Molecular Characterization of the Follistatin (FST) Gene in Quail and its Modulation through Natural Agonists

Solid tumor development is frequently accompanied by energy-deficient conditions such as glucose deprivation and hypoxia. Follistatin (FST), a secretory protein originally identified from ovarian follicular fluid, has been suggested to be involved in tumor development. However, whether it plays a role in cancer cell survival under energy-deprived conditions remains elusive. In this study, we demonstrated that glucose deprivation markedly enhanced the expression and nucleolar localization of FST in HeLa cells. The nucleolar localization of FST relied on its nuclear localization signal (NLS) comprising the residues 64-87. Localization of FST to the nucleolus attenuated rRNA synthesis, a key process for cellular energy homeostasis and cell survival. Overexpression of FST delayed glucose deprivation-induced apoptosis, whereas down-regulation of FST exerted the opposite effect. These functions depended on the presence of an intact NLS because the NLS-deleted mutant of FST lost the rRNA inhibition effect and the cell protective effect. Altogether, we identified a novel nucleolar function of FST, which is of importance in the modulation of cancer cell survival in response to glucose deprivation.

Follistatin (FST) is a secretory glycoprotein and belongs to the TGF-β superfamily. Previously, we found that two single nucleotide polymorphisms (SNPs) of sheep FST gene were significantly associated with wool quality traits in Chinese Merino sheep (Junken type), indicating that FST is involved in the regulation of hair follicle development and hair trait formation. The transcription regulation of human and mouse FST genes has been widely investigated, and many transcription factors have been identified to regulate FST gene. However, to date, the transcriptional regulation of sheep FST is largely unknown. In the present study, genome walking was used to close the genomic gap upstream of the sheep genomic FST gene and to obtain the FST gene promoter sequence. Transcription factor binding site analysis showed sheep FST promoter region contained a conserved putative binding site for signal transducer and activator of transcription 3 (STAT3), located at nucleotides −423 to −416 relative to the first nucleotide (A, +1) of the initiation codon (ATG) of sheep FST gene. The dual-luciferase reporter assay demonstrated that STAT3 inhibited the FST promoter activity and that the mutation of the putative STAT3 binding site attenuated the inhibitory effect of STAT3 on the FST promoter activity. Additionally, chromatin immunoprecipitation assay (ChIP) exhibited that STAT3 is directly bound to the FST promoter. Cell proliferation assay displayed that FST and STAT3 played opposite roles in cell proliferation. Overexpression of sheep FST significantly promoted the proliferation of sheep fetal fibroblasts (SFFs) and human keratinocyte (HaCaT) cells, and overexpression of sheep STAT3 displayed opposite results, which was accompanied by a significantly reduced expression of FST gene (P < 0.05). Taken together, STAT3 directly negatively regulates sheep FST gene and depresses cell proliferation. Our findings may contribute to understanding molecular mechanisms that underlie hair follicle development and morphogenesis.

Reproductive traits are affected by many factors, including ovarian function,hormones, and genetics. Genetic polymorphisms of candidate genes are associatedwith reproductive traits. Several candidate genes are associated with economictraits, including the follistatin (FST) gene. Thus, this studyaimed to evaluate whether the genetic variations in the FSTgene are associated with the reproductive traits in Awassi ewes. The genomic DNAwas extracted from 109 twin ewes and 123 single-progeny ewes. Therefore, 4sequence fragments from the FST gene were amplified usingpolymerase chain reaction (PCR) (exon 2/240, exon 3/268, exon 4/254, and exon5/266 bp, respectively). For a 254 bp amplicon, 3 genotypes were identified: CC,CG, and GG. Sequencing revealed a novel mutation in CG genotypes c.100C > G.The statistical analysis of c.100C > G showed an association withreproductive characteristics. Ewes carrying the c.100C > G had significantly(P ⩽ .01) lower litter sizes, twinning rates, lambingrates, and more days to lambing compared with CG and CC genotypes. Logisticregression analysis confirmed that the c.100C > G single-nucleotidepolymorphism (SNP) is responsible for decreasing litter size. According to theseresults, the variant c.100C > G negatively affects the traits of interest andis associated with lower reproductive traits in Awassi sheep. As a result ofthis study, ewes carrying the c.100C > G SNP have lower litter size and areless prolific.

Out of a panel of 37 candidate genes tested for linkage with polycystic ovary syndrome (PCOS), the strongest evidence of linkage was reported in the follistatin (FST) gene region. Subsequently, a couple of studies outside India investigated the FST gene for the presence of any mutations and its association with PCOS and the results were found to be largely inconsistent probably due to differences in the ethnic backgrounds and small sample sizes. To screen the FST gene for mutations and to establish their association pattern with PCOS among a large cohort of South Indian women. Case-control study. PCOS cases were recruited according to the 2003 Rotterdam diagnostic criteria. All the exons of the FST gene were amplified and analyzed in all the cases and controls for the presence of mutations using polymerase chain reaction (PCR) and direct DNA sequencing. A total of 549 women consisting of 250 PCOS cases and 299 controls were recruited for the study. No mutations were found in any of the exons of the FST gene in our Indian sample which is consistent with an earlier finding among the Asian women from Singapore. Although three of the four cohorts of Caucasian background studied earlier reported variants, none of them could establish a strong association with PCOS. The occurrence of the exonic variants of FST gene seems to be dependent on the ethnic background of the subjects under study and its role in the PCOS pathophysiology cannot be established with hitherto available evidence.

Follistatin (FST), a member of the transforming growth factor beta super-family regulates body growth by inhibiting the binding of myostatin (an inhibitor of growth) with its receptor in chicken. An experiment was conducted to explore ontogenic expression of the follistatin gene, determine polymorphism at the coding region of the gene and estimate its effect on growth traits in native (Aseel) and exotic broiler (PD-1) and layer (White Leghorn) chicken. The significant differences of FST gene expression were observed among the breeds revealing significantly (p < 0.05) higher expression in PD-1 line followed by White Leghorn and Aseel breeds during both embryonic and post-hatch period. The polymorphism at the functional domain of the FST gene was identified with the presence of 4 haplotypes. The follistatin haplogroups had the significant effect on body weights (p < 0.05) at 42 days of age in the White Leghorn, PD-1 and Aseel breeds (h1h1 in PD-1, h1h4 in White Leghorn and h1h2 haplogroups in Aseel breeds had the highest body weights of 770.04 ± 12.96, 246.28 ± 7.60 and 270.00 ± 10.68 g, respectively). It is concluded that the follistatin gene expressed differently during the embryonic and post-embryonic period across the breeds and the coding region of the gene was polymorphic having significant effects on growth traits in chicken.

Myocardial infarction causes mitochondrial atrophy and loss of function by reducing mitochondrial volume. Therefore, researchers are interested in finding a way to reduce the injuries and treat them. The study aims to evaluate the effect of 8 weeks of high-intensity interval training on follistatin (FST) gene expression in the fast and slow twitch muscles of rats with myocardial infarction. The study was conducted in 2020 at the Cardiac Research Center, Shahid Rajaei University of Medical Sciences (Tehran, Iran). For this purpose, 12 male Wistar rats with myocardial infarction were assigned to the experimental group high-intensity interval training (3 days a week for 30 min, each interval consisting of 4 min of running with 85-90% VO2max intensity and 2 min of active recovery with intensity of 50-60% VO2max for 8 weeks) and a control group. Then, the expression of follistatin in fast and slow twitch muscle contraction genes was investigated as triggers and inhibitors of muscle atrophy. Statistical data were analyzed with SPSS18 (α≥0.05). To determine the normality of the data, the Kolmogorov-Smirnov test was used, and in the case of normality of the data distribution, the independent samples t test was used. Independent t test results showed that FST gene expression in the slow twitch (ST) muscle contraction group was significantly decreased compared with the control group (P<0.001). Moreover, the expression of the FST gene in fast twitch muscles was significantly increased in the severe exercise group compared with the control group (P<0.001). Overall, 8 weeks of intense intermittent exercise decreased FST gene expression in slow and fast twitch muscles in rats with myocardial infarction.

ABSTRACTProliferation, differentiation, and estrogen secretion of granulosa cells are the key factors affecting the estrous after weaning in sows. The objective of this study was to evaluate the expression of Follistatin (FST) in the ovary of Xiushui Hang and Duroc sows at weaning and estrus, the effect of FSH on transcript abundance of FST gene in granulosa cells and the role of FST gene in the weaning to estrus using siRNAs targeted to FST gene. In the present study, expression of the FST mRNA was evaluated by real time PCR. The FST mRNA levels showed a reduction from weaning to the estrus in both Xiushui Hang and Duroc sows, and the mRNA levels in Duroc ovary was higher than in Xiushui Hang sows at the beginning of estrus. Granulosa cells were obtained from the two largest follicles around follicular deviation, FST expression was decreased sharply after treatment with FSH (250 ng/ml). Knockdown of FST by siRNA in porcine granulosa cells significantly increased cell proliferation and estrogen secretion. These results indicate that FST gene is a negative regulator of follicle growth and function during the weaning-estrus interval.

Follistatin (FST) is involved in hair follicle morphogenesis. However, its effects on hair traits are not clear. This study was designed to investigate the effects of FST gene single nucleotide polymorphisms (SNP) on wool quality traits in Chinese Merino sheep (Junken Type). We performed gene expression analysis, SNP detection, and association analysis of FST gene with sheep wool quality traits. The real-time RT-PCR analysis showed that FST gene was differentially expressed in adult skin between Chinese Merino sheep (Junken Type) and Suffolk sheep. Immunostaining showed that FST was localized in inner root sheath (IRS) and matrix of hair follicle (HF) in both SF and Suffolk sheep. Sequencing analysis identified a total of seven SNPs (termed SNPs 1–7) in the FST gene in Chinese Merino sheep (Junken Type). Association analysis showed that SNP2 (Chr 16. 25,633,662 G>A) was significantly associated with average wool fiber diameter, wool fineness SD, and wool crimp (P < 0.05). SNP4 (Chr 16. 25,633,569 C>T) was significantly associated with wool fineness SD and CV of fiber diameter (P < 0.05). Similarly, the haplotypes derived from these seven identified SNPs were also significantly associated with average wool fiber diameter, wool fineness SD, CV of fiber diameter, and wool crimp (P < 0.05). Our results suggest that FST influences wool quality traits and its SNPs 2 and 4 might be useful markers for marker-assisted selection and sheep breeding.

Follistatin (FST) is an activin-binding protein that neutralizes the activity of activin. FST also binds other members of the transforming growth factor-beta (TGF-beta) superfamily, including myostatin (MSTN). We report herein on the isolation and characterization of a full-length cDNA sequence predicted to encode FST in a marine fish, the gilthead sea bream Sparus aurata. The deduced amino acid sequence of sea bream FST (saFST) is highly conserved to the counterpart sequences in other vertebrates and contains the N-terminal domain and three FST domains. The deduced mature saFST shows 81-86% identity with FSTs from other vertebrates. It is 290 amino acids long, similar to other fish FSTs and the short isoform of Xenopus FST but longer by two residues than mammalian FST288. Ontogeny of MSTN (a TGF-beta superfamily member and a negative growth regulator of skeletal muscle in mammals), and FST (known to bind MSTN) gene expression revealed the presence of both transcripts throughout larval development. However, a different expression pattern was found in earlier developmental stages; while MSTN could not be detected prior to the day of hatching, FST transcript was detected in embryos 12 h post-fertilization, confirming its role during vertebrate embryonic development. Both FST and MSTN were expressed in many adult tissues, with variable levels of expression, including muscle. Recombinant saFST inhibited saMSTN activity in a reporter gene assay, indicating a similar effect to that reported in mammals.

Barnase thermal titration via molecular dynamics simulations: Detection of early denaturation sites

Follistatin (FST) inhibits the action of activin by interfering with the binding of activin to its receptor. Although the prognostic value of FST in various cancers has been investigated previously, studies rarely focused on hypopharyngeal carcinoma (HPC). In our study, the effect of FST expression on HPC tissues and cell lines was investigated. A total of 60 patients with HPC were recruited for this study. Levels of FST mRNA and protein were measured by quantitative polymerase chain reaction (PCR) and immunohistochemistry in HPC tissue samples and by qPCR in the HPC FaDu cells, as well as immortal nasopharyngeal epithelial cell line NP-69 cells. After silencing the FST expression in FaDu cells using lentivirus-mediated siRNA that was specific for FST mRNA, cell proliferation was determined by a Celigo assay. Tumor growth was monitored in nude mice and viability was determined by a methylthiazoletetrazolium assay. The ratio of cell cycle arrest and apoptosis was evaluated by flow cytometry. The colony formation ability was performed using Giemsa staining. In addition, wound healing and Transwell migration and invasion assays were performed for the analysis of cell motility. FST expression was significantly higher in human HPC tissue and FaDu cells than in normal tissue and NP-69 cells. A higher expression of FST in HPC samples was positively correlated with advanced tumors. Moreover, FST knockdown by shRNA significantly decreased cell growth, colony formation, migration and invasion. Furthermore, FST silencing increased the cell apoptosis percentage, and arrested cell cycle in the S phase in FaDu cells. In addition, FST silencing suppressed tumor growth in vivo. Our results indicated that the FST gene was associated with HPC progression and may serve as a potential therapeutic target for the treatment of HPC.

Background: Using natural ingredients as antivirals can be considered a treatment for SARS-CoV-2. One of the potential plants, mahogany (Swietenia macrophylla King), is widely used in various countries as an antiviral treatment. Paparin-like protease (PLpro) is an essential cysteine ​​protease that regulates viral replication and interferes with the regulation of immune sensing. Objective: This study aims to predict which compounds in the mahogany plant have good affinity, patterns, and stability interaction against the target protein of SARS-CoV-2. Methods: The drug-likeness parameter using SwissADME was used to screen compounds that will be docked against PLpro using the Autodock program. The parameters observed in molecular docking analysis are the value of bond energy and interaction model to amino acid residues. The compounds in mahogany plants that have the best interactions were then analyzed using molecular dynamics simulation methods to determine the stability of their bonds based on the values of Root Mean Square Deviation (RMSD) and Root Mean Square Fluctuation (RMSF). Results: Twenty-two compounds met the drug-likeness requirements. Molecular docking analysis showed that the compounds predicted to have the best binding affinity and have an interaction pattern similar to natural ligands towards the molecular target of PLpro are 7-deacetoxy-7-oxogedunin and 3β-hydroxy-stigmast-5-en-7-one. The molecular dynamics simulation results revealed that based on the RMSD and RMSF values, the compound 3β-hydroxy-stigmast-5-en-7-one showed higher stability than 7-deacetoxy-7-oxogedunin. Conclusion: 3β-hydroxy-stigmast-5-en-7-one and 7-deacetoxy-7-oxogedunin were predicted to have good interaction with PLPro; however, 3β-hydroxy-stigmast-5-en-7-one showed the higher interaction stability.

Objective Recent animal studies have suggested that loss of follistatin (FST) may result in premature cessation of ovarian function. Our objective was to investigate whether mutations in the FST coding region are present in Chinese women with idiopathic premature ovarian failure (POF).Method The matched case-control study took place in the First Affiliated Hospital of Nanjing Medical University with 80 idiopathic POF patients and 80 matched controls. There were no interventions. The entire FST coding region was analyzed by direct sequencing in all subjects.Results Three FST gene variants were identified, namely c.270C> G, c.598G> C and c.953–184A> T (rs722910). The synonymous variant (c.270C> G) in exon 2 was present in the heterozygous state in a single POF patient. The novel c.598G> C missense mutation, located in exon 4 and resulting in an alanine to proline substitution at amino acid 200, was detected in a single healthy control. There was no difference in genotype distribution and allele frequency of the known single nucleotide polymorphism rs722910 between POF patients and controls.Conclusion Although we did not find any evidence that it is a disease-causing gene, our study is the first to evaluate the role of the FST gene in Chinese women with idiopathic POF.

Molecular cloning, expression pattern of follistatin gene and association analysis with growth traits in bighead carp (Hypophthalmichthys nobilis).

Porcine follistatin gene structure supports two forms of mature follistatin produced by alternative splicing