Sexual growth dimorphism is a unique trait of the Chinese tongue sole (Cynoglossus semilaevis) where female fish grow much faster and bigger than their male counterparts. In tongue sole, the transcription factor dmrt1 is essential for testis formation and germ cell maintenance, but its disruption not only causes sterility but also leads to enhanced somatic growth in males. To study how dmrt1 links reproduction and growth, homozygous dmrt1-knockout males were treated with LNP-dmrt1, which carries dmrt1 mRNA inside. After several weeks of repeated injections, histological examination revealed some recovery of spermatogenesis, while some seminiferous tubules contained developing sperms. The transcriptome across the brain, blood, and gonad showed that the dmrt1 knock out changed steroidogenesis, meiosis, and hormone signaling, but LNP-dmrt1 treatment turned on many pathways involved in extracellular matrix interaction and steroid biosynthesis. Using circRNA, miRNA, and mRNA expression data together we built a ceRNA network, showing a couple of main controllers linked to cell cycle and neural signaling. Our results show that although LNP-mediated dmrt1 mRNA delivery was able to partially rebuild testicular structure and reactivate some key pathways related to steroid synthesis and tissue organization, full recovery of normal spermatogenesis was not achieved. The involvement of circRNA–miRNA–mRNA regulatory networks further suggesting that post-transcriptional regulation may be involved in the limited rescue effect, rather than dmrt1 acting alone. These findings deepen our understanding of how reproductive capacity and growth are coordinated in sexually dimorphic fish and provide a potential foundation for developing more effective strategies to improve fertility and breeding efficiency in tongue sole aquaculture.

Xanthomonas campestris is a diverse bacterial plant pathogen that causes disease on many critical crops. However, the mechanisms underlying its virulence have remained elusive. We paired a quantitative virulence screen with a bacterial genome-wide association study (GWAS) to uncover the genetic underpinnings of virulence on this model pathogen. We found that a diverse panel of 67 X. campestris strains varied substantially in their ability to grow and cause disease on the model host A. thaliana Col-0. However, its virulence on this host was not a monophyletic trait. This opened the door to the application of a GWAS to test for convergent genetic loci underlying virulence in X. campestris. After correcting for phylogenetic structure, only four accessory gene families were positively associated with X. campestris virulence: a xopJ family effector, a hypothetical gene, and two genes associated with transposable element mobility. These four genes mapped to a Tn3-family transposon that appears to have been horizontally transferred across multiple distinct X. campestris lineages, enabling the emergence of divergent pathogens in these lineages with the same underlying genetic architecture. Further analysis revealed that these transposons were often nested within larger transposons that also contained additional virulence genes, including effectors xopE, avrBs3, and xopAH, which may further contribute to host specific adaptation within independent lineages. This study demonstrates that the horizontal transfer of an effector harbouring transposon is a primary determinant of X. campestris virulence on the model host A. thaliana. It also highlights the utility of GWASs for identifying bacterial loci associated with virulence, which has far-reaching implications for pathogen surveillance and the engineering of resistant crops.

Sebastes schlegelii (Hilgendorf, 1880), a viviparous fish with annual ovarian remodeling, shows marked activation of Extracellular Matrix (ECM)-receptor interaction signaling post-mating. However, the evolutionary conservation and functional significance of a key ECM remodeler-Matrix metalloproteinase 2 (MMP2) in this unique reproductive system are poorly understood. In this study, we identified the core members of the MMP family from the S. schlegelii genome and investigated their expression patterns and localization in ovarian tissues. The results demonstrated that the gelatinase gene mmp2 (Ssc_10020200) displayed high expression levels in the mated ovaries, with its expression being predominantly located in the ovarian follicular cell layers and follicular placenta and embryos. Both in vitro and in vivo experiments revealed that MMP2 knockdown during the vitellogenic and gestation stages led to significant ovarian developmental abnormalities. Furthermore, MMP2 knockdown induced a reduction in ECM components such as COL4Α1, LAMB1, and FN1B, accompanied by enhanced apoptotic signaling. This study suggests that MMP2 plays a vital role in modulating ECM remodeling and maintaining ovarian tissue homeostasis, further ensuring normal ovarian development and healthy reproductive processes. Our findings elucidate a critical function for MMP2 in ovarian development of viviparous teleost, and may inspire further strategies to enhance fish reproductive health and aquaculture outcomes.

Exogenous estrogen (17β-estradiol, E2) induces partial sex reversal in the Chinese giant salamander (CGS) at low concentrations (25 µg/L), and complete sex reversal at high concentrations (1000 µg/L), suggesting potential risks of skewed sex ratios in wild populations of this critically endangered species. Simultaneously, controlled sex reversal offers a strategy to enhance the aquaculture efficiency of captive CGS. However, the molecular mechanisms underlying this process remain largely unexplored. Here, we generated histological evidence of male-to-female sex reversal in CGS under E2 treatment and characterized dynamic, multi-timepoint transcriptomic changes during this process. Integrating histological and transcriptomic data, we identified 210 days post-hatching (120 days after hormone treatment) as a critical window for male gonadal sex reversal. During this period, E2 upregulates meiosis-related genes, promoting and maintaining the transition from male to female gonads. Concurrently, E2 suppresses DMRT1 expression, inhibiting male-specific differentiation pathways and facilitating testicular feminization. Notably, early E2 exposure increases CYP19A expression, but as E2 accumulates, it gradually supplants role of CYP19A in hormone-mediated regulation, resulting in a progressive decline of CYP19A function in gonads. These findings provide a comprehensive molecular and histological framework for understanding E2-induced sex reversal in CGS, offering crucial insights for the conservation of wild populations and for improving aquaculture practices.

Alfalfa (Medicago sativa L.) is a globally important forage legume and the most widely cultivated sown pasture species in China. Drought, as one of the most common abiotic stresses, limits alfalfa growth and development. Nitric oxide (NO), a key signaling molecule, plays an essential role in plant growth, development, and responses to various abiotic stresses. In this study, exogenous NO was applied to alfalfa seedlings under drought stress, followed by physiological and transcriptomic analyses. The results showed that sodium nitroprusside (SNP)-treated alfalfa seedlings grew better than untreated controls (CK), with improved leaf tissue structure. Meanwhile, SNP treatment increased proline content, reduced malondialdehyde accumulation, and enhanced hydroxyl radical scavenging capacity. Under drought stress, lignin content increased in alfalfa seedlings. Following exogenous NO application, lignin content in alfalfa seedlings further increased. RNA-Seq analysis identified 20,183 differentially expressed genes (DEGs) in alfalfa seedlings treated with PEG, SNP, or PEG + SNP. KEGG enrichment analysis indicated that the DEGs under drought stress were involved in the phenylpropanoid biosynthesis pathway, which regulates lignin biosynthesis and enhances drought tolerance. GO enrichment analysis revealed that these DEGs participated in the response to water deprivation, thereby modulating drought stress tolerance and improving drought resistance. Furthermore, we assessed the transcript-level changes in genes induced by phenylpropanoid biosynthesis in alfalfa. Among them, 124 DEGs were identified as participating in phenylpropanoid biosynthesis, including 10 up-regulated DEGs (three of which encode key enzymes associated with lignin synthesis), while the remaining DEGs were down-regulated. These findings provide new insights into the transcriptomic mechanisms of SNP-mediated drought adaptation in alfalfa and reveal key pathways contributing to drought tolerance in this species.

Microhaplotypes are playing an increasingly important role in forensic evidence analysis. However, the impact of meiotic recombination events on the efficacy of microhaplotypes in paternity testing remains poorly understood. Based on the data from the 1000 Genomes Project and a high-resolution recombination map, we identified microhaplotypes from low recombination-rate regions (recombination rate < 0.1 cM/Mb) and validated their efficacy for paternity testing by assessing their performance in 580 trio families and in simulated datasets. The results showed that: (1) The low-recombination microhaplotypes outperformed their high-recombination counterparts in paternity testing; (2) A panel of 30 microhaplotypes, selected from low-recombination, non-coding regions of the autosomes, achieved a cumulative power of discrimination (CPD) of 1-6.77 × 10− 37 and cumulative probability of exclusion (CPE) of 1-1.72 × 10− 13; (3) Validation using real-world trios demonstrated that low-recombination microhaplotypes had significantly higher cumulative paternity index (CPI) values (p < 2e-16), outperforming conventional STR markers by seven orders of magnitude; (4) Simulation analyses revealed that low-recombination markers could distinguish true parentage with 100% accuracy and had higher paternity testing accuracy compared to high-recombination counterparts; (5) An online microhaplotype database (http://bioinformatics.gqliulab.com/microhaplotype/) was established, enabling users to query information and calculate forensic parameters. An open-access microhaplotype screening tool, MHpatools, was developed and is freely available at http://bioinformatics.gqliulab.com/MHpatools/. Collectively, the established microhaplotype system, database, and analytical tools would be useful in forensic genetics.

The foundation for ensuring walnuts of high yield and quality is excellent rootstock. Numerous studies have shown that the walnut rootstock has a major impact on the grafted varieties’ quality, yield, stress resistance, growth, and development. Elucidating the mechanism of grafting affinity in walnuts is crucial, as it paves the way for selecting premium rootstocks to achieve increased production and efficiency. In this study, a combined analysis of the transcriptome, miRNA, and metabolome was conducted to elucidate the molecular mechanisms underlying graft compatibility between different walnut rootstocks (Juglans sigillata cv. ‘Qianhe 7’ and ‘Panzhou’). A total of 44, 41, 50, and 28 differential miRNAs were identified between ‘Qianhe 7’ vs ‘Panzhou’ at 1st d, 11th d, 17th d, and 30th d after grafting, respectively. These miRNAs target genes associated with plant hormone signaling pathways, particularly those related to IAA, SA, and ABA, such as ARF, SAUR, and RAS1-like genes. The most dominant metabolites in both J. sigillata rootstocks (cv. ‘Qianhe 7’ and ‘Panzhou’) at 1st d after grafting were salicylic acid (SA) and Indole-3-acetic acid (IAA). The concentration of SA increased with the extension of time in ‘Qianhe 7’, while IAA decreased significantly on the 30th d. The IAA and abscisic acid (ABA) contents of ‘Qianhe 7’ were markedly higher than those observed in ‘Panzhou’ at 11th d and 30th d post-grafting, respectively. But the concentration of jasmonic acid (JA) in the ‘Panzhou’ was significantly higher than that observed in the ‘Qianhe 7’ at 17th d post-grafting. These results indicated that SA, ABA, and IAA may play a pivotal role in the adaptability of grafted J. sigillata (‘Qianhe 7’ and ‘Panzhou’), which provides a foundation for understanding walnut graft affinity. A deeper exploration of the synergistic regulatory network between the identified miRNAs and plant hormones is warranted to clarify the underlying mechanisms, thereby informing hormone-based breeding strategies for walnut rootstocks and facilitating the sustainable advancement of the walnut industry.

Megalobrama amblycephala is an economically important fish species in Chinese aquaculture, but its high sensitivity to hypoxia poses a major challenge for farming. This study aims to investigate the role of the E3 ubiquitin ligase parkin in regulating hypoxia signaling in fish. Under hypoxic stress, parkin expression was significantly upregulated in oxygen-sensitive tissues (brain and gills) and embryos of M. amblycephala, consistent with the expression of hypoxia-responsive genes. Ma-parkin was found to exhibit high conservation in its key functional domains. Mechanistically, it directly interacts with Ma-hif-α, promoting their ubiquitination and proteasomal degradation via E3 ligase activity, thereby suppressing hif-mediated transcriptional activation. Critical residues T74 and C333 were identified as essential for this activity; mutations at these sites impaired Ma-hif-α degradation and ubiquitination, as well as HRE transactivation. Furthermore, we found that Ma-hif-α can regulate the transcription of parkin, forming a negative feedback loop to fine-tune the hypoxic response. Our study demonstrates that parkin serves as a key negative regulator of the hypoxia signaling pathway in M. amblycephala, operating through a finely tuned feedback mechanism. The E3 ubiquitin ligase activity of parkin, dependent on critical residues T74 and C333, directly mediates hif-α ubiquitination and degradation. The reciprocal regulation between hif-α and parkin forms a regulatory circuit that modulates the hypoxic response. These findings not only reveal a conserved adaptive mechanism to hypoxia in fish but also identify specific molecular targets for genetic improvement of hypoxia tolerance in aquaculture species.