Additional file 1 of Impacts of longitudinal water curtain cooling system on transcriptome-related immunity in ducks

Supplementary Material

FLJ25439, a novel cytokinesis-associated protein, induces tetraploidization and maintains chromosomal stability via enhancing expression of endoplasmic reticulum stress chaperones

<p>Investigation of the mechanisms leading to aneuploidy and polyploidy is critical to cancer research. Previous studies have provided strong evidence of the importance of tetraploidization as an early step in tumorigenesis. In cancer cells, tetraploid cells may contribute to abnormal mitotic progression, which may be associated with cytokinesis failure. Tetraploidy leads to genomic instability due to centrosome and chromosome over-replication. Until now, the mechanism by which cells maintain tetraploid status has been unknown. Here, we identified a novel D box-containing protein, FLJ25439, which displays a dynamic expression profile during mitosis/cytokinesis with the midbody as the most prominent associated structure. To understand the function of FLJ25439, we established stable cell lines overexpressing FLJ25439. FLJ25439-overexpression cells grew slower and displayed a tetraploid DNA content in comparison with diploid parental cells. They also showed aberrant mitosis and dysregulated expression of p53, pRb and p21, suggesting a defect in cell cycle progression. To explore the molecular mechanisms responsible for FLJ25439-induced tetraploidization, we conducted a comparative analysis of the global protein expression patterns of wild type and overexpressors using proteomics and bioinformatics approaches. Protein category profiling indicated that FLJ25439 is involved in pathways related to anti-apoptosis, protein folding, the cell cycle, and cytoskeleton regulation. Specifically, genotoxic-stress- and ER stress-related chaperone proteins greatly contributed to the FLJ25439 overexpression phenotypes. The results of this study pave the way to our further understanding of the role of this novel cytokinesis-related protein in protecting cells from environmental stress and tetraploid formation.</p

Table_1_Effects of cage vs. net-floor mixed rearing system on goose spleen histomorphology and gene expression profiles.docx

Due to the demands for both environmental protection and modernization of the goose industry in China, the traditional goose waterside rearing systems have been gradually transitioning to the modern intensive dryland rearing ones, such as the net-floor mixed rearing system (MRS) and cage rearing system (CRS). However, the goose immune responses to different dryland rearing systems remain poorly understood. This study aimed to investigate and compare the age-dependent effects of MRS and CRS on the splenic histomorphological characteristics and immune-related genes expression profiles among three economically important goose breeds, including Sichuan White goose (SW), Gang goose (GE), and Landes goose (LD). Morphological analysis revealed that the splenic weight and organ index of SW were higher under CRS than under MRS (p < 0.05). Histological observations showed that for SW and LD, the splenic corpuscle diameter and area as well as trabecular artery diameter were larger under MRS than under CRS at 30 or 43 weeks of age (p < 0.05), while the splenic red pulp area of GE was larger under CRS than under MRS at 43 weeks of age (p < 0.05). Besides, at 43 weeks of age, higher mRNA expression levels of NGF, SPI1, and VEGFA in spleens of SW were observed under MRS than under CRS (p < 0.05), while higher levels of HSPA2 and NGF in spleens of LD were observed under MRS than under CRS (p < 0.05). For GE, there were higher mRNA expression levels of MYD88 in spleens under CRS at 30 weeks of age (p < 0.05). Moreover, our correlation analysis showed that there appeared to be more pronounced positive associations between the splenic histological parameters and expression levels of several key immune-related genes under MRS than under CRS. Therefore, it is speculated that the geese reared under MRS might exhibit enhanced immune functions than those under CRS, particularly for SW and LD. Although these phenotypic differences are assumed to be associated with the age-dependent differential expression profiles of HSPA2, MYD88, NGF, SPI1, and VEGFA in the goose spleen, the underlying regulatory mechanisms await further investigations.</p

Image_1_Effects of cage vs. net-floor mixed rearing system on goose spleen histomorphology and gene expression profiles.tif

Due to the demands for both environmental protection and modernization of the goose industry in China, the traditional goose waterside rearing systems have been gradually transitioning to the modern intensive dryland rearing ones, such as the net-floor mixed rearing system (MRS) and cage rearing system (CRS). However, the goose immune responses to different dryland rearing systems remain poorly understood. This study aimed to investigate and compare the age-dependent effects of MRS and CRS on the splenic histomorphological characteristics and immune-related genes expression profiles among three economically important goose breeds, including Sichuan White goose (SW), Gang goose (GE), and Landes goose (LD). Morphological analysis revealed that the splenic weight and organ index of SW were higher under CRS than under MRS (p < 0.05). Histological observations showed that for SW and LD, the splenic corpuscle diameter and area as well as trabecular artery diameter were larger under MRS than under CRS at 30 or 43 weeks of age (p < 0.05), while the splenic red pulp area of GE was larger under CRS than under MRS at 43 weeks of age (p < 0.05). Besides, at 43 weeks of age, higher mRNA expression levels of NGF, SPI1, and VEGFA in spleens of SW were observed under MRS than under CRS (p < 0.05), while higher levels of HSPA2 and NGF in spleens of LD were observed under MRS than under CRS (p < 0.05). For GE, there were higher mRNA expression levels of MYD88 in spleens under CRS at 30 weeks of age (p < 0.05). Moreover, our correlation analysis showed that there appeared to be more pronounced positive associations between the splenic histological parameters and expression levels of several key immune-related genes under MRS than under CRS. Therefore, it is speculated that the geese reared under MRS might exhibit enhanced immune functions than those under CRS, particularly for SW and LD. Although these phenotypic differences are assumed to be associated with the age-dependent differential expression profiles of HSPA2, MYD88, NGF, SPI1, and VEGFA in the goose spleen, the underlying regulatory mechanisms await further investigations.</p

Table_1_Comparative transcriptomics analysis identifies crucial genes and pathways during goose spleen development.docx

As the largest peripheral lymphoid organ in poultry, the spleen plays an essential role in regulating the body’s immune capacity. However, compared with chickens and ducks, information about the age- and breed-related changes in the goose spleen remains scarce. In this study, we systematically analyzed and compared the age-dependent changes in the morphological, histological, and transcriptomic characteristics between Landes goose (LG; Anser anser) and Sichuan White goose (SWG; Anser cygnoides). The results showed a gradual increase in the splenic weights for both LG and SWG until week 10, while their splenic organ indexes reached the peak at week 6. Meanwhile, the splenic histological indexes of both goose breeds continuously increased with age, reaching the highest levels at week 30. The red pulp (RP) area was significantly higher in SWG than in LG at week 0, while the splenic corpuscle (AL) diameter was significantly larger in LG than in SWG at week 30. At the transcriptomic level, a total of 1710 and 1266 differentially expressed genes (DEGs) between week 0 and week 30 were identified in spleens of LG and SWG, respectively. Meanwhile, a total of 911 and 808 DEGs in spleens between LG and SWG were identified at weeks 0 and 30, respectively. Both GO and KEGG enrichment analysis showed that the age-related DEGs of LG or SWG were dominantly enriched in the Cell cycle, TGF−beta signaling, and Wnt signaling pathways, while most of the breed-related DEGs were enriched in the Neuroactive ligand−receptor interaction, Cytokine−cytokine receptor interaction, ECM−receptor interaction, and metabolic pathways. Furthermore, through construction of protein-protein interaction networks using significant DEGs, it was inferred that three hub genes including BUB1, BUB1B, and TTK could play crucial roles in regulating age-dependent goose spleen development while GRIA2, GRIA4, and RYR2 could be crucial for the breed-specific goose spleen development. These data provide novel insights into the splenic developmental differences between Chinese and European domestic geese, and the identified crucial pathways and genes are helpful for a better understanding of the mechanisms regulating goose immune functions.</p

Table_2_Effects of cage vs. net-floor mixed rearing system on goose spleen histomorphology and gene expression profiles.docx

Due to the demands for both environmental protection and modernization of the goose industry in China, the traditional goose waterside rearing systems have been gradually transitioning to the modern intensive dryland rearing ones, such as the net-floor mixed rearing system (MRS) and cage rearing system (CRS). However, the goose immune responses to different dryland rearing systems remain poorly understood. This study aimed to investigate and compare the age-dependent effects of MRS and CRS on the splenic histomorphological characteristics and immune-related genes expression profiles among three economically important goose breeds, including Sichuan White goose (SW), Gang goose (GE), and Landes goose (LD). Morphological analysis revealed that the splenic weight and organ index of SW were higher under CRS than under MRS (p < 0.05). Histological observations showed that for SW and LD, the splenic corpuscle diameter and area as well as trabecular artery diameter were larger under MRS than under CRS at 30 or 43 weeks of age (p < 0.05), while the splenic red pulp area of GE was larger under CRS than under MRS at 43 weeks of age (p < 0.05). Besides, at 43 weeks of age, higher mRNA expression levels of NGF, SPI1, and VEGFA in spleens of SW were observed under MRS than under CRS (p < 0.05), while higher levels of HSPA2 and NGF in spleens of LD were observed under MRS than under CRS (p < 0.05). For GE, there were higher mRNA expression levels of MYD88 in spleens under CRS at 30 weeks of age (p < 0.05). Moreover, our correlation analysis showed that there appeared to be more pronounced positive associations between the splenic histological parameters and expression levels of several key immune-related genes under MRS than under CRS. Therefore, it is speculated that the geese reared under MRS might exhibit enhanced immune functions than those under CRS, particularly for SW and LD. Although these phenotypic differences are assumed to be associated with the age-dependent differential expression profiles of HSPA2, MYD88, NGF, SPI1, and VEGFA in the goose spleen, the underlying regulatory mechanisms await further investigations.</p

Table_2_Comparative transcriptomics analysis identifies crucial genes and pathways during goose spleen development.docx

As the largest peripheral lymphoid organ in poultry, the spleen plays an essential role in regulating the body’s immune capacity. However, compared with chickens and ducks, information about the age- and breed-related changes in the goose spleen remains scarce. In this study, we systematically analyzed and compared the age-dependent changes in the morphological, histological, and transcriptomic characteristics between Landes goose (LG; Anser anser) and Sichuan White goose (SWG; Anser cygnoides). The results showed a gradual increase in the splenic weights for both LG and SWG until week 10, while their splenic organ indexes reached the peak at week 6. Meanwhile, the splenic histological indexes of both goose breeds continuously increased with age, reaching the highest levels at week 30. The red pulp (RP) area was significantly higher in SWG than in LG at week 0, while the splenic corpuscle (AL) diameter was significantly larger in LG than in SWG at week 30. At the transcriptomic level, a total of 1710 and 1266 differentially expressed genes (DEGs) between week 0 and week 30 were identified in spleens of LG and SWG, respectively. Meanwhile, a total of 911 and 808 DEGs in spleens between LG and SWG were identified at weeks 0 and 30, respectively. Both GO and KEGG enrichment analysis showed that the age-related DEGs of LG or SWG were dominantly enriched in the Cell cycle, TGF−beta signaling, and Wnt signaling pathways, while most of the breed-related DEGs were enriched in the Neuroactive ligand−receptor interaction, Cytokine−cytokine receptor interaction, ECM−receptor interaction, and metabolic pathways. Furthermore, through construction of protein-protein interaction networks using significant DEGs, it was inferred that three hub genes including BUB1, BUB1B, and TTK could play crucial roles in regulating age-dependent goose spleen development while GRIA2, GRIA4, and RYR2 could be crucial for the breed-specific goose spleen development. These data provide novel insights into the splenic developmental differences between Chinese and European domestic geese, and the identified crucial pathways and genes are helpful for a better understanding of the mechanisms regulating goose immune functions.</p

Image_1_Comparative transcriptomics analysis identifies crucial genes and pathways during goose spleen development.tif

As the largest peripheral lymphoid organ in poultry, the spleen plays an essential role in regulating the body’s immune capacity. However, compared with chickens and ducks, information about the age- and breed-related changes in the goose spleen remains scarce. In this study, we systematically analyzed and compared the age-dependent changes in the morphological, histological, and transcriptomic characteristics between Landes goose (LG; Anser anser) and Sichuan White goose (SWG; Anser cygnoides). The results showed a gradual increase in the splenic weights for both LG and SWG until week 10, while their splenic organ indexes reached the peak at week 6. Meanwhile, the splenic histological indexes of both goose breeds continuously increased with age, reaching the highest levels at week 30. The red pulp (RP) area was significantly higher in SWG than in LG at week 0, while the splenic corpuscle (AL) diameter was significantly larger in LG than in SWG at week 30. At the transcriptomic level, a total of 1710 and 1266 differentially expressed genes (DEGs) between week 0 and week 30 were identified in spleens of LG and SWG, respectively. Meanwhile, a total of 911 and 808 DEGs in spleens between LG and SWG were identified at weeks 0 and 30, respectively. Both GO and KEGG enrichment analysis showed that the age-related DEGs of LG or SWG were dominantly enriched in the Cell cycle, TGF−beta signaling, and Wnt signaling pathways, while most of the breed-related DEGs were enriched in the Neuroactive ligand−receptor interaction, Cytokine−cytokine receptor interaction, ECM−receptor interaction, and metabolic pathways. Furthermore, through construction of protein-protein interaction networks using significant DEGs, it was inferred that three hub genes including BUB1, BUB1B, and TTK could play crucial roles in regulating age-dependent goose spleen development while GRIA2, GRIA4, and RYR2 could be crucial for the breed-specific goose spleen development. These data provide novel insights into the splenic developmental differences between Chinese and European domestic geese, and the identified crucial pathways and genes are helpful for a better understanding of the mechanisms regulating goose immune functions.</p

Table_3_Comparative transcriptomics analysis identifies crucial genes and pathways during goose spleen development.docx

As the largest peripheral lymphoid organ in poultry, the spleen plays an essential role in regulating the body’s immune capacity. However, compared with chickens and ducks, information about the age- and breed-related changes in the goose spleen remains scarce. In this study, we systematically analyzed and compared the age-dependent changes in the morphological, histological, and transcriptomic characteristics between Landes goose (LG; Anser anser) and Sichuan White goose (SWG; Anser cygnoides). The results showed a gradual increase in the splenic weights for both LG and SWG until week 10, while their splenic organ indexes reached the peak at week 6. Meanwhile, the splenic histological indexes of both goose breeds continuously increased with age, reaching the highest levels at week 30. The red pulp (RP) area was significantly higher in SWG than in LG at week 0, while the splenic corpuscle (AL) diameter was significantly larger in LG than in SWG at week 30. At the transcriptomic level, a total of 1710 and 1266 differentially expressed genes (DEGs) between week 0 and week 30 were identified in spleens of LG and SWG, respectively. Meanwhile, a total of 911 and 808 DEGs in spleens between LG and SWG were identified at weeks 0 and 30, respectively. Both GO and KEGG enrichment analysis showed that the age-related DEGs of LG or SWG were dominantly enriched in the Cell cycle, TGF−beta signaling, and Wnt signaling pathways, while most of the breed-related DEGs were enriched in the Neuroactive ligand−receptor interaction, Cytokine−cytokine receptor interaction, ECM−receptor interaction, and metabolic pathways. Furthermore, through construction of protein-protein interaction networks using significant DEGs, it was inferred that three hub genes including BUB1, BUB1B, and TTK could play crucial roles in regulating age-dependent goose spleen development while GRIA2, GRIA4, and RYR2 could be crucial for the breed-specific goose spleen development. These data provide novel insights into the splenic developmental differences between Chinese and European domestic geese, and the identified crucial pathways and genes are helpful for a better understanding of the mechanisms regulating goose immune functions.</p