Sour Sensing from the Tongue to the Brain.

The ability to sense sour provides an important sensory signal to prevent the ingestion of unripe, spoiled, or fermented foods. Taste and somatosensory receptors in the oral cavity trigger aversive behaviors in response to acid stimuli. Here, we show that the ion channel Otopetrin-1, a proton-selective channel normally involved in the sensation of gravity in the vestibular system, is essential for sour sensing in the taste system. We demonstrate that knockout of Otop1 eliminates acid responses from sour-sensing taste receptor cells (TRCs). In addition, we show that mice engineered to express otopetrin-1 in sweet TRCs have sweet cells that also respond to sour stimuli. Next, we genetically identified the taste ganglion neurons mediating each of the five basic taste qualities and demonstrate that sour taste uses its own dedicated labeled line from TRCs in the tongue to finely tuned taste neurons in the brain to trigger aversive behaviors.

Vulnerability to APOBEC3G linked to the pathogenicity of deltaretroviruses.

Human retroviruses are derived from simian ones through cross-species transmission. These retroviruses are associated with little pathogenicity in their natural hosts, but in humans, HIV causes AIDS, and human T-cell leukemia virus type 1 (HTLV-1) induces adult T-cell leukemia-lymphoma (ATL). We analyzed the proviral sequences of HTLV-1, HTLV-2, and simian T-cell leukemia virus type 1 (STLV-1) from Japanese macaques (Macaca fuscata) and found that APOBEC3G (A3G) frequently generates G-to-A mutations in the HTLV-1 provirus, whereas such mutations are rare in the HTLV-2 and STLV-1 proviruses. Therefore, we investigated the mechanism of how HTLV-2 is resistant to human A3G (hA3G). HTLV-1, HTLV-2, and STLV-1 encode the so-called antisense proteins, HTLV-1 bZIP factor (HBZ), Antisense protein of HTLV-2 (APH-2), and STLV-1 bZIP factor (SBZ), respectively. APH-2 efficiently inhibits the deaminase activity of both hA3G and simian A3G (sA3G). HBZ and SBZ strongly suppress sA3G activity but only weakly inhibit hA3G, suggesting that HTLV-1 is incompletely adapted to humans. Unexpectedly, hA3G augments the activation of the transforming growth factor (TGF)-ß/Smad pathway by HBZ, and this activation is associated with ATL cell proliferation by up-regulating BATF3/IRF4 and MYC. In contrast, the combination of APH-2 and hA3G, or the combination of SBZ and sA3G, does not enhance the TGF-ß/Smad pathway. Thus, HTLV-1 is vulnerable to hA3G but utilizes it to promote the proliferation of infected cells via the activation of the TGF-ß/Smad pathway. Antisense factors in each virus, differently adapted to control host cellular functions through A3G, seem to dictate the pathogenesis.

EPAS1, a hypoxia- and ferroptosis-related gene, promotes malignant behaviour of cervical cancer by ceRNA and super-enhancer.

Hypoxia and Ferroptosis are associated with the malignant behaviour of cervical cancer. Endothelial PAS domain-containing protein 1 (EPAS1) contributes to the progression of cervical cancer. EPAS1 plays important roles in hypoxia and ferroptosis. Using the GEO dataset, machine-learning algorithms were used to screen for hypoxia- and ferroptosis-related genes (HFRGs) in cervical cancer. EPAS1 was identified as the hub gene. qPCR and WB were used to investigate the expression of EPAS1 in normal and cervical cancer tissues. The proliferation, invasion and migration of EPAS1 cells in HeLa and SiHa cell lines were detected using CCK8, transwell and wound healing assays, respectively. Apoptosis was detected by flow cytometry. A dual-luciferase assay was used to analyse the MALAT1-miR-182-5P-EPAS1 mRNA axis and core promoter elements of the super-enhancer. EPAS1 was significantly overexpressed in cervical cancer tissues. EPAS1 could increase the proliferation, invasion, migration of HeLa and SiHa cells and reduce the apoptosis of HeLa and SiHa cell. According to the double-luciferase assay, EPAS1 expression was regulated by the MALAT1-Mir-182-5p-EPAS1 mRNA axis. EPAS1 is associated with super-enhancers. Double-luciferase assay showed that the core elements of the super-enhancer were E1 and E3. EPAS1, an HFRG, is significantly overexpressed in cervical cancer. EPAS1 promotes malignant behaviour of cervical cancer cells. EPAS1 expression is regulated by super-enhancers and the MALAT1-miR-182-5P- EPAS1 mRNA axis. EPAS1 may be a target for the diagnosis and treatment of cervical cancer.

Identification of the effects of alkalinity exposure on the gills of oriental river prawns, Macrobrachium nipponense.

Macrobrachium nipponense is an important commercial freshwater species in China. However, the ability of alkali tolerance of M. nipponense is insufficient to culture in the major saline-alkali water source in China. Thus, it is urgently needed to perform the genetic improvement of alkali tolerance in this species. In the present study, we aimed to analyse the effects of alkali treatment on gills in this species after 96 h alkalinity exposure under the alkali concentrations of 0 mmol/L, 4 mmol/L, 8 mmol/L, and 12 mmol/L through performing the histological observations, measurement of antioxidant enzymes, metabolic profiling analysis, and transcriptome profiling analysis. The results of the present study revealed that alkali treatment stimulated the contents of malondialdehyde, glutathione, glutathione peroxidase in gills, indicating these antioxidant enzymes plays essential roles in the protection of body from the damage, caused by the alkali treatment. In addition, high concentration of alkali treatment (> 8 mmol/L) resulted in the damage of gill membrane and haemolymph vessel, affecting the normal respiratory function of gill. Metabolic profiling analysis revealed that Metabolic pathways, Biosynthesis of secondary metabolites, Biosynthesis of plant secondary metabolites, Microbial metabolism in diverse environments, Biosynthesis of amino acids were identified as the main enriched metabolic pathways of differentially expressed metabolites, which are consistent with the previous publications, treated by the various environmental factors. Transcriptome profiling analyses revealed that the alkali concentration of 12 mmol/L has more regulatory effects on the changes of gene expression than the other alkali concentrations. KEGG analysis revealed that Phagosome, Lysosome, Glycolysis/Gluconeogenesis, Purine Metabolism, Amino sugar and nucleotide sugar metabolism, and Endocytosis were identified as the main enriched metabolic pathways in the present study, predicting these metabolic pathways may be involved in the adaption of alkali treatment in M. nipponense. Phagosome, Lysosome, Purine Metabolism, and Endocytosis are immune-related metabolic pathways, while Glycolysis/Gluconeogenesis, and Amino sugar and nucleotide sugar metabolism are energy metabolism-related metabolic pathways. Quantitative PCR analyses of differentially expressed genes (DEGs) verified the accuracy of the RNA-Seq. Alkali treatment significantly stimulated the expressions of DEGs from the metabolic pathways of Phagosome and Lysosome, suggesting Phagosome and Lysosome play essential roles in the regulation of alkali tolerance in this species, as well as the genes from these metabolic pathways. The present study identified the effects of alkali treatment on gills, providing valuable evidences for the genetic improvement of alkali tolerance in M. nipponense.

Identification of genes regulated by 20-Hydroxyecdysone in Macrobrachium nipponense using comparative transcriptomic analysis.

BACKGROUND: Macrobrachium nipponense is a freshwater prawn of economic importance in China. Its reproductive molt is crucial for seedling rearing and directly impacts the industry's economic efficiency. 20-hydroxyecdysone (20E) controls various physiological behaviors in crustaceans, among which is the initiation of molt. Previous studies have shown that 20E plays a vital role in regulating molt and oviposition in M. nipponense. However, research on the molecular mechanisms underlying the reproductive molt and role of 20E in M. nipponense is still limited. RESULTS: A total of 240.24 Gb of data was obtained from 18 tissue samples by transcriptome sequencing, with > 6 Gb of clean reads per sample. The efficiency of comparison with the reference transcriptome ranged from 87.05 to 92.48%. A total of 2532 differentially expressed genes (DEGs) were identified. Eighty-seven DEGs associated with molt or 20E were screened in the transcriptomes of the different tissues sampled in both the experimental and control groups. The reliability of the RNA sequencing data was confirmed using Quantitative Real-Time PCR. The expression levels of the eight strong candidate genes showed significant variation at the different stages of molt. CONCLUSION: This study established the first transcriptome library for the different tissues of M. nipponense in response to 20E and demonstrated the dominant role of 20E in the molting process of this species. The discovery of a large number of 20E-regulated strong candidate DEGs further confirms the extensive regulatory role of 20E and provides a foundation for the deeper understanding of its molecular regulatory mechanisms.

Combined targeting of GPX4 and BCR-ABL tyrosine kinase selectively compromises BCR-ABL+ leukemia stem cells.

BACKGROUND: In the ongoing battle against BCR-ABL+ leukemia, despite significant advances with tyrosine kinase inhibitors (TKIs), the persistent challenges of drug resistance and the enduring presence of leukemic stem cells (LSCs) remain formidable barriers to achieving a cure. METHODS: In this study, we demonstrated that Disulfiram (DSF) induces ferroptosis to synergize with TKIs in inhibiting BCR-ABL+ cells, particularly targeting resistant cells and LSCs, using cell models, mouse models, and primary cells from patients. We elucidated the mechanism by which DSF promotes GPX4 degradation to induce ferroptosis through immunofluorescence, co-immunoprecipitation (CO-IP), RNA sequencing, lipid peroxidation assays, and rescue experiments. RESULTS: Here, we present compelling evidence elucidating the sensitivity of DSF, an USA FDA-approved drug for alcohol dependence, towards BCR-ABL+ cells. Our findings underscore DSF's ability to selectively induce a potent cytotoxic effect on BCR-ABL+ cell lines and effectively inhibit primary BCR-ABL+ leukemia cells. Crucially, the combined treatment of DSF with TKIs selectively eradicates TKI-insensitive stem cells and resistant cells. Of particular note is DSF's capacity to disrupt GPX4 stability, elevate the labile iron pool, and intensify lipid peroxidation, ultimately leading to ferroptotic cell death. Our investigation shows that BCR-ABL expression induces alterations in cellular iron metabolism and increases GPX4 expression. Additionally, we demonstrate the indispensability of GPX4 for LSC development and the initiation/maintenance of BCR-ABL+ leukemia. Mechanical analysis further elucidates DSF's capacity to overcome resistance by reducing GPX4 levels through the disruption of its binding with HSPA8, thereby promoting STUB1-mediated GPX4 ubiquitination and subsequent proteasomal degradation. Furthermore, the combined treatment of DSF with TKIs effectively targets both BCR-ABL+ blast cells and drug-insensitive LSCs, conferring a significant survival advantage in mouse models. CONCLUSION: In summary, the dual inhibition of GPX4 and BCR-ABL presents a promising therapeutic strategy to synergistically target blast cells and drug-insensitive LSCs in patients, offering potential avenues for advancing leukemia treatment.

Anti-HTLV-1 immunity combined with proviral load as predictive biomarkers for adult T-cell leukemia-lymphoma.

Human T-cell leukemia virus type 1 (HTLV-1) establishes chronic infection in humans and induces a T-cell malignancy called adult T-cell leukemia-lymphoma (ATL) and several inflammatory diseases such as HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP). Persistent HTLV-1 infection is established under the pressure of host immunity, and therefore the immune response against HTLV-1 is thought to reflect the status of the disease it causes. Indeed, it is known that cellular immunity against viral antigens is suppressed in ATL patients compared to HAM/TSP patients. In this study, we show that profiling the humoral immunity to several HTLV-1 antigens, such as Gag, Env, and Tax, and measuring proviral load are useful tools for classifying disease status and predicting disease development. Using targeted sequencing, we found that several carriers whom this profiling method predicted to be at high risk for developing ATL indeed harbored driver mutations of ATL. The clonality of HTLV-1-infected cells in those carriers was still polyclonal; it is consistent with an early stage of leukemogenesis. Furthermore, this study revealed significance of anti-Gag proteins to predict high risk group in HTLV-1 carriers. Consistent with this finding, anti-Gag cytotoxic T lymphocytes (CTLs) were increased in patients who received hematopoietic stem cell transplantation and achieved remission state, indicating the significance of anti-Gag CTLs for disease control. Our findings suggest that our strategy that combines anti-HTLV-1 antibodies and proviral load may be useful for prediction of the development of HTLV-1-associated diseases.

iProbiotics: a machine learning platform for rapid identification of probiotic properties from whole-genome primary sequences.

Lactic acid bacteria consortia are commonly present in food, and some of these bacteria possess probiotic properties. However, discovery and experimental validation of probiotics require extensive time and effort. Therefore, it is of great interest to develop effective screening methods for identifying probiotics. Advances in sequencing technology have generated massive genomic data, enabling us to create a machine learning-based platform for such purpose in this work. This study first selected a comprehensive probiotics genome dataset from the probiotic database (PROBIO) and literature surveys. Then, k-mer (from 2 to 8) compositional analysis was performed, revealing diverse oligonucleotide composition in strain genomes and apparently more probiotic (P-) features in probiotic genomes than non-probiotic genomes. To reduce noise and improve computational efficiency, 87 376 k-mers were refined by an incremental feature selection (IFS) method, and the model achieved the maximum accuracy level at 184 core features, with a high prediction accuracy (97.77%) and area under the curve (98.00%). Functional genomic analysis using annotations from gene ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG) and Rapid Annotation using Subsystem Technology (RAST) databases, as well as analysis of genes associated with host gastrointestinal survival/settlement, carbohydrate utilization, drug resistance and virulence factors, revealed that the distribution of P-features was biased toward genes/pathways related to probiotic function. Our results suggest that the role of probiotics is not determined by a single gene, but by a combination of k-mer genomic components, providing new insights into the identification and underlying mechanisms of probiotics. This work created a novel and free online bioinformatic tool, iProbiotics, which would facilitate rapid screening for probiotics.

Visible-light-induced decarboxylative cascade cyclization of acryloylbenzamides with N-hydroxyphthalimide esters via EDA complexes.

A visible-light-induced decarboxylative cascade reaction of acryloylbenzamides with alkyl N-hydroxyphthalimide (NHP) esters for the synthesis of various 4-alkyl isoquinolinediones mediated by triphenylphosphine (PPh3) and sodium iodide (NaI) was developed. This operationally simple protocol proceeded via the photoactivation of electron donor-acceptor (EDA) complexes between N-hydroxyphthalimide esters and NaI/PPh3, resulting in multiple carbon-carbon bond formations without the use of precious metal complexes or synthetically elaborate organic dyes, which provided an alternative practical approach to synthesize diverse isoquinoline-1,3(2H,4H)-dione derivatives.

Global prevalence and burden of multidrug-resistant tuberculosis from 1990 to 2019.

BACKGROUND: Tuberculosis(TB) remains a pressing public health challenge, with multidrug-resistant tuberculosis (MDR-TB) emerging as a major threat. And healthcare authorities require reliable epidemiological evidence as a crucial reference to address this issue effectively. The aim was to offer a comprehensive epidemiological assessment of the global prevalence and burden of MDR-TB from 1990 to 2019. METHODS: Estimates and 95% uncertainty intervals (UIs) for the age-standardized prevalence rate (ASPR), age-standardized incidence rate (ASIR), age-standardized disability-adjusted life years rate (ASR of DALYs), and age-standardized death rate (ASDR) of MDR-TB were obtained from the Global Burden of Disease (GBD) 2019 database. The prevalence and burden of MDR-TB in 2019 were illustrated in the population and regional distribution. Temporal trends were analyzed by using Joinpoint regression analysis to calculate the annual percentage change (APC), average annual percentage change (AAPC) and its 95% confidence interval(CI). RESULTS: The estimates of the number of cases were 687,839(95% UIs: 365,512 to 1223,262), the ASPR were 8.26 per 100,000 (95%UIs: 4.61 to 15.20), the ASR of DALYs were 52.38 per 100,000 (95%UIs: 22.64 to 97.60) and the ASDR were 1.36 per 100,000 (95%UIs: 0.54 to 2.59) of MDR-TB at global in 2019. Substantial burden was observed in Africa and Southeast Asia. Males exhibited higher ASPR, ASR of DALYs, and ASDR than females across most age groups, with the burden of MDR-TB increasing with age. Additionally, significant increases were observed globally in the ASIR (AAPC = 5.8; 95%CI: 5.4 to 6.1; P < 0.001), ASPR (AAPC = 5.9; 95%CI: 5.4 to 6.4; P < 0.001), ASR of DALYs (AAPC = 4.6; 95%CI: 4.2 to 5.0; P < 0.001) and ASDR (AAPC = 4.4; 95%CI: 4.0 to 4.8; P < 0.001) of MDR-TB from 1990 to 2019. CONCLUSIONS: This study underscored the persistent threat of drug-resistant tuberculosis to public health. It is imperative that countries and organizations worldwide take immediate and concerted action to implement measures aimed at significantly reducing the burden of TB.

Oral enzyme-responsive nanoprobes for targeted theranostics of inflammatory bowel disease.

BACKGROUND: Inflammatory bowel disease (IBD) is a progressive and debilitating inflammatory disease of the gastrointestinal tract (GIT). Despite recent advances, precise treatment and noninvasive monitoring remain challenging. METHODS: Herein, we developed orally-administered, colitis-targeting and hyaluronic acid (HA)-modified, core-shell curcumin (Cur)- and cerium oxide (CeO2)-loaded nanoprobes (Cur@PC-HA/CeO2 NPs) for computed tomography (CT) imaging-guided treatment and monitoring of IBD in living mice. RESULTS: Following oral administration, high-molecular-weight HA maintains integrity with little absorption in the upper GIT, and then actively accumulates at local colitis sites owing to its colitis-targeting ability, leading to specific CT enhancement lasting for 24 h. The retained NPs are further degraded by hyaluronidase in the colon to release Cur and CeO2, thereby exerting anti-inflammatory and antioxidant effects. Combined with the ability of NPs to regulate intestinal flora, the oral NPs result in substantial relief in symptoms. Following multiple treatments, the gradually decreasing range of the colon with high CT attenuation correlates with the change in the clinical biomarkers, indicating the feasibility of treatment response and remission. CONCLUSION: This study provides a proof-of-concept for the design of a novel theranostic integration strategy for concomitant IBD treatment and the real-time monitoring of treatment responses.

RETINAL MICROVASCULAR CHANGES AND RISK OF CORONARY HEART DISEASE: A Systematic Review and Meta-Analysis.

PURPOSE: To quantify associations between various retinal microvascular changes and the risk of the development of coronary heart disease (CHD). METHODS: PubMed, Embase, Web of Science, and Cochrane Library databases were searched for cohort studies on the association between retinal microvascular changes and incident CHD up to July 31, 2023. The summary risk estimates were estimated using the random-effects model. Subgroup and sensitivity analyses were performed to investigate the potential source of heterogeneity. RESULTS: The authors identified 21 studies that met the inclusion criteria of this meta-analysis through database searching. This study yielded significant associations between retinal microvascular changes, including arteriolar narrowing, venular widening, vessel occlusion, and other retinal vascular signs, and the risk of CHD, with pooled adjusted hazard ratios of 1.20 (95% confidence interval: 1.13-1.27). In sex- and age-stratified analyses, retinal microvascular changes were associated with a greater risk of developing CHD in female patients and younger adults. CONCLUSION: A range of retinal microvascular changes was associated with the risk of CHD, particularly in female patients and younger ages. The results of this study support the concept that retinal microvascular abnormalities may be markers for future CHD. Noninvasive retinal microvascular assessments may be helpful in screening patients with increased CHD risk.

Lag effect of ambient temperature on respiratory emergency department visits in Beijing: a time series and pooled analysis.

BACKGROUND: Although the association between ambient temperature and mortality of respiratory diseases was numerously documented, the association between various ambient temperature levels and respiratory emergency department (ED) visits has not been well studied. A recent investigation of the association between respiratory ED visits and various levels of ambient temperature was conducted in Beijing, China. METHODS: Daily meteorological data, air pollution data, and respiratory ED visits data from 2017 to 2018 were collected in Beijing. The relationship between ambient temperature and respiratory ED visits was explored using a distributed lagged nonlinear model (DLNM). Then we performed subgroup analysis based on age and gender. Finally, meta-analysis was utilized to aggregate the total influence of ambient temperature on respiratory ED visits across China. RESULTS: The single-day lag risk for extreme cold peaked at a relative risk (RR) of 1.048 [95% confidence interval (CI): 1.009, 1.088] at a lag of 21 days, with a long lag effect. As for the single-day lag risk for extreme hot, a short lag effect was shown at a lag of 7 days with an RR of 1.076 (95% CI: 1.038, 1.114). The cumulative lagged effects of both hot and cold effects peaked at lag 0-21 days, with a cumulative risk of the onset of 3.690 (95% CI: 2.133, 6.382) and 1.641 (95% CI: 1.284, 2.098), respectively, with stronger impact on the hot. Additionally, the elderly were more sensitive to ambient temperature. The males were more susceptible to hot weather than the females. A longer cold temperature lag effect was found in females. Compared with the meta-analysis, a pooled effect of ambient temperature was consistent in general. In the subgroup analysis, a significant difference was found by gender. CONCLUSIONS: Temperature level, age-specific, and gender-specific effects between ambient temperature and the number of ED visits provide information on early warning measures for the prevention and control of respiratory diseases.

Medium-term exposure to size-fractioned particulate matter and asthma exacerbations in China: A longitudinal study of asthmatics with poor medication adherence.

BACKGROUND: Studies have shown that short- and long-term exposure to particulate matter (PM) can increase the risk of asthma morbidity and mortality. However, the effect of medium-term exposure remains unknown. We aim to examine the effect of medium-term exposure to size-fractioned PM on asthma exacerbations among asthmatics with poor medication adherence. METHODS: We conducted a longitudinal study in China based on the National Mobile Asthma Management System Project that specifically and routinely followed asthma exacerbations in asthmatics with poor medication adherence from April 2017 to May 2019. High-resolution satellite remote-sensing data were used to estimate each participant's medium-term exposure (on average 90 days) to size-fractioned PM (PM1, PM2.5, and PM10) based on the residential address and the date of the follow-up when asthma exacerbations (e.g., hospitalizations and emergency room visits) occurred or the end of the follow-up. The Cox proportional hazards model was employed to examine the hazard ratio of asthma exacerbations associated with each PM after controlling for sex, age, BMI, education level, geographic region, and temperature. RESULTS: Modelling results revealed nonlinear exposure-response associations of asthma exacerbations with medium-term exposure to PM1, PM2.5, and PM10. Specifically, for emergency room visits, we found an increased hazard ratio for PM1 above 22.8 µg/m3 (1.060, 95 % CI: 1.025-1.096, per 1 µg/m3 increase), PM2.5 above 38.2 µg/m3 (1.032, 95 % CI: 1.010-1.054), and PM10 above 78.6 µg/m3 (1.019, 95 % CI: 1.006-1.032). For hospitalizations, we also found an increased hazard ratio for PM1 above 20.3 µg/m3 (1.055, 95 % CI: 1.001-1.111) and PM2.5 above 39.2 µg/m3 (1.038, 95 % CI: 1.003-1.074). Furthermore, the effects of PM were greater for a longer exposure window (90-180 days) and among participants with a high BMI. CONCLUSION: This study suggests that medium-term exposure to PM is associated with an increased risk of asthma exacerbations in asthmatics with poor medication adherence, with a higher risk from smaller PM.

Long Non-Coding RNA H19 Leads to Upregulation of γ-Globin Gene Expression during Erythroid Differentiation.

Long noncoding RNAs (lncRNAs) are important because they are involved in a variety of life activities and have many downstream targets. Moreover, there is also increasing evidence that some lncRNAs play important roles in the expression and regulation of γ-globin genes. In our previous study, we analyzed genetic material from nucleated red blood cells (NRBCs) extracted from premature and full-term umbilical cord blood samples. Through RNA sequencing (RNA-Seq) analysis, lncRNA H19 emerged as a differentially expressed transcript between the two blood types. While this discovery provided insight into H19, previous studies had not investigated its effect on the γ-globin gene. Therefore, the focus of our study was to explore the impact of H19 on the γ-globin gene. In this study, we discovered that overexpressing H19 led to a decrease in HBG mRNA levels during erythroid differentiation in K562 cells. Conversely, in CD34+ hematopoietic stem cells and human umbilical cord blood-derived erythroid progenitor (HUDEP-2) cells, HBG expression increased. Additionally, we observed that H19 was primarily located in the nucleus of K562 cells, while in HUDEP-2 cells, H19 was present predominantly in the cytoplasm. These findings suggest a significant upregulation of HBG due to H19 overexpression. Notably, cytoplasmic localization in HUDEP-2 cells hints at its potential role as a competing endogenous RNA (ceRNA), regulating γ-globin expression by targeting microRNA/mRNA interactions.

Identification of Candidate Male-Reproduction-Related Genes from the Testis and Androgenic Gland of Macrobrachium nipponense, Regulated by PDHE1, through Transcriptome Profiling Analysis.

The previous publication identified that pyruvate dehydrogenase E1 (PDHE1) positively regulated the process of male reproduction in M. nipponense through affecting the expressions of insulin-like androgenic gland hormone. The present study aimed to identify the potential male-reproduction-related genes that were regulated by PDHE1 through performing the transcriptome profiling analysis in the testis and androgenic gland after the knockdown of the expressions of PDHE1 by the injection of dsPDHE1. Both RNA-Seq and qPCR analysis identified the significant decreases in PDHE1 expressions in the testis and androgenic gland in dsPDHE1-injected prawns compared to those in dsGFP-injected prawns, indicating the efficiency of dsPDHE1 in the present study. Transcriptome profiling analysis identified 56 and 127 differentially expressed genes (DEGs) in the testis and androgenic gland, respectively. KEGG analysis revealed that the energy-metabolism-related pathways represented the main enriched metabolic pathways of DEGs in both the testis and androgenic gland, including pyruvate metabolism, the Citrate cycle (TCA cycle), Glycolysis/Gluconeogenesis, and the Glucagon signaling pathway. Thus, it is predicted that these metabolic pathways and the DEGs from these metabolic pathways regulated by PDHE1 may be involved in the regulation of male reproduction in M. nipponense. Furthermore, four genes were found to be differentially expressed in both the testis and androgenic gland, of which ribosomal protein S3 was down-regulated and uncharacterized protein LOC113829596 was up-regulated in both the testis and androgenic gland in dsPDHE1-injected prawns. The present study provided valuable evidence for the establishment of an artificial technique to regulate the process of male reproduction in M. nipponense.

NPC Intracellular Cholesterol Transporter 1 Regulates Ovarian Maturation and Molting in Female Macrobrachium nipponense.

NPC intracellular cholesterol transporter 1 (NPC1) plays an important role in sterol metabolism and transport processes and has been studied in many vertebrates and some insects, but rarely in crustaceans. In this study, we characterized NPC1 from Macrobrachium nipponense (Mn-NPC1) and evaluated its functions. Its total cDNA length was 4283 bp, encoding for 1344 amino acids. It contained three conserved domains typical of the NPC family (NPC1_N, SSD, and PTC). In contrast to its role in insects, Mn-NPC1 was mainly expressed in the adult female hepatopancreas, with moderate expression in the ovary and heart. No expression was found in the embryo (stages CS-ZS) and only weak expression in the larval stages from hatching to the post-larval stage (L1-PL15). Mn-NPC1 expression was positively correlated with ovarian maturation. In situ hybridization showed that it was mainly located in the cytoplasmic membrane and nucleus of oocytes. A 25-day RNA interference experiment was employed to illustrate the Mn-NPC1 function in ovary maturation. Experimental knockdown of Mn-NPC1 using dsRNA resulted in a marked reduction in the gonadosomatic index and ecdysone content of M. nipponense females. The experimental group showed a significant delay in ovarian maturation and a reduction in the frequency of molting. These results expand our understanding of NPC1 in crustaceans and of the regulatory mechanism of ovarian maturation in M. nipponense.

Functional Study of the Role of the Methyl Farnesoate Epoxidase Gene in the Ovarian Development of Macrobrachium nipponense.

Methyl farnesoate epoxidase (MFE) is a gene encoding an enzyme related to the last step of juvenile hormone biosynthesis. Mn-MFE cDNA has a total length of 1695 bp and an open reading frame (ORF) length of 1482 bp, encoding 493 amino acids. Sequence analysis showed that its amino acid sequence has a PPGP hinge, an FGCG structural domain, and other structural domains specific to the P450 family of enzymes. Mn-MFE was most highly expressed in the hepatopancreas, followed by the ovary and gill, weakly expressed in heart and muscle tissue, and barely expressed in the eyestalk and cranial ganglion. Mn-MFE expression remained stable during the larval period, during which it mainly played a critical role in gonadal differentiation. Expression in the ovary was positively correlated and expression in the hepatopancreas was negatively correlated with ovarian development. In situ hybridization (ISH) showed that the signal was expressed in the oocyte, nucleus, cell membrane and follicular cells, and the intensity of expression was strongest at stage O-IV. The knockdown of Mn-MFE resulted in a significantly lower gonadosomatic index and percentage of ovaries past stage O-III compared to the control group. However, no differences were found in the cumulative frequency of molting between the experimental and control groups. Moreover, the analysis of ovarian tissue sections at the end of the experiment showed differences between groups in development speed but not in subcellular structure. These results demonstrate that Mn-MFE promotes the ovarian development of Macrobrachium nipponense adults but has no effect on molting.

Studies on the Relationships between Growth and Gonad Development during First Sexual Maturation of Macrobrachium nipponense and Associated SNPs Screening.

In this study, we used full-sib families to investigate the association between growth and gonad development during first sexual maturation of M. nipponense. We found that male GSI was significantly negatively correlated with growth traits (p < 0.01) and there were no significant correlations between female GSI (Gonadosomatic index) and growth traits (p > 0.05). HSI (Hepatopancreas index) in both males and females showed no significant correlations with growth traits (p > 0.05). We furthermore investigated the association between the specific allele of Mn-CTS L1 polymorphism and gonad development and growth traits. In total, 35 mutation loci were screened and 16 high-quality single-nucleotide polymorphisms (SNPs) loci were obtained after validation. Four and two SNPs proved to be strongly associated with all growth traits in female and male M. nipponense separately, among which A+118T might be a candidate SNP positively associated with large growth traits. Two and one SNPs were screened, respectively, in males and females to associate with GSI, while three SNPs were detected to associate with female HSI, among which A+1379C may be applied as a potential molecular marker for gene-assisted selection to improve both reproduction speed and growth traits in M. nipponense.

Role of Mn-LIPA in Sex Hormone Regulation and Gonadal Development in the Oriental River Prawn, Macrobrachium nipponense.

This study investigates the role of lysosomal acid lipase (LIPA) in sex hormone regulation and gonadal development in Macrobrachium nipponense. The full-length Mn-LIPA cDNA was cloned, and its expression patterns were analyzed using quantitative real-time PCR (qPCR) in various tissues and developmental stages. Higher expression levels were observed in the hepatopancreas, cerebral ganglion, and testes, indicating the potential involvement of Mn-LIPA in sex differentiation and gonadal development. In situ hybridization experiments revealed strong Mn-LIPA signaling in the spermatheca and hepatopancreas, suggesting their potential role in steroid synthesis (such as cholesterol, fatty acids, cholesteryl ester, and triglycerides) and sperm maturation. Increased expression levels of male-specific genes, such as insulin-like androgenic gland hormone (IAG), sperm gelatinase (SG), and mab-3-related transcription factor (Dmrt11E), were observed after dsMn-LIPA (double-stranded LIPA) injection, and significant inhibition of sperm development and maturation was observed histologically. Additionally, the relationship between Mn-LIPA and sex-related genes (IAG, SG, and Dmrt11E) and hormones (17ß-estradiol and 17α-methyltestosterone) was explored by administering sex hormones to male prawns, indicating that Mn-LIPA does not directly control the production of sex hormones but rather utilizes the property of hydrolyzing triglycerides and cholesterol to provide energy while influencing the synthesis and secretion of self-sex hormones. These findings provide valuable insights into the function of Mn-LIPA in M. nipponense and its potential implications for understanding sex differentiation and gonadal development in crustaceans. It provides an important theoretical basis for the realization of a monosex culture of M. nipponense.