miRNA profiling in intrauterine exosomes of pregnant cattle on day 7

Intrauterine exosomes have been identified to be involved in the embryo development and implantation. The aim of this study was to explore the role of miRNAs in intrauterine exosomes in bovine pregnancy. Intrauterine exosomes were collected from uterine flushing fluids of three donor and three recipient Xianan cows 7 days after fertilization. Intrauterine exosomes miRNAs were extracted and the exosomal miRNAs expression levels were analyzed. Sixty miRNAs differed significantly in their amounts between donors and recipients (p-value 1). Twenty-two miRNAs were upregulated and 38 downregulated in the group of donor cows. The bta-miR-184 was the most significant (PBenjamini-Hochberg < 0.001). A total of 9,775 target genes were predicted using the 60 miRNAs. GO and KEGG analysis showed that the target genes were enriched in several biological processes or pathways associated with embryo implantation and endometrial development, such as cell adhesion, cell junction, focal adhesion, and Rap1 signaling pathway. Our findings suggest that, in cattle early pregnancy stage, these differently expressed miRNAs in intrauterine exosomes involved in embryo implantation and endometrial development, which may exert a significant effect and influence the uterine microenvironment for embryo implantation. These results could provide reference for screening and exploring the intrauterine exosomal miRNA affecting embryo implantation

miR-183/96/182 Cluster Regulates the Development of Bovine Myoblasts through Targeting <i>FoxO1</i>

Muscle development is an important factor affecting meat yield and quality and is coordinated by a variety of the myogenic genes and signaling pathways. Recent studies reported that miRNA, a class of highly conserved small noncoding RNA, is actively involved in regulating muscle development, but many miRNAs still need to be further explored. Here, we identified that the miR-183/96/182 cluster exhibited higher expression in bovine embryonic muscle; meanwhile, it widely existed in other organizations. Functionally, the results of the RT-qPCR, EdU, CCK8 and immunofluorescence assays demonstrated that the miR-183/96/182 cluster promoted proliferation and differentiation of bovine myoblast. Next, we found that the miR-183/96/182 cluster targeted FoxO1 and restrained its expression. Meanwhile, the expression of FoxO1 had a negative correlation with the expression of the miR-183/96/182 cluster during myoblast differentiation. In a word, our findings indicated that the miR-183/96/182 cluster serves as a positive regulator in the proliferation and differentiation of bovine myoblasts through suppressing the expression of FoxO1

The evaluation of 23-bp and 12-bp insertion/deletion within the PRNP gene and their effects on growth traits in healthy Chinese native cattle breeds

Polymorphisms in the prion protein gene (PRNP) have been linked with the occurrence of transmissible spongiform encephalopathies and the expression of some phenotypic traits in healthy animals including cattle. The aim of this study was to verify the 23-bp insertion/deletion (indel) within promoter region and the 12-bp indel within intron 1 of PRNP, as well as to evaluate their associations with growth traits. Two loci of PRNP were genotyped in 1558 healthy Chinese cattle sampled from 6 indigenous breeds. Moreover, phenotypic records for growth were used to find the relationship between PRNP polymorphisms and phenotypic performance. The results confirmed the occurrence of these two indel polymorphisms in the analysed breeds. Association analysis showed that the 23-bp indel was significantly related to the body length and heart girth in 18-months-old Nanyang cattle. The 12-bp indel was significantly related to the growth traits in three cattle breeds, such as the body weight in Xia'nan cattle, the daily gained weight in 12-months-old Nanyang cattle and the cannon circumference and rump length in Ji'an cattle. These findings indicated that these two indels may affect bovine growth traits, which could benefit to healthy cattle selection and breeding through marker-assisted selection. Abbreviations AI: artificial insemination; ANOVA: analysis of variance; Bp: base pair; BSE: bovine spongiform encephalopathy; D: deletion; DD: deletion/deletion; GLM: generalized linear model; reaction; He: heterozygosity; Ho: homozygosity; HWE: Hardy–Weinberg equilibrium; I: insertion; ID: insertion/deletion; II: insertion/insertion; Indel: insertion/deletion; MAS: marker-assisted selection; Ne: effective allele numbers; PCR: polymerase chain reaction; PIC: polymorphism information content; PrP: Prion protein (PrP); PRNP: the prion protein; RP58: repressor protein with a predicted molecular mass of 58 kDa; SNP: single nucleotide polymorphism; Sp1: specific protein 1; SPSS: statistical product and service solutions; TSE: transmissible spongiform encephalopathie

Copy Number Variations and Expression Levels of Guanylate-Binding Protein 6 Gene Associated with Growth Traits of Chinese Cattle

Association studies have indicated profound effects of copy number variations (CNVs) on various phenotypes in different species. In this study, we identified the CNV distributions and expression levels of guanylate-binding protein 6 (GBP6) associated with the growth traits of Chinese cattle. The results showed that the phenotypic values of body size and weight of Xianan (XN) cattle were higher than those of Nanyang (NY) cattle. The medium CNV types were mostly identified in the XN and NY breeds, but their CNV distributions were significantly different (adjusted p &lt; 0.05). The association analysis revealed that the body weight, cannon circumference and chest circumference of XN cattle had significantly different values in different CNV types (p &lt; 0.05), with CNV gain types (Log22&minus;&Delta;&Delta;Ct &gt; 0.5) displaying superior phenotypic values. We also found that transcription levels varied in different tissues (p &lt; 0.001) and the CNV gain types showed the highest relative gene expression levels in the muscle tissue, consistent with the highest phenotypic values of body weight and cannon circumference among the three CNV types. Consequently, our results suggested that CNV gain types of GBP6 could be used as the candidate markers in the cattle-breeding program for growth traits

Doublecortin and JIP3 are neural-specific counteracting regulators of dynein-mediated retrograde trafficking

Mutations in the microtubule (MT)-binding protein doublecortin (DCX) or in the MT-based molecular motor dynein result in lissencephaly. However, a functional link between DCX and dynein has not been defined. Here, we demonstrate that DCX negatively regulates dynein-mediated retrograde transport in neurons from Dcx-/y or Dcx-/y;Dclk1-/- mice by reducing dynein’s association with MTs and disrupting the composition of the dynein motor complex. Previous work showed an increased binding of the adaptor protein C-Jun-amino-terminal kinase-interacting protein 3 (JIP3) to dynein in the absence of DCX. Using purified components, we demonstrate that JIP3 forms an active motor complex with dynein and its cofactor dynactin with two dyneins per complex. DCX competes with the binding of the second dynein, resulting in a velocity reduction of the complex. We conclude that DCX negatively regulates dynein-mediated retrograde transport through two critical interactions by regulating dynein binding to MTs and regulating the composition of the dynein motor complex

Global Transcriptome Analysis During Adipogenic Differentiation and Involvement of Transthyretin Gene in Adipogenesis in Cattle

<p>Adipose tissue plays central role in determining the gustatory quality of beef, but traditional Chinese beef cattle have low levels of fat content. We applied RNA-seq to study the molecular mechanisms underlying adipocyte differentiation in Qinchuan cattle. A total of 18,283 genes were found to be expressed in preadipocytes and mature adipocytes, respectively. 470 of which were significantly differentially expressed genes (DEGs) [false discovery rate (FDR) values < 0.05 and fold change ≥ 2]. In addition, 4534 alternative splicing (AS) events and 5153 AS events were detected in preadipocytes and adipocytes, respectively. We constructed a protein interaction network, which suggested that collagen plays an important role during bovine adipogenic differentiation. We characterized the function of the most down-regulated DEG (P < 0.001) among genes we have detected by qPCR, namely, the transthyretin (TTR) gene. Overexpression of TTR appears to promote the expression of the peroxisome proliferator activated receptor γ (PPARγ) (P < 0.05) and fatty acid binding Protein 4 (FABP4) (P < 0.05). Hence, TTR appears to be involved in the regulation of bovine adipogenic differentiation. Our study represents the comprehensive approach to explore bovine adipocyte differentiation using transcriptomic data and reports an involvement of TTR during bovine adipogenic differentiation. Our results provide novel insights into the molecular mechanisms underlying bovine adipogenic differentiation.</p

Spider-Inspired Multicomponent 3D Printing Technique for Next-Generation Complex Biofabrication

The shortage of tissue resources is currently a serious challenge that limits the clinical therapy to patients with tissue loss or end-stage organ failure. The booming development of 3D printing offers unprecedented hope for tissue engineering since it can construct cells and biomaterials into a 3D tissue-mimicking object with precise control over size and shape. However, it is still challenging to fabricate artificial living tissues or organs due to the extreme complexity of biological tissues. Herein, we propose a new concept of spider-inspired 3D printing technique (SI-3DP) for continuous multicomponent 3D printing based on in situ gelation at a multibarrel printing nozzle. The printing process allows for rapid construction of 3D architectures composed of different inks in the desired position. To present the potential in biomedical applications, the SI-DIP also prints vessel-like hollow hydrogel microfibers and cell-laden hollow fibers, indicating good biocompatibility of this technique. The newly developed SI-3DP technique is envisioned to promote the development of next-generation complex biofabrication