KED gene expression in early response to wounding stress in tomato plants.

The wounding-responsive KED gene, named for its coding for a lysine (K), glutamic acid (E), and aspartic acid (D)-rich protein, is widely present among land plants. However, little is known about its regulation or function. In this study, we found that transcription of the tomato (Solanum lycopersicum) KED gene, SlKED, was rapidly and transiently elevated by wounding or ethephon treatment. Compared to the wild-type plants, the CRISPR/Cas9-mediated SlKED knockout plants did not exhibit altered expression patterns for genes involved in hormone biosynthesis or stress signaling, suggesting a lack of pleiotropic effect on other stress-responsive genes. Conversely, jasmonic acid did not appear to directly regulate SlKED expression. Wounded leaves of the KED-lacking plants exhibited higher binding of Evans blue dye than the wild-type, indicating a possible role for KED in healing damaged tissues. The SlKED knockout plants showed a similar dietary effect as the wild-type on the larval growth of tobacco hornworm. But a higher frequency of larval mandible (mouth) movement was recorded during the first 2 minutes of feeding on the wounded KED-lacking SlKED knockout plants than on the wounded KED-producing wild-type plants, probably reflecting an initial differential response by the feeding larvae to the SlKED knockout plants. Our findings suggest that SlKED may be an ethylene-mediated early responder to mechanical stress in tomato, acting downstream of the wound stress response pathways. Although its possible involvement in response to other biotic and abiotic stresses is still unclear, we propose that SlKED may play a role in plant's rapid, short-term, early wounding responses, such as in cellular damage healing.

Spike protein recognition of mammalian ACE2 predicts the host range and an optimized ACE2 for SARS-CoV-2 infection.

SARS-CoV-2 causes the recent global COVID-19 public health emergency. ACE2 is the receptor for both SARS-CoV-2 and SARS-CoV. To predict the potential host range of SARS-CoV-2, we analyzed the key residues of ACE2 for recognizing S protein. We found that most of the selected mammals including pets (dog and cat), pangolin and Circetidae mammals remained the most of key residues for association with S protein from SARS-CoV and SARS-CoV-2. The interaction interface between cat/dog/pangolin/Chinese hamster ACE2 and SARS-CoV/SARS-CoV-2 S protein was simulated through homology modeling. We identified that N82 in ACE2 showed a closer contact with SARS-CoV-2 S protein than M82 in human ACE2. Our finding will provide important insights into the host range of SARS-CoV-2 and a new strategy to design an optimized ACE2 for SARS-CoV-2 infection.

SARS-CoV-2 spike protein favors ACE2 from Bovidae and Cricetidae.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causes the recent COVID-19 public health crisis. Bat is the widely believed original host of SARS-CoV-2. However, its intermediate host before transmitting to humans is not clear. Some studies proposed pangolin, snake, or turtle as the intermediate hosts. Angiotensin-converting enzyme 2 (ACE2) is the receptor for SARS-CoV-2, which determines the potential host range for SARS-CoV-2. On the basis of structural information of the complex of human ACE2 and SARS-CoV-2 receptor-binding domain (RBD), we analyzed the affinity to S protein of the 20 key residues in ACE2 from mammal, bird, turtle, and snake. Several ACE2 proteins from Primates, Bovidae, Cricetidae, and Cetacea maintained the majority of key residues in ACE2 for associating with SARS-CoV-2 RBD. The simulated structures indicated that ACE2 proteins from Bovidae and Cricetidae were able to associate with SARS-CoV-2 RBD. We found that nearly half of the key residues in turtle, snake, and bird were changed. The simulated structures showed several key contacts with SARS-CoV-2 RBD in turtle and snake ACE2 were abolished. This study demonstrated that neither snake nor turtle was the intermediate hosts for SARS-CoV-2, which further reinforced the concept that the reptiles are resistant against infection of coronavirus. This study suggested that Bovidae and Cricetidae should be included in the screening of intermediate hosts for SARS-CoV-2.

Novel Regulatory Mechanisms for the SoxC Transcriptional Network Required for Visual Pathway Development.

What pathways specify retinal ganglion cell (RGC) fate in the developing retina? Here we report on mechanisms by which a molecular pathway involving Sox4/Sox11 is required for RGC differentiation and for optic nerve formation in mice in vivo, and is sufficient to differentiate human induced pluripotent stem cells into electrophysiologically active RGCs. These data place Sox4 downstream of RE1 silencing transcription factor in regulating RGC fate, and further describe a newly identified, Sox4-regulated site for post-translational modification with small ubiquitin-related modifier (SUMOylation) in Sox11, which suppresses Sox11's nuclear localization and its ability to promote RGC differentiation, providing a mechanism for the SoxC familial compensation observed here and elsewhere in the nervous system. These data define novel regulatory mechanisms for this SoxC molecular network, and suggest pro-RGC molecular approaches for cell replacement-based therapies for glaucoma and other optic neuropathies.SIGNIFICANCE STATEMENT Glaucoma is the most common cause of blindness worldwide and, along with other optic neuropathies, is characterized by loss of retinal ganglion cells (RGCs). Unfortunately, vision and RGC loss are irreversible, and lead to bilateral blindness in ∼14% of all diagnosed patients. Differentiated and transplanted RGC-like cells derived from stem cells have the potential to replace neurons that have already been lost and thereby to restore visual function. These data uncover new mechanisms of retinal progenitor cell (RPC)-to-RGC and human stem cell-to-RGC fate specification, and take a significant step toward understanding neuronal and retinal development and ultimately cell-transplant therapy.

The Use of "Omics" in Lactation Research in Dairy Cows.

"Omics" is the application of genomics, transcriptomics, proteomics, and metabolomics in biological research. Over the years, tremendous amounts of biological information has been gathered regarding the changes in gene, mRNA and protein expressions as well as metabolites in different physiological conditions and regulations, which has greatly advanced our understanding of the regulation of many physiological and pathophysiological processes. The aim of this review is to comprehensively describe the advances in our knowledge regarding lactation mainly in dairy cows that were obtained from the "omics" studies. The "omics" technologies have continuously been preferred as the technical tools in lactation research aiming to develop new nutritional, genetic, and management strategies to improve milk production and milk quality in dairy cows.

Enantioselective cascade biocatalysis for deracemization of 2-hydroxy acids using a three-enzyme system.

BACKGROUND: Enantiopure 2-hydroxy acids are key intermediates for the synthesis of pharmaceuticals and fine chemicals. We present an enantioselective cascade biocatalysis using recombinant microbial cells for deracemization of racemic 2-hydroxy acids that allows for efficient production of enantiopure 2-hydroxy acids. RESULTS: The method was realized by a single recombinant Escherichia coli strain coexpressing three enzymes: (S)-2-hydroxy acid dehydrogenase, (R)-2-keto acid reductase and glucose dehydrogenase. One enantiomer [(S)-2-hydroxy acid] is firstly oxidized to the keto acid with (S)-2-hydroxy acid dehydrogenase, while the other enantiomer [(R)-2-hydroxy acid] remains unchanged. Then, the keto acid obtained reduced to the opposite enantiomer with (R)-2-keto acid reductase plus cofactor regeneration enzyme glucose dehydrogenase subsequently. The recombinant E. coli strain coexpressing the three enzymes was proven to be a promising biocatalyst for the cascade bioconversion of a structurally diverse range of racemic 2-hydroxy acids, giving the corresponding (R)-2-hydroxy acids in up to 98.5 % conversion and >99 % enantiomeric excess. CONCLUSIONS: In summary, a cascade biocatalysis was successfully developed to prepare valuable (R)-2-hydroxy acids with an efficient three-enzyme system. The developed elegant cascade biocatalysis possesses high atom efficiency and represents a promising strategy for production of highly valued (R)-2-hydroxy acids.

Effects of Chinese Propolis in Protecting Bovine Mammary Epithelial Cells against Mastitis Pathogens-Induced Cell Damage.

Chinese propolis (CP), an important hive product, can alleviate inflammatory responses. However, little is known regarding the potential of propolis treatment for mastitis control. To investigate the anti-inflammatory effects of CP on bovine mammary epithelial cells (MAC-T), we used a range of pathogens to induce cellular inflammatory damage. Cell viability was determined and expressions of inflammatory/antioxidant genes were measured. Using a cell-based reporter assay system, we evaluated CP and its primary constituents on the NF-κB and Nrf2-ARE transcription activation. MAC-T cells treated with bacterial endotoxin (lipopolysaccharide, LPS), heat-inactivated Escherichia coli, and Staphylococcus aureus exhibited significant decreases in cell viability while TNF-α and lipoteichoic acid (LTA) did not. Pretreatment with CP prevented losses in cell viability associated with the addition of killed bacteria or bacterial endotoxins. There were also corresponding decreases in expressions of proinflammatory IL-6 and TNF-α mRNA. Compared with the mastitis challenged cells, enhanced expressions of antioxidant genes HO-1, Txnrd-1, and GCLM were observed in CP-treated cells. CP and its polyphenolic active components (primarily caffeic acid phenethyl ester and quercetin) had strong inhibitive effects against NF-κB activation and increased the transcriptional activity of Nrf2-ARE. These findings suggest that propolis may be valuable in the control of bovine mastitis.

Nanoparticle-mediated signaling endosome localization regulates growth cone motility and neurite growth.

Understanding neurite growth regulation remains a seminal problem in neurobiology. During development and regeneration, neurite growth is modulated by neurotrophin-activated signaling endosomes that transmit regulatory signals between soma and growth cones. After injury, delivering neurotrophic therapeutics to injured neurons is limited by our understanding of how signaling endosome localization in the growth cone affects neurite growth. Nanobiotechnology is providing new tools to answer previously inaccessible questions. Here, we show superparamagnetic nanoparticles (MNPs) functionalized with TrkB agonist antibodies are endocytosed into signaling endosomes by primary neurons that activate TrkB-dependent signaling, gene expression and promote neurite growth. These MNP signaling endosomes are trafficked into nascent and existing neurites and transported between somas and growth cones in vitro and in vivo. Manipulating MNP-signaling endosomes by a focal magnetic field alters growth cone motility and halts neurite growth in both peripheral and central nervous system neurons, demonstrating signaling endosome localization in the growth cone regulates motility and neurite growth. These data suggest functionalized MNPs may be used as a platform to study subcellular organelle localization and to deliver nanotherapeutics to treat injury or disease in the central nervous system.

Enzymatically prepared neoagarooligosaccharides improve gut health and function through promoting the production of spermidine by Faecalibacterium in chickens.

Maintaining animal gut health through modulating the gut microbiota is a constant need when antibiotics are not used in animal feed during the food animal production process. Prebiotics is regarded as one of the most promising antibiotic alternatives for such purpose. As an attractive prebiotic, the role and mechanisms of neoagarooligosaccharides (NAOS) in promoting animal growth and gut health have not been elucidated. In this study, we first cloned and expressed marine bacterial ß-agarase in yeast to optimize the NAOS preparation and then investigated the role and the underlying mechanisms of the prepared NAOS in improving chicken gut health and function. The marine bacterial ß-agarase PDE13B was expressed in Pichia pastoris GS115 and generated even-numbered NAOS. Dietary the prepared NAOS promoted chicken growth and improved intestinal morphology, its barrier, and digestion capabilities, and absorption function. Metagenomic analysis indicated that NAOS modulated the chicken gut microbiota structure and function, and microbial interactions, and promoted the growth of spermidine-producing bacteria especially Faecalibacterium. Through integration of gut metagenome, gut content metabolome, and gut tissue transcriptome, we established connections among NAOS, gut microbes, spermidine, and chicken gut gene expression. The spermidine regulation of genes related to autophagy, immunity, and inflammation was further confirmed in chicken embryo intestinal epithelium cells. We also verified that NAOS can be utilized by Faecalibacterium prausnitzii to grow and produce spermidine in in vitro experiments. Collectively, we provide a systematic investigation of the role of NAOS in regulating gut health and demonstrate the microbial spermidine-mediated mechanism involved in prebiotic effects of NAOS, which lays foundation for future use of NAOS as a new antibiotic alternative in animal production.

Soluble adenylyl cyclase activity is necessary for retinal ganglion cell survival and axon growth.

cAMP is a critical second messenger mediating activity-dependent neuronal survival and neurite growth. We investigated the expression and function of the soluble adenylyl cyclase (sAC, ADCY10) in CNS retinal ganglion cells (RGCs). We found sAC protein expressed in multiple RGC compartments including the nucleus, cytoplasm and axons. sAC activation increased cAMP above the level seen with transmembrane adenylate cyclase (tmAC) activation. Electrical activity and bicarbonate, both physiologic sAC activators, significantly increased survival and axon growth, whereas pharmacologic or siRNA-mediated sAC inhibition dramatically decreased RGC survival and axon growth in vitro, and survival in vivo. Conversely, RGC survival and axon growth were unaltered in RGCs from AC1/AC8 double knock-out mice or after specifically inhibiting tmACs. These data identify a novel sAC-mediated cAMP signaling pathway regulating RGC survival and axon growth, and suggest new neuroprotective or regenerative strategies based on sAC modulation.

[Progress on the miRNA related with mammary gland development and lactation].

MicroRNAs (miRNAs) are non-coding RNAs that play important roles in post transcriptional regulation. They are involved in the regulation of mammary gland development and lactation. In this paper, we summarized the expression pattern of miRNAs which varied with tissues and lactation stages, and the functions of several miRNAs are also briefly reviewed. The objective of this work is to give reference for further study on miRNAs in mammary gland, and to provide theoretical basis and ideas for the use of miRNAs in improving healthy development of mammary gland and regulating the efficiency of lactation and the quality of milk.

Ginkgo biloba extract alleviates fatty liver hemorrhagic syndrome in laying hens via reshaping gut microbiota.

BACKGROUND: Ginkgo biloba extract (GBE) is evidenced to be effective in the prevention and alleviation of metabolic disorders, including obesity, diabetes and fatty liver disease. However, the role of GBE in alleviating fatty liver hemorrhagic syndrome (FLHS) in laying hens and the underlying mechanisms remain to be elucidated. Here, we investigated the effects of GBE on relieving FLHS with an emphasis on the modulatory role of GBE in chicken gut microbiota. RESULTS: The results showed that GBE treatment ameliorated biochemical blood indicators in high-fat diet (HFD)-induced FLHS laying hen model by decreasing the levels of TG, TC, ALT and ALP. The lipid accumulation and pathological score of liver were also relieved after GBE treatment. Moreover, GBE treatment enhanced the antioxidant activity of liver and serum by increasing GSH, SOD, T-AOC, GSH-PX and reducing MDA, and downregulated the expression of genes related to lipid synthesis (FAS, LXRα, GPAT1, PPARγ and ChREBP1) and inflammatory cytokines (TNF-α, IL-6, TLR4 and NF-κB) in the liver. Microbial profiling analysis revealed that GBE treatment reshaped the HFD-perturbed gut microbiota, particularly elevated the abundance of Megasphaera in the cecum. Meanwhile, targeted metabolomic analysis of SCFAs revealed that GBE treatment significantly promoted the production of total SCFAs, acetate and propionate, which were positively correlated with the GBE-enriched gut microbiota. Finally, we confirmed that the GBE-altered gut microbiota was sufficient to alleviate FLHS by fecal microbiota transplantation (FMT). CONCLUSIONS: We provided evidence that GBE alleviated FLHS in HFD-induced laying hens through reshaping the composition of gut microbiota. Our findings shed light on mechanism underlying the anti-FLHS efficacy of GBE and lay foundations for future use of GBE as additive to prevent and control FLHS in laying hen industry.

Evolutionary analysis of KED-rich proteins in plants.

During the course of evolution, organisms have developed genetic mechanisms in response to various environmental stresses including wounding from mechanical damage or herbivory-caused injury. A previous study of wounding response in the plant tobacco identified a unique wound-induced gene, aptly named KED due to its coding for a protein that has an unusually high content of amino acids lysine (K), glutamic acid (E) and aspartic acid (D). However, by far little is known about this intriguing gene. In this study, we investigated the evolutionary aspects of the KED-rich coding genes. We found that a consistent pattern of wound-induced KED gene expression is maintained across representative species of angiosperm and gymnosperm. KED genes can be identified in species from all groups of land plants (Embryophyta). All the KED proteins from vascular plants (Tracheophyta) including angiosperm, gymnosperm, fern and lycophyte share a conserved 19-amino acid domain near the C-terminus, whereas bryophytes (moss, liverwort and hornwort) possess KED-rich, multi-direct-repeat sequences that are distinct from the vascular plant KEDs. We detected KED-rich sequences in Charophyta species but not in Chlorophyta wherever genome sequences are available. Our studies suggest diverse and complex evolution pathways for land plant KED genes. Vascular plant KEDs exhibit high evolutionary conservation, implicating their shared function in response to wounding stress. The extraordinary enrichment of amino acids K, E and D in these groups of distinct and widely distributed proteins may reflect the structural and functional requirement for these three residues during some 600 million years of land plant evolution.

Untargeted metabolomics description of propolis's in vitro antibacterial mechanisms against Clostridium perfringens.

Propolis is a natural resinous substance that is collected by honeybees (Apis mellifera) with promising antibacterial effects. Here, we examined the antibacterial activity of Chinese propolis against Clostridium perfringens, a bacterial pathogen that threatens food safety and causes intestinal erosion. The inhibitory effects of the ethanolic extract of Chinese propolis (CPE) on human-associated C. perfringens strains were determined by using the circle of inhibition, the minimum inhibitory concentrations, and bactericidal concentrations. CPE also induced morphological elongation, bacterial cell wall damage, and intracellular material leakage in C. perfringens. Untargeted HPLC-qTOF-MS-based metabolomics analysis of the bacterial metabolic compounds revealed that propolis triggered glycerophospholipid metabolism, one carbon pool by folate, and d-glutamine and d-glutamate metabolism alterations in C. perfringens. Finally, caffeic acid phenethyl ester was identified as the key active ingredient in CPE. This study suggested the usage of propolis as an alternative to antibiotics in controlling C. perfringens.

Mining chicken ileal microbiota for immunomodulatory microorganisms.

The gut microbiota makes important contributions to host immune system development and resistance to pathogen infections, especially during early life. However, studies addressing the immunomodulatory functions of gut microbial individuals or populations are limited. In this study, we explore the systemic impact of the ileal microbiota on immune cell development and function of chickens and identify the members of the microbiota involved in immune system modulation. We initially used a time-series design with six time points to prove that ileal microbiota at different succession stages is intimately connected to immune cell maturation. Antibiotics perturbed the microbiota succession and negatively affected immune development, whereas early exposure to the ileal commensal microbiota from more mature birds promoted immune cell development and facilitated pathogen elimination after Salmonella Typhimurium infection, illustrating that early colonization of gut microbiota is an important driver of immune development. Five bacterial strains, Blautia coccoides, Bacteroides xylanisolvens, Fournierella sp002159185, Romboutsia lituseburensis, and Megamonas funiformis, which are closely related to the immune system development of broiler chickens, were then screened out and validated for their immunomodulatory properties. Our results provide insight into poultry immune system-microbiota interactions and also establish a foundation for targeted immunological interventions aiming to combat infectious diseases and promote poultry health and production.

Enzymatically prepared alginate oligosaccharides improve broiler chicken growth performance by modulating the gut microbiota and growth hormone signals.

BACKGROUND: Alginate oligosaccharide (AOS) holds great potential as a novel feed supplement in farm animals. However, the effects of AOS on chicken health and the underlying mechanisms are not fully understood. This study aimed to optimize the enzymatic preparation of AOS by using bacterial alginate lyases expressed in yeast, investigate the effects of the prepared AOS on the growth performance and gut health of broiler chickens, and reveal the underlying mechanisms. RESULTS: Five alginate lyases from bacteria were cloned into Pichia pastoris GS115 and the alginate lyase PDE9 was expressed at relatively high yield, activity and stability in P. pastoris. Animal trials were carried out using 320 1-day-old male Arbor Acres broilers (four groups; 8 replicates/group × 10 chicks/replicate) receiving either a basal diet or the same diet supplemented with 100, 200 and 400 mg/kg PDE9-prepared AOS for 42 d. The results showed that dietary supplementation of 200 mg/kg AOS displayed the highest activity in promoting the birds' ADG and ADFI (P < 0.05). AOS ameliorated the intestinal morphology, absorption function and barrier function, as indicated by the enhanced (P < 0.05) intestinal villus height, maltase activity, and the expression of PEPT, SGLT1, ZNT1, and occludin. AOS also increased serum insulin-like growth factor-1, ghrelin (P < 0.05), and growth hormone (P < 0.1). Moreover, the concentrations of acetate, isobutyrate, isovalerate, valerate, and total SCFAs in cecum of birds fed AOS were significantly higher than the control birds (P < 0.05). Metagenomic analysis indicated that AOS modulated the chicken gut microbiota structure, function, and microbial interactions and promoted the growth of SCFAs-producing bacteria, for example, Dorea sp. 002160985; SCFAs, especially acetate, were found positively correlated with the chicken growth performance and growth-related hormone signals (P < 0.05). We further verified that AOS can be utilized by Dorea sp. to grow and to produce acetate in vitro. CONCLUSIONS: We demonstrated that the enzymatically produced AOS effectively promoted broiler chicken growth performance by modulating the chicken gut microbiota structure and function. For the first time, we established the connections among AOS, chicken gut microbiota/SCFAs, growth hormone signals and chicken growth performance.

Inulin-enriched Megamonas funiformis ameliorates metabolic dysfunction-associated fatty liver disease by producing propionic acid.

Accumulated evidence supports the beneficial role of inulin in alleviating metabolic dysfunction-associated fatty liver disease (MAFLD) by modulating gut microbiota. However, the underlying mechanisms are not fully understood. Here we used high-fat diet (HFD)-induced laying hen model of MAFLD to investigate the effect of inulin on ameliorating MAFLD and found that the inulin-enriched Megamonas genus was inversely correlated with hepatic steatosis-related parameters. Oral administration of a newly isolated commensal bacterium by culturomics, M. funiformis CML154, to HFD-fed hens and mice ameliorated MAFLD, changed liver gene expression profiles, and increased intestinal propionate concentration. Further evidence demonstrated that the anti-MAFLD effect of M. funiformis CML154 is attributed to propionate-mediated activation of the APN-AMPK-PPARα signaling pathway, thereby inhibiting fatty acid de novo synthesis and promoting ß-oxidation. These findings establish the causal relationships among inulin, M. funiformis, and MAFLD, and suggest that M. funiformis CML154 is a probiotic candidate for preventative or therapeutic intervention of MAFLD.

Integrated multi-omics reveals the roles of cecal microbiota and its derived bacterial consortium in promoting chicken growth.

IMPORTANCE: The improvement of chicken growth performance is one of the major concerns for the poultry industry. Gut microbes are increasingly evidenced to be associated with chicken physiology and metabolism, thereby influencing chicken growth and development. Here, through integrated multi-omics analyses, we showed that chickens from the same line differing in their body weight were very different in their gut microbiota structure and host-microbiota crosstalk; microbes in high body weight (HBW) chickens contributed to chicken growth by regulating the gut function and homeostasis. We also verified that a specific bacterial consortium consisting of isolates from the HBW chickens has the potential to be used as chicken growth promoters. These findings provide new insights into the potential links between gut microbiota and chicken phenotypes, shedding light on future manipulation of chicken gut microbiota to improve chicken growth performance.

Quantitative microbiome profiling reveals the developmental trajectory of the chicken gut microbiota and its connection to host metabolism.

Revealing the assembly and succession of the chicken gut microbiota is critical for a better understanding of its role in chicken physiology and metabolism. However, few studies have examined dynamic changes of absolute chicken gut microbes using the quantitative microbiome profiling (QMP) method. Here, we revealed the developmental trajectory of the broiler chicken gut bacteriome and mycobiome by combining high-throughput sequencing with a microbial load quantification assay. We showed that chicken gut microbiota abundance and diversity reached a plateau at 7 days posthatch (DPH), forming segment-specific community types after 1 DPH. The bacteriome was more impacted by deterministic processes, and the mycobiome was more affected by stochastic processes. We also observed stage-specific microbes in different gut segments, and three microbial occurrence patterns including "colonization," "disappearance," and "core" were defined. The microbial co-occurrence networks were very different among gut segments, with more positive associations than negative associations. Furthermore, we provided links between the absolute changes in chicken gut microbiota and their serum metabolite variations. Time-course untargeted metabolomics revealed six metabolite clusters with different changing patterns of abundance. The foregut microbiota had more connections with chicken serum metabolites, and the gut microbes were closely related to chicken lipid and amino acid metabolism. The present study provided a full landscape of chicken gut microbiota development in a quantitative manner, and the associations between gut microbes and chicken serum metabolites further highlight the impact of gut microbiota in chicken growth and development.

Expression profiles of microRNAs from lactating and non-lactating bovine mammary glands and identification of miRNA related to lactation.

BACKGROUND: MicroRNAs (miRNAs) have been implicated in the regulation of milk protein synthesis and development of the mammary gland (MG). However, the specific functions of miRNAs in these regulations are not clear. Therefore, the elucidation of miRNA expression profiles in the MG is an important step towards understanding the mechanisms of lactogenesis. RESULTS: Two miRNA libraries were constructed from MG tissues taken from a lactating and a non-lactating Holstein dairy cow, respectively, and the short RNA sequences (18-30 nt) in these libraries were sequenced by Solexa sequencing method. The libraries included 885 pre-miRNAs encoding for 921 miRNAs, of which 884 miRNAs were unique sequences and 544 (61.5%) were expressed in both periods. A custom-designed microarray assay was then performed to compare miRNA expression patterns in the MG of lactating and non-lactating dairy cows. A total of 56 miRNAs in the lactating MG showed significant differences in expression compared to non-lactating MG (P<0.05). Integrative miRNA target prediction and network analysis approaches were employed to construct an interaction network of lactation-related miRNAs and their putative targets. Using a cell-based model, six miRNAs (miR-125b, miR-141, miR-181a, miR-199b, miR-484 and miR-500) were studied to reveal their possible biological significance. CONCLUSION: Our study provides a broad view of the bovine MG miRNA expression profile characteristics. Eight hundred and eighty-four miRNAs were identified in bovine MG. Differences in types and expression levels of miRNAs were observed between lactating and non-lactating bovine MG. Systematic predictions aided in the identification of lactation-related miRNAs, providing insight into the types of miRNAs and their possible mechanisms in regulating lactation.