GenoMycAnalyzer: a web-based tool for species and drug resistance prediction for Mycobacterium genomes.

BACKGROUND: Drug-resistant tuberculosis (TB) is a major threat to global public health. Whole-genome sequencing (WGS) is a useful tool for species identification and drug resistance prediction, and many clinical laboratories are transitioning to WGS as a routine diagnostic tool. However, user-friendly and high-confidence automated bioinformatics tools are needed to rapidly identify M. tuberculosis complex (MTBC) and non-tuberculous mycobacteria (NTM), detect drug resistance, and further guide treatment options. RESULTS: We developed GenoMycAnalyzer, a web-based software that integrates functions for identifying MTBC and NTM species, lineage and spoligotype prediction, variant calling, annotation, drug-resistance determination, and data visualization. The accuracy of GenoMycAnalyzer for genotypic drug susceptibility testing (gDST) was evaluated using 5,473 MTBC isolates that underwent phenotypic DST (pDST). The GenoMycAnalyzer database was built to predict the gDST for 15 antituberculosis drugs using the World Health Organization mutational catalogue. Compared to pDST, the sensitivity of drug susceptibilities by the GenoMycAnalyzer for first-line drugs ranged from 95.9% for rifampicin (95% CI 94.8-96.7%) to 79.6% for pyrazinamide (95% CI 76.9-82.2%), whereas those for second-line drugs ranged from 98.2% for levofloxacin (95% CI 90.1-100.0%) to 74.9% for capreomycin (95% CI 69.3-80.0%). Notably, the integration of large deletions of the four resistance-conferring genes increased gDST sensitivity. The specificity of drug susceptibilities by the GenoMycAnalyzer ranged from 98.7% for amikacin (95% CI 97.8-99.3%) to 79.5% for ethionamide (95% CI 76.4-82.3%). The incorporated Kraken2 software identified 1,284 mycobacterial species with an accuracy of 98.8%. GenoMycAnalyzer also perfectly predicted lineages for 1,935 MTBC and spoligotypes for 54 MTBC. CONCLUSIONS: GenoMycAnalyzer offers both web-based and graphical user interfaces, which can help biologists with limited access to high-performance computing systems or limited bioinformatics skills. By streamlining the interpretation of WGS data, the GenoMycAnalyzer has the potential to significantly impact TB management and contribute to global efforts to combat this infectious disease. GenoMycAnalyzer is available at http://www.mycochase.org .

Prevalence and Molecular Characterization of Vancomycin Variable Enterococcus faecium Isolated From Clinical Specimens.

Vancomycin variable Enterococcus (VVE) bacteria are phenotypically susceptible to vancomycin, but they harbor the vanA gene. We aimed to ascertain the prevalence of VVE among clinically isolated vancomycin-susceptible Enterococcus faecium (VSE) isolates, as well as elucidate the molecular characteristics of the vanA gene cluster within these isolates. Notably, we investigated the prevalence and structure of the vanA gene cluster of VVE. Between June 2021 and May 2022, we collected consecutive, non-duplicated vancomycin-susceptible Enterococcus faecium (VSE) samples. Real-time PCR was performed to detect the presence of vanA, vanB, and vanC. Overlapping PCR with sequencing and whole-genome sequencing were performed for structural analysis. Sequence types (STs) were determined by multilocus sequence typing. Exposure testing was performed to assess the ability of the isolates to acquire vancomycin resistance. Among 282 VSE isolates tested, 20 isolates (7.1%) were VVE. Among them, 17 isolates had partial deletions in the IS1216 or IS1542 sequences in vanS (N=10), vanR (N=5), or vanH (N=2). All VVE isolates belonged to the CC17 complex and comprised five STs, namely ST17 (40.0%), ST1421 (25.0%), ST80 (25.0%), ST787 (5.0%), and ST981 (5.0%). Most isolates were related to three hospital-associated clones (ST17, ST1421, and ST80). After vancomycin exposure, 18 of the 20 VVEs acquired vancomycin resistance. Considering the high reversion rate, detecting VVE by screening VSE for vanA is critical for appropriate treatment and infection control.

Spatial architectures of somatic mutations in normal prostate, benign prostatic hyperplasia and coexisting prostate cancer.

This study aimed to identify somatic mutations in nontumor cells (NSMs) in normal prostate and benign prostatic hyperplasia (BPH) and to determine their relatedness to prostate cancer (PCA). From 22 PCA patients, two prostates were sampled for 3-dimensional mapping (50 normal, 46 BPH and 1 PCA samples), and 20 prostates were trio-sampled (two normal or BPH samples and one PCA sample) and analyzed by whole-genome sequencing. Normal and BPH tissues harbored several driver NSMs and copy number alterations (CNAs), including in FOXA1, but the variations exhibited low incidence, rare recurrence, and rare overlap with PCAs. CNAs, structural variants, and mutation signatures were similar between normal and BPH samples, while BPHs harbored a higher mutation burden, shorter telomere length, larger clone size, and more private NSMs than normal prostates. We identified peripheral-zonal dominance and right-side asymmetry in NSMs, but the asymmetry was heterogeneous between samples. In one normal prostate, private oncogenic RAS-signaling NSMs were detected, suggesting convergence in clonal maintenance. Early embryonic mutations exhibited two distinct distributions, characterized as layered and mixed patterns. Our study identified that the BPH genome differed from the normal prostate genome but was still closer to the normal genome than to the PCA genome, suggesting that BPH might be more related to aging or environmental stress than to tumorigenic processes.

No Interference of H9 Extract on Trastuzumab Pharmacokinetics in Their Combinations.

Trastuzumab is used to treat breast cancer patients overexpressing human epidermal growth factor receptor 2, but resistance and toxicity limit its uses, leading to attention to trastuzumab combinations. Recently, the synergistic effect of trastuzumab and H9 extract (H9) combination against breast cancer has been reported. Because drug exposure determines its efficacy and toxicity, the question of whether H9 changes trastuzumab exposure in the body has been raised. Therefore, this study aimed to characterize trastuzumab pharmacokinetics and elucidate the effect of H9 on trastuzumab pharmacokinetics at a combination dose that shows synergism in mice. As a result, trastuzumab showed linear pharmacokinetics after its intravenous administration from 1 to 10 mg/kg. In the combination of trastuzumab and H9, single and 2-week treatments of oral H9 (500 mg/kg) did not influence trastuzumab pharmacokinetics. In the multiple-combination treatments of trastuzumab and H9 showing their synergistic effect (3 weeks of trastuzumab with 2 weeks of H9), the pharmacokinetic profile of trastuzumab was comparable to that of 3 weeks of trastuzumab alone. In tissue distribution, the tissue to plasma ratios of trastuzumab below 1.0 indicated its limited distributions within the tissues, and these patterns were unaffected by H9. These results suggest that the systemic and local exposures of trastuzumab are unchanged by single and multiple-combination treatments of H9.

A Novel Anticancer Peptide Derived from Bryopsis plumosa Regulates Proliferation and Invasion in Non-Small Cell Lung Cancer Cells.

The discovery of new highly effective anticancer drugs with few side effects is a challenge for drug development research. Natural or synthetic anticancer peptides (ACPs) represent a new generation of anticancer agents with high selectivity and specificity. The rapid emergence of chemoradiation-resistant lung cancer has necessitated the discovery of novel anticancer agents as alternatives to conventional therapeutics. In this study, we synthesized a peptide containing 22 amino acids and characterized it as a novel ACP (MP06) derived from green sea algae, Bryopsis plumosa. Using the ACP database, MP06 was predicted to possess an alpha-helical secondary structure and functionality. The anti-proliferative and apoptotic effects of the MP06, determined using the cytotoxicity assay and Annexin V/propidium iodide staining kit, were significantly higher in non-small-cell lung cancer (NSCLC) cells than in non-cancerous lung cells. We confirmed that MP06 suppressed cellular migration and invasion and inhibited the expression of N-cadherin and vimentin, the markers of epithelial-mesenchymal transition. Moreover, MP06 effectively reduced the metastasis of tumor xenografts in zebrafish embryos. In conclusion, we suggest considering MP06 as a novel candidate for the development of new anticancer drugs functioning via the ERK signaling pathway.

Monitoring the Outcomes of Systemic Chemotherapy Including Immune Checkpoint Inhibitor for HER2-Positive Metastatic Gastric Cancer by Liquid Biopsy.

PURPOSE: For precision medicine, exploration and monitoring of molecular biomarkers are essential. However, in advanced gastric cancer (GC) with visceral lesions, an invasive procedure cannot be performed repeatedly for the follow-up of molecular biomarkers. MATERIALS AND METHODS: To verify the clinical implication of serial liquid biopsies targeting circulating tumor DNA (ctDNA) on treatment response, we conducted targeted deep sequencing for serially collected ctDNA of 15 HER2-positive metastatic GC patients treated with anti-PD-1 inhibitor in combination with standard systemic treatment. RESULTS: In the baseline ctDNAs, 14 patients (93%) harbored more than one genetic alteration. A number of mutations in well-known cancer-related genes, such as KRAS and PIK3CA, were identified. Copy number alterations were identified in eight GCs (53.3%), and amplification of the ERBB2 gene (6/15, 40.0%) was the most recurrent. When we calculated the mean variant allele frequency (VAF) of mutations in each ctDNA as the molecular tumor burden index (mTBI), the mTBI trend was largely consistent with the VAF profiles in both responder and non-responder groups. Notably, in the longitudinal analysis of ctDNA, mTBI provided 2-42 weeks (mean 13.4 weeks) lead time in the detection of disease progression compared to conventional follow-up with CT imaging. CONCLUSION: Our data indicate that the serial genetic alteration profiling of ctDNA is feasible to predict treatment response in HER2-positive GC patients in a minimally invasive manner. Practically, ctDNA profiles are useful not only for the molecular diagnosis of GC but also for the selection of GC patients with poor prognosis for systemic treatment (ClinicalTrials.gov identifier: NCT02901301).

pncA Large Deletion is the Characteristic of Pyrazinamide-Resistant Mycobacterium tuberculosis belonging to the East Asian Lineage.

BACKGROUND: Pyrazinamide (PZA) is often used as an add-on agent in the treatment of multidrug-resistant tuberculosis, regardless of phenotypic drug susceptibility testing (pDST) results. However, evaluating the effectiveness of PZA is challenging because of its low pH activity, which can result in unreliable pDST results. This study aimed to investigate the genomic characteristics associated with PZA resistance that can be used to develop genotypic DST. MATERIALS AND METHODS: A publicly available whole genome sequencing (WGS) dataset of 10,725 Mycobacterium tuberculosis complex genomes (3,326 phenotypically PZA-resistant and 7,399 phenotypically PZA-susceptible isolates) were analyzed. RESULTS: In total, 2,934 pncA non-silent mutations were identified in 2,880 isolates (26.9%). Detected mutations were found throughout the entire coding region of pncA in a scattered pattern, of which the most frequent mutation was p.Q10P (n = 278), followed by p.H57D (n = 167) and c.-11A>G (n = 122). The sensitivity and specificity of the group 1 or 2 mutations reported by the World Health Organization (WHO) mutational catalogue were 73.0% and 98.9%, respectively. We further identified 18 novel pncA mutations that were significantly associated with phenotypically PZA-resistant. In addition to these mutations, we identified 102 large deletions in the pncA gene, and all but two isolates were phenotypically resistant to PZA isolates. Notably, pncA deletions were mutually exclusive to pncA mutations, and more than half of the isolates with pncA large deletions belonged to the East Asian lineage (67.6%). The sensitivity, specificity, positive predictive value, and negative predictive value of the pooled variants (group 1 or 2 mutations, novel resistance-associated mutations, and large deletions of the pncA gene) were 79.0%, 98.9%, 97.0%, and 91.3%, respectively. The area under the curve (AUC) value for the pooled variants was significantly higher than the AUC value for the group 1 or 2 mutations (P <0.001), indicating that the pooled variants have a better discriminative ability for predicting PZA resistance. CONCLUSION: Using WGS, we found that the pncA mutations are scattered without specific mutational hotspots, and large deletions associated with PZA resistance are more common in the East Asian lineage of M. tuberculosis isolates. Our data also demonstrated the reliability of group 1 or 2 mutations presented in the WHO mutation catalogue and the need for further investigation on group 3 mutations, contributing to the evaluation of the current knowledge base on mutations associated with the PZA-resistant M. tuberculosis complex.

Targeted deep sequencing reveals genomic alterations of actinic keratosis/cutaneous squamous cell carcinoma in situ and cutaneous squamous cell carcinoma.

Actinic keratosis (AK) and cutaneous squamous cell carcinoma in situ (CIS) are two of the most common precursors of cutaneous squamous cell carcinoma (cSCC). However, the genomic landscape of AK/CIS and the drivers of cSCC progression remain to be elucidated. The aim of our study was to investigate the genomic alterations between AK/CIS and cSCC in terms of somatic mutations and copy number alterations (CNAs). We performed targeted deep sequencing of 160 cancer-related genes with a median coverage of 515× for AK (N = 9), CIS (N = 9), cSCC lesions (N = 13), and matched germline controls from 17 patients. cSCC harboured higher abundance of total mutations, driver mutations and CNAs than AK/CIS. Driver mutations were found in TP53 (81%), NOTCH1 (32%), RB1 (26%) and CDKN2A (19%). All AK/CIS and cSCC lesions (93.5%), except two, harboured TP53 or NOTCH1 mutations, some of which were known oncogenic mutations or reported mutations in normal skin. RB1 driver mutations were found in CIS/cSCC (36.4%) but not in AK. CDKN2A driver mutations were found more frequently in cSCC (30.8%) than in AK/CIS (11.1%). Among recurrent (≥3 samples) CNAs (gain in MYC and PIK3CA/SOX2/TP63; loss in CDKN2A and RB1), MYC (8q) gain and CDKN2A (9p) loss were more frequently detected in cSCC (30.8%) than in AK/CIS (11.1%). Ultraviolet was responsible for the majority of somatic mutations in both AK/CIS and cSCC. Our study revealed that AK/CIS lesions harbour prevalent TP53 or NOTCH1 mutations and that additional somatic mutations and CNAs may lead to cSCC progression in AK/CIS lesions.

Generation of ints14 Knockout Zebrafish using CRISPR/Cas9 for the Study of Development and Disease Mechanisms.

INTS14/VWA9, a component of the integrator complex subunits, plays a pivotal role in regulating the fate of numerous nascent RNAs transcribed by RNA polymerase II, particularly in the biogenesis of small nuclear RNAs and enhancer RNAs. Despite its significance, a comprehensive mutation model for developmental research has been lacking. To address this gap, we aimed to investigate the expression patterns of INTS14 during zebrafish embryonic development. We generated ints14 mutant strains using the CRISPR/Cas9 system. We validated the gRNA activity by co-injecting Cas9 protein and a single guide RNA into fertilized zebrafish eggs, subsequently confirming the presence of a 6- or 9-bp deletion in the ints14 gene. In addition, we examined the two mutant alleles through PCR analysis, T7E1 assay, TA-cloning, and sequencing. For the first time, we used the CRISPR/Cas9 system to create a model in which some sequences of the ints14 gene were removed. This breakthrough opens new avenues for in-depth exploration of the role of ints14 in animal diseases. The mutant strains generated in this study can provide a valuable resource for further investigations into the specific consequences of ints14 gene deletion during zebrafish development. This research establishes a foundation for future studies exploring the molecular mechanisms underlying the functions of ints14, its interactions with other genes or proteins, and its broader implications for biological processes.

Effect of Copper Chelators via the TGF-ß Signaling Pathway on Glioblastoma Cell Invasion.

Glioblastoma multiforme (GBM) is a fast-growing and aggressive type of brain cancer. Unlike normal brain cells, GBM cells exhibit epithelial-mesenchymal transition (EMT), which is a crucial biological process in embryonic development and cell metastasis, and are highly invasive. Copper reportedly plays a critical role in the progression of a variety of cancers, including brain, breast, and lung cancers. However, excessive copper is toxic to cells. D-penicillamine (DPA) and triethylenetetramine (TETA) are well-known copper chelators and are the mainstay of treatment for copper-associated diseases. Following treatment with copper sulfate and DPA, GBM cells showed inhibition of proliferation and suppression of EMT properties, including reduced expression levels of N-cadherin, E-cadherin, and Zeb, which are cell markers associated with EMT. In contrast, treatment with copper sulfate and TETA yielded the opposite effects in GBM. Genes, including TGF-ß, are associated with an increase in copper levels, implying their role in EMT. To analyze the invasion and spread of GBM, we used zebrafish embryos xenografted with the GBM cell line U87. The invasion of GBM cells into zebrafish embryos was markedly inhibited by copper treatment with DPA. Our findings suggest that treatment with copper and DPA inhibits proliferation and EMT through a mechanism involving TGF-ß/Smad signaling in GBM. Therefore, DPA, but not TETA, could be used as adjuvant therapy for GBM with high copper concentrations.

Evaluation of a Rapid and Simple Method for Assessing Retinal Vessel Structures in Adult Zebrafish.

The evaluation of retinal vascular structures is important for analyzing various ophthalmic diseases. Conventional trypsin digestion was used for separating retinal vasculatures in mouse, rat, and other animal models; however, the trypsin method alone is technically difficult to perform and has not been reported in zebrafish to date. In this study, we introduced a rapid and convenient method that allows the investigation of fine vessel structures at a cellular level in the relatively intact retinal vasculature of adult zebrafish. Using an anti-ZO-1 antibody, tight junction structures in retinal vessels were examined in detail and several different cell types constituting blood vessels in arterial and capillary areas were identified. In addition, using cell type-specific antibodies, we identified smooth muscle cells, blood cells, and endothelial cells in the retinal vasculature. Finally, using the hyperglycemic model, we observed the dilation of retinal vessels, the downregulation of tight junction proteins, and the reduction in smooth muscle cells. Based on these results, we provide a rapid and convenient method for the study of retinal vasculature disease in the zebrafish animal model.

Single-cell analysis of multiple myelomas refines the molecular features of bortezomib treatment responsiveness.

Both the tumor and tumor microenvironment (TME) are crucial for pathogenesis and chemotherapy resistance in multiple myeloma (MM). Bortezomib, commonly used for MM treatment, works on both MM and TME cells, but innate and acquired resistance easily develop. By single-cell RNA sequencing (scRNA-seq), we investigated bone marrow aspirates of 18 treatment-naïve MM patients who later received bortezomib-based treatments. Twelve plasma and TME cell types and their subsets were identified. Suboptimal responders (SORs) to bortezomib exhibited higher copy number alteration burdens than optimal responders (ORs). Forty-four differentially expressed genes for SORs based on scRNA-seq data were further analyzed in an independent cohort of 90 treatment-naïve MMs, where 24 genes were validated. A combined model of three clinical variables (older age, low absolute lymphocyte count, and no autologous stem cell transplantation) and 24 genes was associated with bortezomib responsiveness and poor prognosis. In T cells, cytotoxic memory, proliferating, and dysfunctional subsets were significantly enriched in SORs. Moreover, we identified three monocyte subsets associated with bortezomib responsiveness and an MM-specific NK cell trajectory that ended with an MM-specific subset. scRNA-seq predicted the interaction of the GAS6-MERTK, ALCAM-CD6, and BAG6-NCR gene networks. Of note, tumor cells from ORs and SORs were the most prominent sources of ALCAM on effector T cells and BAG6 on NK cells, respectively. Our results indicate that the complicated compositional and molecular changes of both tumor and immune cells in the bone marrow (BM) milieu are important in the development and acquisition of resistance to bortezomib-based treatment of MM.

Differential transcriptome profile underlying risky choice in a rat gambling task.

Background and aims: Proper measurement of expected risk is important for making rational decisions, and maladaptive decision making may underlie various psychiatric disorders. However, differentially expressed genetic profiling involved in this process is still largely unknown. A rodent version of the gambling task (rGT) has been developed to measure decision-making by adopting the same principle of Iowa Gambling Task in humans. In the present study, we examined using next-generation sequencing (NGS) technique whether there are differences in gene expression profiles in the medial prefrontal cortex (mPFC) and the nucleus accumbens (NAc) when rats make different choices toward risk in rGT. Methods: Rats were trained in a touch screen chamber to learn the relationships between 4 different light signals on the window of the screen and accompanied reward outcomes or punishments set up with different magnitudes and probabilities. Once they showed a stabilized pattern of preference upon free choice, rats were classified into risk-averse or risk-seeking groups. After performing the rGT, rats were decapitated, the mPFC and the NAc was dissected out, and NGS was performed with the total RNA extracted. Results: We found that 477 and 36 genes were differentially expressed (approximately 75 and 83% out of them were downregulated) in the mPFC and the NAc, respectively, in risk-seeking compared to risk-averse rats. Among those, we suggested a few top ranked genes that may contribute to promoting risky choices. Discussion and conclusions: Our findings provide insights into transcriptional components underlying risky choices in rats.

Anti-stress Effect of Octopus Cephalotocin in Rats.

Cephalotocin is a bioactivity-regulating peptide expressed in octopus (Octopus vulgaris). The peptide sequence of cephalotocin is very similar to the peptide sequence of mammalian vasopressin, and cephalotocin has been proposed to mainly activate arginine vasopressin 1b receptor (Avpr1b) in the brain. However, the effects of cephalotocin on mammalian behavior have not been studied. In the current study, cephalotocin significantly reduced both the frequency and amplitude of spontaneous excitatory postsynaptic currents (sEPSCs) recorded from not only cultured neuronal cells from postnatal Sprague-Dawley (SD) rats but also hippocampal slices from 4-week-old male C57BL/6 mice. Intraperitoneal (IP) injection did not affect the open field behaviors of C57BL/6 mice. Cephalotocin was directly infused into the hippocampus because the normalized Avpr1b staining intensity divided by the DAPI staining intensity indicated that Avpr1b expression tended to be high in the hippocampus. A hippocampal infusion of 1 mg/kg cephalotocin via an implanted cannula exerted an anti-stress effect, significantly reducing the immobility time in the tail suspension test (TST). The present results provide evidence that the effects of cephalotocin on the activity of hippocampal neurons are related to ameliorating stress, suggesting that cephalotocin may be developed as an anti-stress biomodulator that functions by affecting the brain.

Spatiotemporal dynamics of macrophage heterogeneity and a potential function of Trem2hi macrophages in infarcted hearts.

Heart failure (HF) is a frequent consequence of myocardial infarction (MI). Identification of the precise, time-dependent composition of inflammatory cells may provide clues for the establishment of new biomarkers and therapeutic approaches targeting post-MI HF. Here, we investigate the spatiotemporal dynamics of MI-associated immune cells in a mouse model of MI using spatial transcriptomics and single-cell RNA-sequencing (scRNA-seq). We identify twelve major immune cell populations; their proportions dynamically change after MI. Macrophages are the most abundant population at all-time points (>60%), except for day 1 post-MI. Trajectory inference analysis shows upregulation of Trem2 expression in macrophages during the late phase post-MI. In vivo injection of soluble Trem2 leads to significant functional and structural improvements in infarcted hearts. Our data contribute to a better understanding of MI-driven immune responses and further investigation to determine the regulatory factors of the Trem2 signaling pathway will aid the development of novel therapeutic strategies for post-MI HF.

Single-cell RNA sequencing reveals the existence of pro-metastatic subpopulation within a parental B16 murine melanoma cell line.

The mechanism of melanoma metastasis is poorly understood, especially at the single-cell level. To understand the evolution from primary melanoma to metastasis, we investigated single-cell transcriptome profiles of parental B16 melanoma cells (B16F0) and its highly metastatic subclone (B16F10). Genomic alterations between cells were also analyzed by whole-exome sequencing. We identified 274 differentially expressed genes (DEGs) in B16F10, including upregulated genes related to metastasis, Lgals3, Sparc, Met, and Tmsb4x, and downregulated Mitf pathway genes, Ptgds, Cyb5a, and Cd63. The proportion of cycling cells and cells highly expressing Kdm5b was significantly high in B16F10 cells. Among the five subclusters of B16 cells (C1-5), C3/C4 clusters comprised both B16F0 and B16F10 cells and exhibited intermediate DEG patterns, whereas the C5 cluster mostly comprised B16F10 and showed typical metastatic characteristics. In trajectory analysis, the C4 cluster in B16F0, which showed unique characteristics (mainly cycling cells and upregulation of Mitf pathway genes), have transition potential to the C5 cluster (B16F10). Regarding genomic alterations, stepwise evolution with shared mutations, including Braf, Pten, and Trp53, and further specific alterations led to metastatic development. Our results provide deeper understanding of melanoma metastasis at the single-cell level, thus aiding further studies in melanoma metastasis control.

An Octopus-Derived Peptide with Antidiuretic Activity in Rats.

Discovering new drug candidates with high efficacy and few side effects is a major challenge in new drug development. The two evolutionarily related peptides oxytocin (OXT) and arginine vasopressin (AVP) are known to be associated with a variety of physiological and psychological processes via the association of OXT with three types of AVP receptors. Over decades, many synthetic analogs of these peptides have been designed and tested for therapeutic applications; however, only a few studies of their natural analogs have been performed. In this study, we investigated the bioactivity and usefulness of two natural OXT/AVP analogs that originate from the marine invertebrate Octopus vulgaris, named octopressin (OTP) and cephalotocin (CPT). By measuring the intracellular Ca2+ or cyclic AMP increase in each OXT/AVP receptor subtype-overexpressing cell, we found that CPT, but not OTP, acts as a selective agonist of human AVP type 1b and 2 receptors. This behavior is reminiscent of desmopressin, the most widely prescribed antidiuretic drug in the world. Similar to the case for desmopressin, a single intravenous tail injection of CPT into Sprague-Dawley rats reduced urine output and increased urinary osmolality. In conclusion, we suggest that CPT has a significant antidiuretic effect and that CPT might be beneficial for treating urological conditions such as nocturia, enuresis, and diabetes insipidus.

Chromosome-level genome assembly of the shuttles hoppfish, Periophthalmus modestus.

BACKGROUND: The shuttles hoppfish (mudskipper), Periophthalmus modestus, is one of the mudskippers, which are the largest group of amphibious teleost fishes, which are uniquely adapted to live on mudflats. Because mudskippers can survive on land for extended periods by breathing through their skin and through the lining of the mouth and throat, they were evaluated as a model for the evolutionary sea-land transition of Devonian protoamphibians, ancestors of all present tetrapods. RESULTS: A total of 39.6, 80.2, 52.9, and 33.3 Gb of Illumina, Pacific Biosciences, 10X linked, and Hi-C data, respectively, was assembled into 1,419 scaffolds with an N50 length of 33 Mb and BUSCO score of 96.6%. The assembly covered 117% of the estimated genome size (729 Mb) and included 23 pseudo-chromosomes anchored by a Hi-C contact map, which corresponded to the top 23 longest scaffolds above 20 Mb and close to the estimated one. Of the genome, 43.8% were various repetitive elements such as DNAs, tandem repeats, long interspersed nuclear elements, and simple repeats. Ab initio and homology-based gene prediction identified 30,505 genes, of which 94% had homology to the 14 Actinopterygii transcriptomes and 89% and 85% to Pfam familes and InterPro domains, respectively. Comparative genomics with 15 Actinopterygii species identified 59,448 gene families of which 12% were only in P. modestus. CONCLUSIONS: We present the high quality of the first genome assembly and gene annotation of the shuttles hoppfish. It will provide a valuable resource for further studies on sea-land transition, bimodal respiration, nitrogen excretion, osmoregulation, thermoregulation, vision, and mechanoreception.