Temporal and spatial expression of Smads and their correlation with YAP1 expression in sheep.

The temporal and spatial patterns of Smad and Yes-associated protein 1 (YAP1) expression were investigated in skeletal muscle (gastrocnemius muscle and extensor digitorum longus) at different growth stages (2 days old, 2 and 6 months old) in Hu sheep. Smads were differentially expressed in sheep skeletal muscle, with high expression in the gastrocnemius muscle and lower expression in the extensor digitorum longus. Expression of Smad2, Smad3, and Smad4 at the 2-day-old stage was significantly higher than at other stages (P < 0.05). The expression of Smad7 in 2-day-old sheep was lower than in 6-month-old sheep, with the lowest levels at 2 months. Smad expression was higher in males than in females at the 2-day-old stage, and expression in 2- and 6-month-old males was lower than that in 2-day-old females. Smad3 expression was higher in the 2-day- and 2-month-old males than in the females. There was a positive correlation (P < 0.01) between YAP1 and Smad2 expression in gastrocnemius muscle at the 2-month-old stage. YAP1 and Smad4/7 expression were positively correlated (P < 0.01) in extensor digitorum longus at the 2-day-old stage. YAP1 expression was negatively correlated with Smad7 in the extensor digitorum longus at 6 months. A significant difference between Smad2 and Smad3 (P < 0.01) expression in muscle was observed, consistent with Smad3 and Smad4 expression, indicating that these inhibit transforming growth factor-ß signaling in the same way. There was a positive correlation (P < 0.01) between YAP1 and MSTN expression, suggesting that YAP1 participates in muscle growth in sheep.

Discovery of differentially expressed genes in cashmere goat (Capra hircus) hair follicles by RNA sequencing.

The mammalian hair follicle (HF) is a unique, highly regenerative organ with a distinct developmental cycle. Cashmere goat (Capra hircus) HFs can be divided into two categories based on structure and development time: primary and secondary follicles. To identify differentially expressed genes (DEGs) in the primary and secondary HFs of cashmere goats, the RNA sequencing of six individuals from Arbas, Inner Mongolia, was performed. A total of 617 DEGs were identified; 297 were upregulated while 320 were downregulated. Gene ontology analysis revealed that the main functions of the upregulated genes were electron transport, respiratory electron transport, mitochondrial electron transport, and gene expression. The downregulated genes were mainly involved in cell autophagy, protein complexes, neutrophil aggregation, and bacterial fungal defense reactions. According to the Kyoto Encyclopedia of Genes and Genomes database, these genes are mainly involved in the metabolism of cysteine and methionine, RNA polymerization, and the MAPK signaling pathway, and were enriched in primary follicles. A microRNA-target network revealed that secondary follicles are involved in several important biological processes, such as the synthesis of keratin-associated proteins and enzymes involved in amino acid biosynthesis. In summary, these findings will increase our understanding of the complex molecular mechanisms of HF development and cycling, and provide a basis for the further study of the genes and functions of HF development.

Candidate genes for the development of hair follicles in Hu sheep.

The aim of this study was to detect candidate genes for the development of hair follicles in the Hu sheep breed. Seven genes have been detected in large, medium, and small wave follicles of Hu sheep using gene chip technology. The histological features of the follicles of newborn Hu-lambs were combined with fluorescence quantitative PCR technology to detect the correlation between the expression of the seven genes and hair follicle development. Among the genes studied, matrix metalloproteinase 2 (MMP2), bone morphogenetic protein-7 (BMP7), and sideroflexin 1 (SFXN1) showed a significantly different pattern of expression in large, medium, and small wave follicles (P < 0.05). The expression of MMP2 had a significant positive correlation with secondary follicles in large waves (P < 0.05), while the expression of BMP7 had a significant correlation with primary follicle diameter in small wave follicles, and a highly significant positive correlation with the number of secondary follicles in the small waves (P < 0.01). The expression of SFXN1 was significantly and positively correlated with the diameters of small wave primary follicles; it also showed a highly significant positive correlation with secondary follicle diameters. Although other genes are associated with hair follicles, their expression in large, medium, and small wave follicles was not significant. We propose that BMP7, MMP2, and SFXN1 genes could be important candidate genes for use in breeding Hu lambs with early coat development.

Analysis of the spatiotemporal expression of major genes in the TGF-ß/Smad signaling pathway and correlation analysis using Hu sheep muscle tissue.

The mRNA expression levels of key genes (Smads, MSTN, and MyoG) in the TGF-ß/Smad signaling pathway in Hu sheep at different growth stages (2 days, 2 months, and 6 months of age) and in different skeletal muscles (longissimus dorsi muscle and soleus muscle) and different genders were detected; and correlation of the Smad family (Smad2, Smad3, Smad4, and Smad7), MSTN, MyoG expressions was analyzed in Hu sheep. The results showed that the expression of Smads was higher in the soleus muscle than in the longissimus dorsi muscle; the expressions of Smad2, Smad3, and Smad4 were significantly higher in 2-day-old sheep than in sheep belonging to the other age groups (P < 0.05); the expressions of Smad2, Smad4, and Smad7 were higher in rams than in 2-day-old ewes, but lower in rams than in 2-month-old and 6-month-old ewes; and the expression of Smad3 was higher in rams than in 2-day-old and 2-month-old ewes, but lower in rams than in 6-month-old ewes. In the 2 different muscle tissues, expression of Smad2 was significantly positively correlated (P < 0.01) with that of Smad3. The expression of Smad3 was significantly positively correlated (P < 0.01) with that of Smad4, which showed that the Smad family genes could have an inhibitory effect on the TGF-ß/Smad signaling pathway.

Sheep YAP1 temporal and spatial expression trend and its relation with MyHCs expression.

RT-PCR was used to study the temporal and spatial pattern of Yes-associated protein 1 (YAP1) and myosin heavy chain (MyHC) expression in four different skeletal muscles (i.e., longissimus dorsi muscle, soleus muscle, gastrocnemius muscle, and extensor digitorum longus) and three growth stages (i.e., 2 days old, 2 and 6 months old) of Hu Sheep. The results showed that YAP1 was differentially expressed in skeletal muscles of sheep, that expression increased gradually with age, and that there were high levels of expression in the gastrocnemius muscle and lower levels in the longissimus dorsi muscle. MyHCI was expressed at high levels in the soleus muscle and at lower levels in the longissimus dorsi muscle. In contrast, MyHCIIA and MyHCIIX were expressed at high levels in the extensor digitorum longus and at lower levels in the soleus muscle. The expression of MyHCI and MyHCIIA decreased with increasing age while that of MyHCIIX increased. YAP1 expression was negatively correlated with MyHCII (P < 0.01) and positively correlated with MyHCIIX (P < 0.01) across all growth stages and skeletal muscle types studied. We speculate that after birth, the thicker muscle fiber diameter is associated with the high expression of MyHCIIX. Therefore, we conclude that YAP1 expression affects sheep muscle fiber development after birth and provides important genetic information for the selection candidate genes for sheep muscle growth.

Cyclin D1 G870A polymorphism is associated with an increased risk of multiple myeloma.

Cyclin D1 is an important cell cycle regulator implicated in the pathogenesis of many cancer types. In particular, translocation and overexpression of cyclin D1 are common events in multiple myeloma (MM), suggesting that it may drive the initiation and progression of this malignancy. However, the association between genetic polymorphisms of cyclin D1 and the risk for developing MM remains poorly characterized. We performed a case-control study with 67 patients with MM and 66 healthy controls in a Chinese population. The cancer-associated G870A single nucleotide polymorphism (SNP) of CCND1, the gene encoding cyclin D1, was determined by polymerase chain reaction and restriction fragment length polymorphism. We found that there was a strong association between the homozygous variant allele  (GG) and MM susceptibility, with an odds ratio (OR) of 4.679 [95% confidence interval (CI): 1.081-5.647]. When further stratified analysis was performed, the increased risk was only evident in individuals over 60 years old (OR=3.297; 95%CI: 1.058-10.276). Our results suggest that the CCND1 G870A SNP may be critically involved in MM development.

Correlation between sheep YAP1 temporal and spatial expression trends and MSTN and MyoG gene expression.

The aim of the current study was to investigate the effects of Yes-associated protein 1 (YAP1) gene expression after birth on the development of muscle and the relationship between YAP1 and myostatin (MSTN) and myogenin (MyoG). Reverse transcription polymerase chain reaction was used to analyze the trends in YAP1, MSTN, and MyoG temporal and spatial expression levels in various skeletal muscles (i.e., longissimus dorsi muscle, soleus muscle, gastrocnemius muscle, and extensor digitorum longus) and across 3 different growth stages (i.e., 2 days old, 2 and 6 months old) of Hu Sheep. The results showed that YAP1 expression was significantly different in the skeletal muscles of sheep; the expression level gradually increased with age; it was highly expressed in the gastrocnemius muscle and minimally expressed in the longissimus dorsi muscle. MSTN, a negative regulator of skeletal muscle development, was minimally expressed in the soleus muscle and might be related to the enlargement of muscle fiber diameter. MyoG, an important factor in regulating skeletal muscle development, was minimally expressed in the longissimus dorsi muscle and extensor digitorum longus, and highly expressed in the gastrocnemius and soleus muscles; it might inhibit the enlargement of muscle fiber diameter after birth. YAP1 expression was significantly (P < 0.05) or extremely significantly (P < 0.01) and positively correlated with MSTN and MyoG at 2 days old, 2 and 6 months old. YAP1 expression was related to muscle fiber development after birth and might be a candidate gene for the regulation of muscle growth.

Expression of the RORα gene in Inner Mongolian cashmere goat hair follicles.

The expression of retinoid-acid-related orphan receptor α (RORα) was evaluated at the mRNA level using real-time polymerase chain reaction (qRT-PCR), and its expression localization was determined by in situ hybridization of adult Inner Mongolian cashmere goats at different times of the year. In situ hybridization demonstrated that RORαwas expressed in secondary hair follicles of the hair shaft, inner root sheath, outer root sheath, medulla, and other parts that are target organs of the RORαreceptor gene. qRT-PCR results showed that there was no significant difference in the RORa mRNA abundance in February, April, August, and October (P > 0.05), and the only difference occurred in December relative to February, August, and October (P < 0.05). This difference revealed that melatonin possibly promotes cashmere growth through the nuclear receptor RORα. This study provides a good foundation for future studies on the relationship between the melatonin receptor and cashmere growth; in addition, it provides new insights for increased cashmere production and quality.

Hair follicle transcriptome profiles during the transition from anagen to catagen in Cashmere goat (Capra hircus).

Previous molecular genetic studies of the goat hair life cycle have focused primarily on a limited number of genes and proteins. To identify additional genes that may play important roles in hair follicle cycle regulation, Illumina sequencing technology was used to catalog differential gene expression profiles in the hair growth cycle (anagen to catagen) of goat, comparing the primary hair follicle with the secondary hair follicle. There were 13,769 and 12,240 unigenes assembled from the reads obtained from primary hair follicle and secondary hair follicle, respectively. Genes encoding keratin proteins and keratin-associated proteins were the most highly expressed. A total of 5899 genes were differentially expressed in anagen vs catagen primary hair follicles, with 532 genes up-regulated and 5367 genes down-regulated. A total of 5208 genes were differentially expressed in anagen vs catagen secondary hair follicle, including 545 genes that were up-regulated and 4663 genes that were down-regulated. Numerous hair growth genes are expressed in the goat hair follicle, of which 73 genes showed co-up-regulation in both hair follicles during the anagen stage. Many of these up-regulated genes, such as STC2, VEGFR, and ROR2, are known to be transfactors in the process of cell differentiation and in the cell cycle. The differential gene expression profiles between primary hair follicles and secondary hair follicles obtained provide a foundation for future studies examining the network of gene expression controlling hair growth cycle in Cashmere goat.

Association between DLK1 and IGF-I gene expression and meat quality in sheep.

The aim of the present study was to detect delta-like 1 ho-molog (DLK1) and insulin-like growth factor-I (IGF-I) gene expression in the longissimus dorsi of Hu sheep at different growth stages and study the association between these genes and meat quality. The diameter and density of muscle fibers and tenderness of the longissimus dorsi were measured. Growth stage, but not sex, significantly affected DLK1 and IGF-I expression. DLK1 and IGF-I expression in the sheep longissimus dorsi gradually increased with growth, but also decreased during some periods. These results suggest that different growth stages significantly affect DLK1 and IGF-I gene expression in sheep muscle tissue. The ex-pression of DLK1 and IGF-I genes were positively and significantly (P < 0.01) correlated with muscle fiber diameter and muscle fiber shear stress, and negatively and significantly (P < 0.01) correlated with muscle fiber density. Muscle fiber diameter was positively and significantly (P < 0.01) correlated with muscle fiber shear stress, and negatively and significantly (P < 0.01) correlated with muscle fiber density. In addition, DLK-1 expression was significantly (P < 0.01) and positively correlated with IGF-I expression.

Developmental changes in IGF-I and MyoG gene expression and their association with meat traits in sheep.

In the present study, real time-polymerase chain reaction was applied to analyze the expression of IGF-I and MyoG genes in Hu sheep longissimus dorsi at different growth stages and their association with meat traits. Expression of the IGF-I gene in Hu sheep differed significantly between males and females at the two day-old (0.01 < P < 0.05), one-month old (0.01 < P < 0.05), and three month-old (P < 0.01) stages. IGF-I gene expression in male longissimus muscles was higher than that of females at all growth stages, except for the three month-old stage. There was no significant difference (P > 0.05) between males and females at any growth stage in expression of the MyoG gene. MyoG gene expression in male longissimus muscles tended to be higher than that of females at all growth stages, except for the six month-old stage. IGF-I gene expression was significantly and positively correlated with live weight (P < 0.01) and carcass weight (0.01< P < 0.05), and was non-significantly positively correlated with net meat weight (P > 0.05). In contrast, MyoG gene expression was non-significantly and positively correlated with live weight, carcass, and net meat weight (P > 0.05). Carcass traits showed highly significant positive correlations (P < 0.01). Furthermore, expressions of IGF-I and MyoG genes showed highly significant positive correlations (P < 0.01). We conclude that the expressions of IGF-I and MyoG genes are significantly and positively correlated with early muscle traits of Hu sheep.