacna2d3−/− mice showed a severely impaired Per1 expression rhythm in the SCN under LL, but not under DD. Cultured SCN slices from Per1-luc transgenic Cacna2d3−/− mice revealed reduced synchrony of Per1-luc gene expression rhythms among SCN neurons. These findings suggest that α2δ3 is essential for synchronized cellular oscillations in the SCN and thereby contributes to enhancing the sustainability of circadian rhythms in behavior.

Adenosine Diphosphate and the P2Y13 Receptor Are Involved in the Autophagic Protection of Ex Vivo Perfused Livers From Fasted Rats: Potential Benefit for Liver Graft Preservation.

Studies on how to protect livers perfused ex vivo can help design strategies for hepatoprotection and liver graft preservation. The protection of livers isolated from 24-hour versus 18-hour starved rats has been previously attributed to autophagy, which contributes to the energy-mobilizing capacity ex vivo. Here, we explored the signaling pathways responsible for this protection. In our experimental models, 3 major signaling candidates were considered in view of their abilities to trigger autophagy: high mobility group box 1 (HMGB1), adenosine monophosphate-activated protein kinase (AMPK), and purinergic receptor P2Y13. To this end, ex vivo livers isolated from starved rats were perfused for 135minutes, after which perfusate samples were studied for protein release and biopsies were performed for evaluating signaling protein contents. For HMGB1, no significant difference was observed between livers isolated from rats starved for 18 and 24hours at perfusion times of both 0 and 135minutes. The phosphorylated and total forms of AMPK, but not their ratios, were significantly higher in 24-hour fasted than in 18-hour fasted livers. However, although the level of phosphorylated AMPK increased, perfusing ex vivo 18-hour fasted livers with 1mM 5-aminoimidazole-4-carboxamide ribonucleotide, an AMPK activator, did not protect the livers. In addition, the adenosine diphosphate (ADP; and not adenosine monophosphate [AMP]) to AMP + ADP + adenosine triphosphate ratio increased in the 24-hour starved livers compared with that in the 18-hour starved livers. Moreover, perfusing 24-hour starved livers with 0.1mM 2-[(2-chloro-5-nitrophenyl)azo]-5-hydroxy-6-methyl-3-[(phosphonooxy)methyl]-4-pyridinecarboxaldehyde (MRS2211), a specific antagonist of the P2Y13 receptor, induced an increase in cytolysis marker levels in the perfusate samples and a decrease in the levels of autophagic marker microtubule-associated proteins 1 light chain 3 II (LC3II)/actin (and a loss of p62/actin decrease), indicating autophagy inhibition and a loss of protection. The P2Y13 receptor and ADP (a physiological activator of this receptor) are involved in the protection of ex vivo livers. Therapeutic opportunities for improving liver graft preservation through the stimulation of the ADP/P2Y13 receptor axis are further discussed.

Abstract PD9-04: Identification of biomarkers associated with therapeutic resistance: Quantitative protein/phosphoprotein analysis of ~750 patients across 8 arms of the neoadjuvant I-SPY 2 TRIAL for high-risk early stage breast cancer

Abstract Background: The goal of I-SPY 2 is to rapidly test novel therapies in addition to standard of care in high-risk breast cancer patients. It has resulted in increasing response rates, where pCR rates in TNBC and HR-HER2+ subsets have reached ~60% and ~75%, respectively. Yet, there remains a sizeable subset of non-responders, especially among HR+ patients. Identification of ‘universal’ resistance mechanisms may guide rational selection of agents to improve these patient's outcomes. Thus, we analyzed reverse phase protein array (RPPA) based quantitative protein/phosphoprotein data across arms to assess whether there are common mechanisms rendering these cancers resistant to all agent classes tested to date. Methods: 736 patients (260 HR+HER2-, 252 TN, 142 HR+HER2-, and 82 HR-HER2+; over 8 arms: 194 Ctr, 105 neratinib (N), 63 veliparib/carboplatin (VC), 128 AMG386 (anti-ANG1/2), 87 MK2206 (anti-AKT), 43 TH/pertuzumab (P), 49 TDM1/P, and 67 pembrolizumab (Pembro)) with pCR and RPPA data at the pre-treatment time point were considered for this analysis. 141 RPPA endpoints representing key cancer pathways were assessed for association with pCR using logistic regression modeling, with HR, HER2 and treatment arm as covariates (likelihood ratio test; p<0.05). Analysis was also performed in HR/HER2 subsets and within treatment arms. Markers were analyzed individually; multiple comparison correction (Benjamini-Hochberg) was applied to p-values. Our analysis is exploratory, and does not adjust for other biomarkers outside this study. Results: Prior to FDR correction, high levels of Cyclin D1, a cell cycle protein implicated in estrogen-mediated DNA damage repair, associate with non-pCR in the population as a whole and within all subtypes except for the HR-HER2+ subset; an association that retains significance after FDR correction overall as well as in HER2- and HR+HER2- subsets. Within individual arms, high Cyclin D1 predicted non-response in VC, control, and AMG386; and trends toward association in Pembro and N. In addition, high quantitative ER and phospho-androgen receptor (pAR; S650) associate with non-pCR in the population as a whole and in the HR+HER2- subset. For both ER and pAR the strongest association with non-pCR was in the Pembro arm. Candidates for universal sensitivity signals include immune proteins JAK-STAT (pSTAT5 (Y694) and pSTAT1 (Y701)) overall; and pERBB2/pEGFR for HER2+ patients. Conclusions: High levels of Cyclin D1, but not other cell cycle proteins, predict non-response to chemo-/targeted-therapy across arms and subtypes, suggesting that agents specifically targeting Cyclin D1 may increase chemo-sensitivity. ER/phospho-AR as global resistance signals suggest inclusion of anti-AR agents in combination therapy, and the need for new endocrine-based approaches. Citation Format: Julia Wulfkuhle, Denise Wolf, Christina Yau, Lamorna Brown-Swigart, Rosa I Gallagher, Gillian Hirst, Laura Sit, Smita Asare, I-SPY 2 TRIAL Investigators, Nola Hylton, Angela DeMichele, Douglas Yee, Jo Chien, Hope Rugo, John Park, Kathy Albain, Rita Nanda, Debu Tripathy, Richard Schwab, Don Berry, Laura Esserman, Laura van t' Veer, Emanual Petricoin, III. Identification of biomarkers associated with therapeutic resistance: Quantitative protein/phosphoprotein analysis of ~750 patients across 8 arms of the neoadjuvant I-SPY 2 TRIAL for high-risk early stage breast cancer [abstract]. In: Proceedings of the 2020 San Antonio Breast Cancer Virtual Symposium; 2020 Dec 8-11; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2021;81(4 Suppl):Abstract nr PD9-04.

Cardiac Gq Receptors and Calcineurin Activation Are Not Required for the Hypertrophic Response to Mechanical Left Ventricular Pressure Overload.

RationaleGq-coupled receptors are thought to play a critical role in the induction of left ventricular hypertrophy (LVH) secondary to pressure overload, although mechano-sensitive channel activation by a variety of mechanisms has also been proposed, and the relative importance of calcineurin- and calmodulin kinase II (CaMKII)-dependent hypertrophic pathways remains controversial.ObjectiveTo determine the mechanisms regulating the induction of LVH in response to mechanical pressure overload.Methods and ResultsTransgenic mice with cardiac-targeted inhibition of Gq-coupled receptors (GqI mice) and their non-transgenic littermates (NTL) were subjected to neurohumoral stimulation (continuous, subcutaneous angiotensin II (AngII) infusion for 14 days) or mechanical pressure overload (transverse aortic arch constriction (TAC) for 21 days) to induce LVH. Candidate signaling pathway activation was examined. As expected, LVH observed in NTL mice with AngII infusion was attenuated in heterozygous (GqI+/–) mice and absent in homozygous (GqI–/–) mice. In contrast, LVH due to TAC was unaltered by either heterozygous or homozygous Gq inhibition. Gene expression of atrial natriuretic peptide (ANP), B-type natriuretic peptide (BNP) and α-skeletal actin (α-SA) was increased 48 h after AngII infusion or TAC in NTL mice; in GqI mice, the increases in ANP, BNP and α-SA in response to AngII were completely absent, as expected, but all three increased after TAC. Increased nuclear translocation of nuclear factor of activated T-cells c4 (NFATc4), indicating calcineurin pathway activation, occurred in NTL mice with AngII infusion but not TAC, and was prevented in GqI mice infused with AngII. Nuclear and cytoplasmic CaMKIIδ levels increased in both NTL and GqI mice after TAC but not AngII infusion, with increased cytoplasmic phospho- and total histone deacetylase 4 (HDAC4) and increased nuclear myocyte enhancer factor 2 (MEF2) levels.ConclusionCardiac Gq receptors and calcineurin activation are required for neurohumorally mediated LVH but not for LVH induced by mechanical pressure overload (TAC). Rather, TAC-induced LVH is associated with activation of the CaMKII-HDAC4-MEF2 pathway.

The HARPS search for southern extra-solar planets – XLV. Two Neptune mass planets orbiting HD 13808: a study of stellar activity modelling’s impact on planet detection

ABSTRACT We present a comprehensive analysis of 10 yr of HARPS radial velocities (RVs) of the K2V dwarf star HD 13808, which has previously been reported to host two unconfirmed planet candidates. We use the state-of-the-art nested sampling algorithm PolyChord to compare a wide variety of stellar activity models, including simple models exploiting linear correlations between RVs and stellar activity indicators, harmonic models for the activity signals, and a more sophisticated Gaussian process regression model. We show that the use of overly simplistic stellar activity models that are not well-motivated physically can lead to spurious ‘detections’ of planetary signals that are almost certainly not real. We also reveal some difficulties inherent in parameter and model inference in cases where multiple planetary signals may be present. Our study thus underlines the importance both of exploring a variety of competing models and of understanding the limitations and precision settings of one’s sampling algorithm. We also show that at least in the case of HD 13808, we always arrive at consistent conclusions about two particular signals present in the RV, regardless of the stellar activity model we adopt; these two signals correspond to the previously reported though unconfirmed planet candidate signals. Given the robustness and precision with which we can characterize these two signals, we deem them secure planet detections. In particular, we find two planets orbiting HD 13808 at distances of 0.11, 0.26 au with periods of 14.2, 53.8 d, and minimum masses of 11, 10 M⊕.

Characterization of Copy-Number Variations and Possible Candidate Genes in Recurrent Pregnancy Losses.

It is well established that embryonic chromosomal abnormalities (both in the number of chromosomes and the structure) account for 50% of early pregnancy losses. However, little is known regarding the potential differences in the incidence and distribution of chromosomal abnormalities between patients with sporadic abortion (SA) and recurrent pregnancy loss (RPL), let alone the role of submicroscopic copy-number variations (CNVs) in these cases. The aim of the present study was to systematically evaluate the role of embryonic chromosomal abnormalities and CNVs in the etiology of RPL compared with SA. Over a 3-year period, 1556 fresh products of conception (POCs) from miscarriage specimens were investigated using single nucleotide polymorphism array (SNP-array) and CNV sequencing (CNV-seq) in this study, along with further functional enrichment analysis. Chromosomal abnormalities were identified in 57.52% (895/1556) of all cases. Comparisons of the incidence and distributions of chromosomal abnormalities within the SA group and RPL group and within the different age groups were performed. Moreover, 346 CNVs in 173 cases were identified, including 272 duplications, 2 deletions and 72 duplications along with deletions. Duplications in 16q24.3 and 16p13.3 were significantly more frequent in RPL cases, and thereby considered to be associated with RPL. There were 213 genes and 131 signaling pathways identified as potential RPL candidate genes and signaling pathways, respectively, which were centered primarily on six functional categories. The results of the present study may improve our understanding of the etiologies of RPL and assist in the establishment of a population-based diagnostic panel of genetic markers for screening RPL amongst Chinese women.

A Search for Technosignatures around 31 Sun-like Stars with the Green Bank Telescope at 1.15–1.73 GHz

Abstract We conducted a search for technosignatures in 2018 and 2019 April with the L-band receiver (1.15–1.73 GHz) of the 100 m diameter Green Bank Telescope. These observations focused on regions surrounding 31 Sun-like stars near the plane of the Galaxy. We present the results of our search for narrowband signals in this data set, as well as improvements to our data processing pipeline. Specifically, we applied an improved candidate signal detection procedure that relies on the topographic prominence of the signal power, which nearly doubles the signal detection count of some previously analyzed data sets. We also improved the direction-of-origin filters that remove most radio frequency interference (RFI) to ensure that they uniquely link signals observed in separate scans. We performed a preliminary signal injection and recovery analysis to test the performance of our pipeline. We found that our pipeline recovers 93% of the injected signals over the usable frequency range of the receiver and 98% if we exclude regions with dense RFI. In this analysis, 99.73% of the recovered signals were correctly classified as technosignature candidates. Our improved data processing pipeline classified over 99.84% of the ∼26 million signals detected in our data as RFI. Of the remaining candidates, 4539 were detected outside of known RFI frequency regions. The remaining candidates were visually inspected and verified to be of anthropogenic nature. Our search compares favorably to other recent searches in terms of end-to-end sensitivity, frequency drift rate coverage, and signal detection count per unit bandwidth per unit integration time.

Altered miRNA expression profiling in enamel organ of fluoride affected rat embryos

Evidence has shown that miRNAs could play a role in dental fluorosis, but there is no study has investigated the global expression miRNA profiles of fluoride-exposed enamel organ. In this study, we analysed the differentially expressed (DE) miRNAs between fluoride-treated and control enamel organ for the first time and found several candidate miRNAs and signaling pathways worthy of further research. Thirty Wistar rats were randomly distributed into three groups and exposed to drinking water with different fluoride contents for 10 weeks and during the gestation. The three groups were a control group (distilled water), medium fluoride group (75 mg/L NaF), and high fluoride group (150 mg/L NaF). On the embryonic day 19.5, the mandible was dissected for histological analysis, and the enamel organ of the mandibular first molar tooth germ was collected for miRNA sequencing (miRNA-seq) and quantitative real-time PCR analysis (qRT-PCR). Typical dental fluorosis was observed in the incisors of the prepregnant rats. In addition to the disorganized structure of enamel organ cells, 39 DE miRNAs were identified in the fluoride groups compared with the control group, and good agreement between the miRNA-seq data and qRT-PCR data was found. The functional annotation of the target genes of 39 DE miRNAs showed significant enrichment in metabolic process, cell differentiation, calcium signaling pathway, and mitogen-activated protein kinase(MAPK) signaling pathway terms. This study provides a theoretical reference for an extensive understanding of the mechanism of fluorosis and potential valuable miRNAs as therapeutic targets in fluorosis.

State estimation method using median of multiple candidates for observation signals including outliers

The state estimation problem for systems in which observation outputs include outliers is addressed herein. When the observation output has outliers, the accuracy of the state estimation is dramati...

Global Earthquake Forecasting System (GEFS): The challenges ahead

We conclude this special issue on the Global Earthquake Forecasting System (GEFS) by briefly reviewing and analyzing the claims of non-seismic precursors made in the present volume, and by reflecting on the current limitations and future directions to take. We find that most studies presented in this special volume, taken individually, do not provide strong enough evidence of non-seismic precursors to large earthquakes. The majority of the presented results are hampered by the fact that the task at hand is susceptible to potential biases in data selection and possible overfitting. The most encouraging results are obtained for ground-based geoelectric signals, although the probability gain is likely small compared to an earthquake clustering baseline. The only systematic search on satellite data available so far, those of the DEMETER mission, did not find a robust precursory pattern. The conclusion that we can draw is that the overall absence of convincing evidence is likely due to a deficit in systematically applying robust statistical methods and in integrating scientific knowledge of different fields. Most authors are specialists of their field while the study of earthquake precursors requires a system approach combined with the knowledge of many specific characteristics of seismicity. Relating non-seismic precursors to earthquakes remains a challenging multidisciplinary field of investigation. The plausibility of these precursors predicted by models of lithosphere-atmosphere-ionosphere coupling, together with the suggestive evidence collected here, call for further investigations. The primary goal of the GEFS is thus to build a global database of candidate signals, which could potentially improve earthquake predictability (if the weak signals observed are real and false positives sufficiently uncorrelated between different data sources). Such a stacking of disparate and voluminous data will require big data storage and machine learning pipelines, which has become feasible only recently. This special issue compiled an eclectic list of non-seismic precursor candidates, which is in itself a valuable source of information for seismologists, geophysicists and other scientists who may not be familiar with such types of investigations. It also forms the foundation for a coherent, multi-disciplinary collaboration on earthquake prediction.

An automated tool to facilitate consistent test-driven development of trigger selections for LHCb’s Run 3

Upon its restart in 2022, the LHCb experiment at the LHC will run at higher instantaneous luminosity and utilize an unprecedented full-software trigger, promising greater physics reach and efficiency. On the flip side, conforming to offline data storage constraints becomes far more challenging. Both of these considerations necessitate a set of highly optimised trigger selections. We therefore present HltEfficiencyChecker: an automated extension to the LHCb trigger application, facilitating trigger development before data-taking driven by trigger rates and efficiencies. Since the default in 2022 will be to persist only the event’s signal candidate to disk, discarding the rest of the event, we also compute efficiencies where the decision was due to the true MC signal, evaluated by matching it to the trigger candidate hit-by-hit. This matching procedure – which we validate here – demonstrates that the distinction between a “trigger” and a “trigger-on-signal” is crucial in characterising the performance of a trigger selection.

Genome-Wide Identification of Populus Malectin/Malectin-Like Domain-Containing Proteins and Expression Analyses Reveal Novel Candidates for Signaling and Regulation of Wood Development.

Malectin domain (MD) is a ligand-binding protein motif of pro- and eukaryotes. It is particularly abundant in Viridiplantae, where it occurs as either a single (MD, PF11721) or tandemly duplicated domain (PF12819) called malectin-like domain (MLD). In herbaceous plants, MD- or MLD-containing proteins (MD proteins) are known to regulate development, reproduction, and resistance to various stresses. However, their functions in woody plants have not yet been studied. To unravel their potential role in wood development, we carried out genome-wide identification of MD proteins in the model tree species black cottonwood (Populus trichocarpa), and analyzed their expression and co-expression networks. P. trichocarpa had 146 MD genes assigned to 14 different clades, two of which were specific to the genus Populus. 87% of these genes were located on chromosomes, the rest being associated with scaffolds. Based on their protein domain organization, and in agreement with the exon-intron structures, the MD genes identified here could be classified into five superclades having the following domains: leucine-rich repeat (LRR)-MD-protein kinase (PK), MLD-LRR-PK, MLD-PK (CrRLK1L), MLD-LRR, and MD-Kinesin. Whereas the majority of MD genes were highly expressed in leaves, particularly under stress conditions, eighteen showed a peak of expression during secondary wall formation in the xylem and their co-expression networks suggested signaling functions in cell wall integrity, pathogen-associated molecular patterns, calcium, ROS, and hormone pathways. Thus, P. trichocarpa MD genes having different domain organizations comprise many genes with putative foliar defense functions, some of which could be specific to Populus and related species, as well as genes with potential involvement in signaling pathways in other tissues including developing wood.

Indole-3-acetic acid synthesized through the indole-3-pyruvate pathway promotes Candida tropicalis biofilm formation.

We previously found that the elevated abundance of the fungus Candida tropicalis is positively correlated with the bacteria Escherichia coli and Serratia marcescens in Crohn's disease patients and the three pathogens, when co-cultured, form a robust mixed-species biofilm. The finding suggests that these three pathogens communicate and promote biofilm formation, possibly through secretion of small signaling molecules. To identify candidate signaling molecules, we carried out a metabolomic analysis of the single-species and triple-species cultures of the three pathogens. This analysis identified 15 metabolites that were highly increased in the triple-species culture. One highly induced metabolite was indole-3-acetic acid (IAA), which has been shown to induce filamentation of certain fungi. We thus tested the effect of IAA on biofilm formation of C. tropicalis and demonstrated that IAA promotes biofilm formation of C. tropicalis. Then, we carried out isotope tracing experiments using 13C-labeled-tryptophan as a precursor to uncover the biosynthesis pathway of IAA in C. tropicalis. The results indicated that C. tropicalis synthesizes IAA through the indole-3-pyruvate pathway. Further studies using inhibitors of the indole-3-pyruvate pathway are warranted to decipher the mechanisms by which IAA influences biofilm formation.

An investigation of the very rare {K}^{+}to {pi}^{+}nu overline{nu} decay

The NA62 experiment reports an investigation of the {K}^{+}to {pi}^{+}nu overline{nu} mode from a sample of K+ decays collected in 2017 at the CERN SPS. The experiment has achieved a single event sensitivity of (0.389 ± 0.024) × 10−10, corresponding to 2.2 events assuming the Standard Model branching ratio of (8.4 ± 1.0) × 10−11. Two signal candidates are observed with an expected background of 1.5 events. Combined with the result of a similar analysis conducted by NA62 on a smaller data set recorded in 2016, the collaboration now reports an upper limit of 1.78 × 10−10 for the {K}^{+}to {pi}^{+}nu overline{nu} branching ratio at 90% CL. This, together with the corresponding 68% CL measurement of ( {0.48}_{-0.48}^{+0.72} ) × 10−10, are currently the most precise results worldwide, and are able to constrain some New Physics models that predict large enhancements still allowed by previous measurements.

Low‐complexity PTS schemes for PAPR reduction in OFDM systems

For original partial transmit sequence (O-PTS), the conjugate operations can be applied to achieve the purpose of ameliorating the peak-to-average power ratio (PAPR) performance. But in this way, an observation has been found that some of these candidate signals has the same PAPR value that brings about great redundant computations. Based on this, a low-complexity PTS is put forward to abate the PAPR of orthogonal frequency division multiplexing (OFDM) systems, thereby avoiding redundant candidate signals. In proposed scheme, a novel scheme employing conjugate operations for generating candidate signals with different PAPR values is given, which avoids the generation of candidate signals with the same PAPR value and saves the needless computations. Meanwhile, the grouping is involved for further reduction in computational complexity. Simulation results illustrate that our scheme has the similar or superior PAPR performance with the remarkable reduction in computational complexity by comparison with O-PTS.

Reflecting Modulation

Reconfigurable intelligent surface (RIS) has emerged as a promising technique for future wireless communication networks. How to reliably transmit information in a RIS-based communication system arouses much interest. This paper proposes a reflecting modulation (RM) scheme for RIS-based communications, where both the reflecting patterns and transmit signals can carry information. Depending on that the transmitter and RIS jointly or independently deliver information, RM is further classified into two categories: jointly mapped RM (JRM) and separately mapped RM (SRM). JRM and SRM are naturally superior to existing schemes, because the transmit signal vectors, reflecting patterns, and bit mapping methods of JRM and SRM are more flexibly designed. To enhance transmission reliability, this paper proposes a discrete optimization-based joint signal mapping, shaping, and reflecting (DJMSR) design for JRM and SRM to minimize the bit error rate (BER) with a given transmit signal candidate set and a given reflecting pattern candidate set. To further improve the performance, this paper optimizes multiple reflecting patterns and their associated transmit signal sets in continuous fields for JRM and SRM. Numerical results show that JRM and SRM with the proposed system optimization methods considerably outperform existing schemes in BER.

Identification of Potential Biomarkers of Polycystic Ovary Syndrome via Integrated Bioinformatics Analysis.

Polycystic ovary syndrome (PCOS) is a life-long reproductive, endocrine, and metabolic disorder that affects up to 17% of women of reproductive age. However, the effect of granulosa cells (GCs) transcriptome changes on oocyte capacity and follicular development in patients with PCOS has not been elucidated. This study aims to analyze transcriptome changes in GCs of PCOS from different perspectives and explore potential biomarkers for the diagnosis and treatment of PCOS. The gene expression profiles of GSE34526 and GSE107746 were obtained from the GEO database. Differentially expressed genes (DEGs) and key signaling pathways were identified. Gene Set Enrichment Analysis (GSEA) revealed that Toll-like receptors, NOD-like receptors, and NOTCH signaling pathways were obviously enriched in GCs of PCOS. We further analyzed DEGs from three aspects: transcription factors (TFs), secreted proteins, and follicular development. Compared with normal GCs, the DEGs encoding TFs and secretory proteins in GCs of PCOS remarkably changed. Besides, HAS2 and CBLN1, which are highly expressed in preovulatory follicular GCs and may trigger ovulation, were significantly decreased in GCs of PCOS. This study found candidate genes and signaling pathways in PCOS, providing new insights and foundations for the etiology of PCOS. Besides, HSA2 and CBLN1 may be potential therapeutic biomarkers for ovulation disorders in PCOS.

Gravitational wave probes of dark matter: challenges and opportunities

In this white paper, we discuss the prospects for characterizing and identifying dark matter using gravitational waves, covering a wide range of dark matter candidate types and signals. We argue that present and upcoming gravitational wave probes offer unprecedented opportunities for unraveling the nature of dark matter and we identify the most urgent challenges and open problems with the aim of encouraging a strong community effort at the interface between these two exciting fields of research.

In Silico Investigation of Signal Peptide Sequences to Enhance Secretion of CD44 Nanobodies Expressed in Escherichia coli.

The selection of a suitable signal peptide that can direct recombinant proteins from the cytoplasm to the extracellular space is an important criterion affecting the production of recombinant proteins in Escherichia coli, a widely used host. Nanobodies are currently attracting the attention of scientists as antibody alternatives due to their specific properties and feasibility of production in E. coli. CD44 nanobodies constitute a potent therapeutic agent that can block CD44/HA interaction in cancer and inflammatory diseases. This molecule may also function as a drug against cancer cells and has been produced previously in E. coli without a signal peptide sequence. The goal of this project was to find a suitable signal peptide to direct CD44 nanobody extracellular secretion in E. coli that will potentially lead to optimization of experimental methods and facilitate downstream steps such as purification. We analyzed 40 E. coli derived signal peptides retrieved from the Signal Peptide database and selected the best candidate signal peptides according to relevant criteria including signal peptide probability, stability, and physicochemical features, which were evaluated using signalP software version 4.1 and the ProtParam tool, respectively. In this in silico study, suitable candidate signal peptide(s) for CD44 nanobody secretory expression were identified. CSGA, TRBC, YTFQ, NIKA, and DGAL were selected as appropriate signal peptides with acceptable D-scores, and appropriate physicochemical and structural properties. Following further analysis, TRBC was selected as the best signal peptide to direct CD44 nanobody expression to the extracellular space of E. coli. The selected signal peptide, TRBC is the most suitable to promote high-level secretory production of CD44 nanobodies in E. coli and potentially will be useful for scaling up CD44 nanobody production in experimental research as well as in other CD44 nanobody applications. However, experimental work is needed to confirm the data.

The ex planta signal activity of a Medicago ribosomal uL2 protein suggests a moonlighting role in controlling secondary rhizobial infection.

We recently described a regulatory loop, which we termed autoregulation of infection (AOI), by which Sinorhizobium meliloti, a Medicago endosymbiont, downregulates the root susceptibility to secondary infection events via ethylene. AOI is initially triggered by so-far unidentified Medicago nodule signals named signal 1 and signal 1' whose transduction in bacteroids requires the S. meliloti outer-membrane-associated NsrA receptor protein and the cognate inner-membrane-associated adenylate cyclases, CyaK and CyaD1/D2, respectively. Here, we report on advances in signal 1 identification. Signal 1 activity is widespread as we robustly detected it in Medicago nodule extracts as well as in yeast and bacteria cell extracts. Biochemical analyses indicated a peptidic nature for signal 1 and, together with proteomic analyses, a universally conserved Medicago ribosomal protein of the uL2 family was identified as a candidate signal 1. Specifically, MtRPuL2A (MtrunA17Chr7g0247311) displays a strong signal activity that requires S. meliloti NsrA and CyaK, as endogenous signal 1. We have shown that MtRPuL2A is active in signaling only in a non-ribosomal form. A Medicago truncatula mutant in the major symbiotic transcriptional regulator MtNF-YA1 lacked most signal 1 activity, suggesting that signal 1 is under developmental control. Altogether, our results point to the MtRPuL2A ribosomal protein as the candidate for signal 1. Based on the Mtnf-ya1 mutant, we suggest a link between root infectiveness and nodule development. We discuss our findings in the context of ribosomal protein moonlighting.