Deciphering the environmental adaptation and functional trait of core and noncore bacterial communities in impacted coral reef seawater.

Microorganisms play pivotal roles in different biogeochemical cycles within coral reef waters. Nevertheless, our comprehension of the microbially mediated processes following environmental perturbation is still limited. To gain a deeper insight into the environmental adaptation and nutrient cycling, particularly within core and noncore bacterial communities, it is crucial to understand reef ecosystem functioning. In this study, we delved into the microbial community structure and function of seawater in a coral reef under different degrees of anthropogenic disturbance. To achieve this, we harnessed the power of 16S rRNA gene high-throughput sequencing and metagenomics techniques. The results showed that a continuous temporal succession but little spatial heterogeneity in the bacterial communities of core and noncore taxa and functional profiles involved in nitrogen (N) and phosphorus (P) cycling. Eutrophication state (i.e., nutrient concentration and turbidity) and temperature played pivotal roles in shaping both the microbial community composition and functional traits of coral reef seawater. Within this context, the core subcommunity exhibited a remarkably broader habitat niche breadth, stronger phylogenetic signal and lower environmental sensitivity when compared to the noncore taxa. Null model analysis further revealed that the core subcommunity was governed primarily by stochastic processes, while deterministic processes played a more significant role in shaping the noncore subcommunity. Furthermore, our observations indicated that changes in function related to N cycling were correlated to the variations in noncore taxa, while core taxa played a more substantial role in critical processes such as P cycling. Collectively, these findings facilitated our knowledge about environmental adaptability of core and noncore bacterial taxa and shed light on their respective roles in maintaining diverse nutrient cycling within coral reef ecosystems.

Comparison of biochemical composition, nutritional quality, and metals concentrations between males and females of three different Crassostrea sp.

Gametogenesis can significantly affect the biochemical composition of oysters, but little research on the difference between sexes. Therefore, we conducted the first in-depth study on the composition differences between males and females of three different Crassostrea sp.. The results showed that females had higher glycogen, lipid, Cu and Zn contents than males, while males had higher protein and taurine contents than females at maturity, which might be related to special meiosis pattern of eggs and less energy was required for female gametogenesis. In addition, both males and females had well-balanced essential amino acid compositions. The omega-3: omega-6 (n-3: n-6) ratio of males was significantly higher than that of females, indicating that the nutritional quality of males was higher. These results provide a reliable and refined theoretical and research basis for revealing the nutritional quality, extracting beneficial ingredients, and developing functional food of Crassostrea sp., and provide data support for the sex-regulated breeding of oysters.

Novel GSK-3 kinase inhibitor Pym-5 induces GSK-3ß rather than GSK-3α-dependent melanogenesis in murine melanoma cells.

BACKGROUND: Glycogen synthase kinase-3 (GSK-3) inhibitors are considered to activate Wnt/ß-Catenin, which remains a controversial topic in melanoma treatment. OBJECTIVE: Here, we have developed Pym-5, an attractive GSK-3 inhibitor. Using Pym-5 as a chemical tool to probe the GSK-3 biology, we aimed to investigate the potential of GSK-3 inhibition as a strategy of melanoma treatment and underlying mechanisms. METHODS: Using pigment B16 and B16BL6 murine melanoma model in vitro and a zebrafish pigmentation model in vivo, we investigated Pym-5-meditaed activation of Wnt/ß-Catenin, melanogenesis and antitumor response in melanoma treatment. RESULTS: We found that Pym-5 delayed the growth and promoted melanogenesis of melanoma cells. Pym-5 activated the transcription of ß-Catenin and responsive targets genes (AXIN2 and MITF), melanin biosynthesis genes (TYR, TYRP1 and TYRP2) and eventually elevated the production of melanin. Interestingly, genetic inactivation of GSK-3ß, but not its paralogue GSK-3α, compromised Pym-5-mediated melanogenesis in B16 and B16BL6 cells. CONCLUSION: These data provide insight into the potential therapeutic benefits obtained from activation of Wnt/ß-Catenin signaling pathway and how Pym-5 can regulate melanin production and the rationale for future clinical application of GSK-3 inhibitor in melanoma patients.

Tanshinone IIA attenuates the stemness of breast cancer cells via targeting the miR-125b/STARD13 axis.

BACKGROUND: Tanshinone II A is an effective component extracted from Salvia miltiorrhiza and the roles of Tanshinone IIA in regulating the stemness of tumor cells remain unclear. This work aims to explore the roles and underlying mechanisms of Tanshinone IIA in breast cancer stemness. METHODS: In vitro mammary spheroid formation, flow cytometry assay on CD24-/CD44+ sub-population, ALDH activity detection, cell viability assay and western blot analysis, and in vivo tumor-initiating analysis were performed to examine the effects of Tanshinone IIA on the stemness of breast cancer cells. MiRNAs-based transcriptome sequencing and data analysis, online dataset analysis, luciferase reporter assay combined with rescuing experiments were constructed to explore the underlying mechanisms. RESULTS: Tanshinone IIA attenuated the stemness of breast cancer cells, evident by downregulating the expression of stemness markers, hindering the capacity of spheroid formation, decreasing the CD24-/CD44+ sub-population in a concentration-dependent manner and reducing the tumor-initiating ability of breast cancer cells. Additionally, Tanshinone IIA enhanced adriamycin sensitivity and attenuated adriamycin resistance of breast cancer cells. Combined with miRNAs-based transcriptome sequencing assay, it was found that Tanshinone IIA downregulated miR-125b level and upregulated its target gene STARD13 (StAR-related lipid transfer protein 13) level, thus inactivating the miR-125b/STARD13 axis, which had been previously confirmed to promote breast cancer progression. Notably, miR-125b overexpression enhanced the stemness of breast cancer cells, and miR-125b overexpression or STARD13 knockdown impaired the inhibitory effects of Tanshinone IIA on the stemness of breast cancer cells. CONCLUSIONS: Tanshinone IIA could attenuate the stemness of breast cancer cells via targeting the miR-125b/STARD13 axis.

MDM2 is involved in the regulation of p53 expression in the immune response of oyster Crassostrea hongkongensis.

MDM2 (mouse double-minute) and p53 form a negative feedback loop and play a prominent role in preventing the induction of uncontrolled apoptosis. To better understand their potential roles in oyster Crassostrea hongkongensis, MDM2 and p53 homologs were first isolated and cloned in C. hongkongensis (named ChMDM2 and Chp53), and their mRNA expression patterns in tissues and developmental stages were analyzed. Multiple sequence alignment analysis and phylogenetic analysis of ChMDM2 and Chp53 displayed a high degree of homology and conservation. In addition, exposure to Vibrio coralliilyticus resulted in DNA damage and apoptosis in the hemocytes of C. hongkongensis, and found that the mRNA expression level of ChMDM2 was decreased, while the relative expression of Chp53 was significantly increased in the hemocytes and gills. Furthermore, fluorescence from ChMDM2-EGFP and Chp53-Red were found to be distributed in the nucleus of HEK293T cells. Besides, dual-luciferase reporter assays showed that ChMDM2 antagonized with Chp53 and participates in p53 signaling pathway. In addition, the interaction between ChMDM2 and Chp53 was confirmed strongly by Co-immunoprecipitation assays. Furthermore, the results of RNAi showed that ChMDM2 and Chp53 participated in apoptosis which induced infection of V. coralliilyticus. Taken together, our results characterized the features of ChMDM2 and Chp53, which played a critical role in apoptosis of C. hongkongensis.

Assessment of the juvenile vulnerability of symbiont-bearing giant clams to ocean acidification.

Ocean acidification (OA) severely affects marine bivalves, especially their calcification processes. However, very little is known about the fate of symbiont-bearing giant clams in the acidified oceans, which hinders our ability to develop strategies to protect this ecologically and economically important group in coral reef ecosystems. Here, we explored the integrated juvenile responses of fluted giant clam Tridacna squamosa (Lamarck, 1819) to acidified seawater at different levels of biological organization. Our results revealed that OA did not cause a significant reduction in survival and shell growth performance, indicating that T. squamosa juveniles are tolerated to moderate acidification. Yet, significantly reduced net calcification rate demonstrated the calcifying physiology sensitivity to OA, in line with significant declines in symbiont photosynthetic yield and zooxanthellae density which in turn lowered the amount of energy supply for energetically expensive calcification processes. Subsequent transcriptome sequencing and comparative analysis of differentially expressed genes revealed that the regulation of calcification processes, such as transport of calcification substrates, acid-base regulation, synthesis of organic matrix in the calcifying fluid, as well as metabolic depression were the major response to OA. Taken together, the integration of physiological and molecular responses can provide a comprehensive understanding of how the early life history stages of giant clams respond to OA and make an important leap forward in assessing their fate under future ocean conditions.

Seasonal variations in biochemical composition and nutritional quality of Crassostrea hongkongensis, in relation to the gametogenic cycle.

Variations in the biochemical composition and nutritional quality with annual changes in gonad development were investigated to identify the optimal harvesting time of C. hongkongensis. The glycogen levels in the mantle, muscle, and gonad-visceral mass were significantly lower in June than in December, associated with changes in the expressions of ChGS and ChGP. Protein content consistently exceeded 52% of dry weight. The only significant change in protein levels was an increase between April and June in the gonad-visceral mass, which was associated with the gonadal transition from proliferation to maturation. Moreover, C. hongkongensis consistently had a well-balanced essential amino acid profile, meeting the essential amino acid requirements of preschool children. The lipid content and fatty acid composition of C. hongkongensis varied with the reproductive cycle, but the omega-3:omega-6 ratio was consistently higher than those of C. gigas and C. virginica. In summary, the optimal harvest time of C. hongkongensis was during the inactive stage of most gonads (from August to February at Beihai).

Tetraploid induction of Crassostrea hongkongensis and C. sikamea by inhibiting the polar body 1 release in diploid fertilized eggs.

The production of an all-triploid population by mating tetraploid males with diploid females is the best and most fundamental method for the large-scale production of triploid oysters. Obtaining a stable tetraploid population is essential for guaranteed production in industrialized triploid cultivation. C. hongkongensis and C. sikamea are important oyster breeding species in southern China, and have great economic value. However, there are not any published data on inducing tetraploid C. hongkongensis or C. sikamea. Therefore, we investigated tetraploid induction in these two oyster species by inhibiting the PB1 release in diploid fertilized eggs using Cytochalasin B (CB) under 31 °C, 15 ‰ salinity. The results confirmed that the optimal tetraploid induction conditions for C. hongkongensis were a CB concentration of 0.50 mg/L with induction starting at 9.0 min after fertilization, and stopping at 21.0 min after fertilization; the induction efficiency index reached 0.123 under these conditions. The optimal tetraploid induction conditions for C. sikamea were a CB concentration of 0.50 mg/L, with induction starting at 7.5 min after fertilization and stopping at 18 min after fertilization; the induction efficiency index could be as high as 0.281 under these conditions. However, we confirmed that the tetraploid rate decreased with larval growth, and no tetraploids were detected in the juvenile period of either C. hongkongensis or C. sikamea. This may be attributed to the very low survival of the tetraploid larvae induced by this method, especially as most tetraploid larvae died during the first three days. In summary, it is simple to directly induce tetraploid C. hongkongensis and C. sikamea larvae by inhibiting the PB1 release of diploid zygotes, but the low survival rate makes it challenging to obtain viable juvenile tetraploids.

Comparative transcriptome analysis of three gonadal development stages reveals potential genes involved in gametogenesis of the fluted giant clam (Tridacna squamosa).

BACKGROUND: Gonad development and differentiation is an essential function for all sexually reproducing species, and many aspects of these developmental processes are highly conserved among the metazoa. However, the mechanisms underlying gonad development and gametogenesis remain unclear in Tridacna squamosa, a large-size bivalve of great ecological value. They are protandrous simultaneous hermaphrodites, with the male gonad maturing first, eventually followed by the female gonads. In this study, nine gonad libraries representing resting, male and hermaphrodite stages in T. squamosa were performed to identify the molecular mechanisms. RESULTS: Sixteen thousand four hundred ninety-one unigenes were annotated in the NCBI non-redundant protein database. Among the annotated unigenes, 5091 and 7328 unigenes were assigned to Gene Ontology categories and the Kyoto Encyclopedia of Genes and Genomes (KEGG) Pathway database, respectively. A total of 4763 differentially expressed genes (DEGs) were identified by comparing male to resting gonads, consisting of 3499 which were comparatively upregulated in males and 1264 which were downregulated in males. Six hundred-ninteen DEGs between male and hermaphroditic gonads were identified, with 518 DEGs more strongly expressed in hermaphrodites and 101 more strongly expressed in males. GO (Gene Ontology) and KEGG pathway analyses revealed that various biological functions and processes, including functions related to the endocrine system, oocyte meiosis, carbon metabolism, and the cell cycle, were involved in regulating gonadal development and gametogenesis in T. squamosa. Testis-specific serine/threonine kinases 1 (TSSK1), TSSK4, TSSK5, Doublesex- and mab-3-related transcription factor 1 (DMRT1), SOX, Sperm surface protein 17 (SP17) and other genes were involved in male gonadal development in Tridacna squamosal. Both spermatogenesis- (TSSK4, spermatogenesis-associated protein 17, spermatogenesis-associated protein 8, sperm motility kinase X, SP17) and oogenesis-related genes (zona pellucida protein, Forkhead Box L2, Vitellogenin, Vitellogenin receptor, 5-hydroxytryptamine, 5-hydroxytryptamine receptor) were simultaneously highly expressed in the hermaphroditic gonad to maintain the hermaphroditism of T. squamosa. CONCLUSION: All these results from our study will facilitate better understanding of the molecular mechanisms underlying giant clam gonad development and gametogenesis, which can provided a base on obtaining excellent gametes during the seed production process for giant clams.

Doxorubicin Loaded Chitosan-W18 O49 Hybrid Nanoparticles for Combined Photothermal-Chemotherapy.

Combined treatment is more effective than single treatment against most forms of cancer. In this work, doxorubicin loaded chitosan-W18 O49 nanoparticles combined with the photothermal therapy and chemotherapy are fabricated through the electrostatic interaction between positively charged chitosan and negatively charged W18 O49 nanoparticles. The in vitro and in vivo behaviors of these nanoparticles are examined by dynamic light scattering, transmission electron microscopy, cytotoxicity, near-infrared fluorescence imaging, and tumor growth inhibition experiment. These nanoparticles have a mean size around 110 nm and show a pH sensitive drug release behavior. After irradiation by the 980 nm laser, these nanoparticles show more pronounced cytotoxicity against HeLa cells than that of free doxorubicin or photothermal therapy alone. The in vivo experiments confirm that their antitumor ability is significantly improved, resulting in superior efficiency in impeding tumor growth and extension of the lifetime of mice.