Long noncoding RNA BMPR1B-AS1 stability regulated by IGF2BP2 affects the decidualization in endometriosis patients through the SMAD1/5/9 pathway.

Endometriosis (EMs)-related infertility commonly has decreased endometrial receptivity and normal decidualization is the basis for establishing and maintaining endometrial receptivity. However, the potential molecular regulatory mechanisms of impaired endometrial decidualization in patients with EMs have not been fully clarified. We confirmed the existence of reduced endometrial receptivity in patients with EMs by scanning electron microscopy and quantitative real-time PCR. Here we identified an lncRNA, named BMPR1B-AS1, which is significantly downregulated in eutopic endometrium in EMs patients and plays an essential role in decidual formation. Furthermore, RNA pull-down, mass spectrometry, RNA immunoprecipitation, and rescue analyses revealed that BMPR1B-AS1 positively regulates decidual formation through interaction with the RNA-binding protein insulin-like growth factor 2 mRNA-binding protein 2 (IGF2BP2). Downregulation of IGF2BP2 led to a decreased stability of BMPR1B-AS1 and inhibition of activation of the SMAD1/5/9 pathway, an inhibitory effect which diminished decidualization in human endometrial stromal cells (hESCs) decidualization. In conclusion, our identified a novel regulatory mechanism in which the IGF2BP2-BMPR1B-AS1-SMAD1/5/9 axis plays a key role in the regulation of decidualization, providing insights into the potential link between abnormal decidualization and infertility in patients with EMs, which will be of clinical significance for the management and treatment of infertility in patients with EMs.

Association of polymorphism in the promotor area of the caprine BMPR1B gene with litter size and body measurement traits in Damani goats.

This study aimed to explore polymorphisms in the promoter region of the caprine BMPR1B (Bone morphogenetic protein receptor 1 beta) gene and its association with body measurement and litter size traits in Damani does. A total of 53 blood samples were collected to analyze the association between the BMPR1B gene polymorphism and 11 phenotypic traits in Damani female goats. The results revealed that three novel SNPs were identified in the promoter region of the caprine BMPR1B gene, including g.67 A > C (SNP1), g.170 G > A(SNP2), and g.501A > T (SNP3), among which the SNP1 and SNP2 were significantly (p < 0.05) associated with litter size and body measurement traits in Damani goats. In SNP1 the AC genotype could be used as a marker for litter size, and the CC genotype for body weight in Damani goats. In SNP2, the genotype GG was significantly (p < 0.05) associated with ear and head length. Therefore, we can conclude from the present study, that genetic variants AC and CC of the caprine BMPR1B gene could be used as genetic markers for economic traits through marker-assisted selection for the breed improvement program of the Damani goat.

Survey of the relationship between polymorphisms within the BMPR1B gene and sheep reproductive traits.

The BMPRIB gene is one of the main genes that can be used as a molecular genetic marker for the early selection of highly productive ewes. It is well-documented that the p.Q249R (g.746A > G) is the first mutation in the kinase domain of the BMPR1B gene that is highly related to increased ovulation rate and litter size. It is likely that the presence of the p.Q249R mutation in the sheep population is one of the factors contributing to the outstanding productivity of the sheep. Moreover, in recent years, researchers have been explored other polymorphisms in the BMPR1B gene with respect to reproductive traits in sheep. Therefore, we carried out the current study to evaluate the association between polymorphisms in this gene and sheep litter size from all appropriate studies. As a result, among 41 polymorphisms in the ovine BMPRIB gene, eight variants, including p.Q249R (g.746A > G), g.29362047T > C, g.29427689G > A, BMPR1B-2 (ss:1960972599), g.29382337G > A, g.29382340G > A, rs1092293287 (10 bp insertion/deletion) and g.29380965A > G were found to be associated with litter size in sheep. This systematic analysis presents the most current data evidence for BMPRIB polymorphisms, highlighting the need for further large-scale studies to determine more important variants.

FecB mutation and litter size are associated with a 90-base pair deletion in BMPR1B in East Friesian and Hu crossbred sheep.

Litter size is a critical economic trait in livestock, but only a few studies have focused on associated indel mutations in BMPR1B, a key regulator of ovulation and litter size in sheep. We evaluated the effects of BMPR1B mutations on the reproductive performance of sheep. We used Hu, East Friesian, and East Friesian/Hu crossbred sheep as experimental subjects and identified a novel 90 bp deletion in BMPR1B, which coincides with the c.746A > G (FecB mutation) genotype. The correlation between the two loci and litter size was then evaluated. We identified three genotypes for the Del-90bp locus, namely, II, ID, and DD, and three genotypes for the c.746A > G locus, namely ++, B+, and BB. Both Del-90bp and c.746A > G significantly affected the litter size of Hu and East Friesian/Hu crossbred sheep. Linkage disequilibrium analysis revealed a strong linkage disequilibrium between these loci in Hu sheep and the F1 population (r2 > 0.33), which suggests that detecting this 90 bp deletion might be a simple method to identify the likely carriers of c.746A > G. However, the function of this 90-bp deletion still needs further exploration. We provide genetic data that can be used as a reference for the breeding of improved prolific traits in sheep.

PCTAIRE Protein Kinase 1 (PCTK1) Suppresses Proliferation, Stemness, and Chemoresistance in Colorectal Cancer through the BMPR1B-Smad1/5/8 Signaling Pathway.

Colorectal cancer (CRC) is the third most common cancer and a leading cause of cancer-related mortality worldwide. Even with advances in therapy, CRC mortality remains high. Therefore, there is an urgent need to develop effective therapeutics for CRC. PCTAIRE protein kinase 1 (PCTK1) is an atypical member of the cyclin-dependent kinase (CDK) family, and the function of PCTK1 in CRC is poorly understood. In this study, we found that patients with elevated PCTK1 levels had a better overall survival rate in CRC based on the TCGA dataset. Functional analysis also showed that PCTK1 suppressed cancer stemness and cell proliferation by using PCTK1 knockdown (PCTK1-KD) or knockout (PCTK1-KO) and PCTK1 overexpression (PCTK1-over) CRC cell lines. Furthermore, overexpression of PCTK1 decreased xenograft tumor growth and knockout of PCTK1 significantly increased in vivo tumor growth. Moreover, knockout of PCTK1 was observed to increase the resistance of CRC cells to both irinotecan (CPT-11) alone and in combination with 5-fluorouracil (5-FU). Additionally, the fold change of the anti-apoptotic molecules (Bcl-2 and Bcl-xL) and the proapoptotic molecules (Bax, c-PARP, p53, and c-caspase3) was reflected in the chemoresistance of PCTK1-KO CRC cells. PCTK1 signaling in the regulation of cancer progression and chemoresponse was analyzed using RNA sequencing and gene set enrichment analysis (GSEA). Furthermore, PCTK1 and Bone Morphogenetic Protein Receptor Type 1B (BMPR1B) in CRC tumors were negatively correlated in CRC patients from the Timer2.0 and cBioPortal database. We also found that BMPR1B was negatively correlated with PCTK1 in CRC cells, and BMPR1B expression was upregulated in PCTK1-KO cells and xenograft tumor tissues. Finally, BMPR1B-KD partially reversed cell proliferation, cancer stemness, and chemoresistance in PCTK1-KO cells. Moreover, the nuclear translocation of Smad1/5/8, a downstream molecule of BMPR1B, was increased in PCTK1-KO cells. Pharmacological inhibition of Smad1/5/8 also suppressed the malignant progression of CRC. Taken together, our results indicated that PCTK1 suppresses proliferation and cancer stemness and increases the chemoresponse of CRC through the BMPR1B-Smad1/5/8 signaling pathway.

Progress on the effect of FecB mutation on BMPR1B activity and BMP/SMAD pathway in sheep.

BMPR1B is the first major gene of litter size identified in sheep. However, the molecular mechanism of the FecB mutation that increases the ovulation rate in sheep is still unclear. In recent years, it has been demonstrated that BMPR1B activity is regulated by the small molecule repressor protein FKBP1A, which acts as a key activity switch of the BMPR1B in the BMP/SMAD pathway. The FecB mutation is located close to the binding site of FKBP1A and BMPR1B. In this review, we summarize the structure of BMPR1B and FKBP1A proteins, and clarify the spatial interactive domains of the two proteins with respect to the location of the FecB mutation. Then the relationship between the FecB mutation and the degree of affinity of the two proteins are predicted. Finally, the hypothesis that FecB mutation causes change of activity in BMP/SMAD pathway by affecting the intensity of the interactions between BMPR1B and FKBP1A is proposed. This hypothesis provides a new clue to investigate the molecular mechanism of FecB mutation affecting ovulation rate and litter size in sheep.

A unique 15-bp InDel in the first intron of BMPR1B regulates its expression in Taihu pigs.

BACKGROUND: BMPR1B (Bone morphogenetic protein receptor type-1B) is a receptor in the bone morphogenetic protein (BMP) family and has been identified as a candidate gene for reproductive traits in pigs. Our previous study in Taihu pigs found a specific estrogen response element (ERE) in the first intron of the BMPR1B gene that is associated with the number born alive trait. However, little is known about the mechanism by which the ERE regulates the expression of BMPR1B in the endometrium. RESULTS: Here, a 15-bp InDel (insertion/deletion) (AGCCAGAAAGGAGGA) was identified as a unique variation in Taihu pigs, and was shown to be responsible for the binding of the type I receptor of estrogen (ESR1) to the ERE using dual-luciferase assays. Four BMPR1B transcripts (T1, T2, T3, and T4) were identified by 5' RACE in endometrial tissue. Expression of T3 and T4 in the endometrium of Meishan pigs was significantly higher than in Duroc pigs during pregnancy. Luciferase assays showed that three distinct BMPR1B promoters may drive expression of T1, T3, and T4. Interestingly, ERE-mediated enhancement of T4 promoter activity significantly increased expression of Transcript T4 in the endometrium of Taihu pigs (P < 0.05). In contrast, the ERE inhibited activity of the T3 promoter and decreased expression of the T3 transcript in the Duroc background (P < 0.05). In summary, we identified a 15-bp InDel in the Taihu ERE that can be used as a molecular marker for the number born alive trait, characterized the 5' untranslated regions (UTRs) of BMPR1B transcripts in the endometrium, and determined how the transcripts are processed by alternative splicing events. CONCLUSIONS: Our results provide a foundation for understanding the transcriptional regulation of BMPR1B and its contributions to the unique breeding prolificacy characteristics of Taihu pigs.

METTL3-mediated LINC00657 promotes osteogenic differentiation of mesenchymal stem cells via miR-144-3p/BMPR1B axis.

N6-methyladenosine (m6A) modification plays a crucial role in the progression of osteoporosis (OP). The study aimed to explore the effects of methyltransferase-like 3 (METTL3) in OP. The levels of METTL3, LINC00657, miR-144-3p and BMPR1B were detected using qPCR. Osteogenesis was assessed using alizarin red and alkaline phosphatase (ALP) staining assays. The protein expression of Bglap, Runx2 and Col1a1 was measured by western blot. The targets of LINC00657 and miR-144-3p were screened by bioinformatic analysis. The interaction between miR-144-3p and LINC00657 or BMPR1B was analyzed by dual-luciferase reporter assay and RNA pull-down assay. The results showed that METTL3 was downregulated in OP. METTL3 mediated m6A methylation of LINC00657 to promote the development of osteogenesis. Further study indicated that LINC00657 functioned as a ceRNA to upregulate BMPR1B via sponging miR-144-3p. Additionally, BMPR1B knockdown alleviated the effects of METTL3 on osteogenesis of bone marrow mesenchymal stem cells (BMSCs). Taken together, METTL3 facilitated osteogenic differentiation of BMSCs via the LINC00657/miR-144-3p/BMPR1B axis. Our findings may provide a novel insight of m6A methylation in the development of OP.

Endothelin-1 alters BMP signaling to promote proliferation of pulmonary artery smooth muscle cells.

Pulmonary arterial hypertension (PAH) is characterized by abnormal outgrowth of pulmonary artery smooth muscle cells (PASMCs) of the media. Abundant expression of endothelin-1 (ET-1) and activated p38 mitogen-activated protein kinase (p38MAPK) has been observed in PAH patients. p38MAPK has been implicated in cell proliferation. An unspecified disturbance in bone morphogenetic protein (BMP) signaling may be involved in the development of PAH. Type I receptors (BMPR1A and BMPR1B) and type II receptor (BMPR2) transduce signals via two distinct pathways, i.e., canonical and non-canonical pathways, activating Smad1/5/8 and p38MAPK, respectively. BMPR1B expression was previously reported to be enhanced in the PASMCs of patients with idiopathic PAH. BMP15 binds specifically to BMPR1B. We assessed the effects of ET-1 on BMP receptor expression and cell proliferation. BMP2 increased BMPR1B expression in human PASMCs after pretreatment with ET-1 in vitro. Although BMP2 alone did not affect PASMC proliferation, BMP2 treatment after ET-1 pretreatment significantly accelerated PASMC proliferation. PH-797804, a selective p38MAPK inhibitor, abrogated this proliferation. Similarly, after ET-1 pretreatment, BMP15 significantly accelerated the proliferation of PASMCs, whereas stimulation with BMP15 alone did not. In conclusion, in PASMCs, ET-1 exposure under pathological conditions alters BMP signaling to activate p38MAPK, resulting in cell proliferation.

Expression of steroidogenic enzymes and TGFß superfamily members in follicular cells of prepubertal gilts with distinct endocrine profiles.

Regulation of the transforming growth factor beta (TGFß) superfamily by gonadotrophins in swine follicular cells is not fully understood. This study evaluated the expression of steroidogenic enzymes and members of the TGFß superfamily in prepubertal gilts allocated to three treatments: 1200 IU eCG at D -3 (eCG); 1200 IU eCG at D -6 plus 500 IU hCG at D -3 (eCG + hCG); and the control, composed of untreated gilts. Blood samples and ovaries were collected at slaughter (D0) and follicular cells were recovered thereafter. Relative gene expression was determined by real-time PCR. Serum progesterone levels were greater in the eCG + hCG group compared with the other groups (P < 0.01). No differences were observed in the expression of BMP15, BMPR1A, BMPR2, FSHR, GDF9, LHCGR and TGFBR1 (P > 0.05). Gilts from the eCG group presented numerically greater mean expression of CYP11A1 mRNA than in the control group that approached statistical significance (P = 0.08) and greater expression of CYP19A1 than in both the eCG and the control groups (P < 0.05). Expression of BMPR1B was lower in the eCG + hCG treatment group compared with the control (P < 0.05). In conclusion, eCG treatment increased the relative expression of steroidogenic enzymes, whereas treatment with eCG + hCG increased serum progesterone levels. Although most of the evaluated TGFß members were not regulated after gonadotrophin treatment, the downregulation of BMPR1B observed after treatment with eCG + hCG and suggests a role in luteinization regulation.

Combined approaches identify known and novel genes associated with sheep litter size and non-seasonal breeding.

Improvement of ewe reproduction is considerable by appropriately increasing litter size and sustaining non-seasonal breeding. However, their genetic makeups have not been entirely elucidated. Genome-wide analyses of 821 individuals were performed by combining three genomic approaches (genome-wide association study, XP-nSL, and runs of homozygosity). Consequently, 35 candidate genes including three domestication genes (TSHR, GTF2A1, and KITLG) were identified. Other than the FecB mutation at BMPR1B, we described a significant association of a missense mutation rs406686139 at seasonal lambing-associated TSHR gene with litter size. Some promising novel genes may be relevant for sheep reproduction by multitude biological processes, such as FETUB functioning in fertilization, HNRNPA1 in oogenesis, DCUN1D1 in spermatogenesis, and HRG in fertility outcome. The present study suggests that improvement of ewe reproduction is attributed to selective breeding, and casts light on the genetic basis and improvement of sheep reproduction.

Association between novel variants in BMPR1B gene and litter size in Mongolia and Ujimqin sheep breeds.

Prolificacy is an important trait of animals, specifically for sheep. The Bone morphogenetic protein receptor 1B (BMPR1B) is a major gene affecting the litter size of many sheep breeds. The well-known FecB mutation (Q249R) was associated fully with the hyper prolific phenotype of Booroola Merino. However, the identification of variation in all exonic regions of BMPR1B was rare. In this study, we sequenced all exonic regions of BMPR1B gene of Mongolia sheep breed, and ten novel variants were detected by direct sequencing. Among them, the litter size of the Mongolia ewes with the CC genotype was significantly higher (0.34 additional lambs, p < .05) than those with the TT genotype of the g.29346567C>T single nucleotide polymorphism (SNP). The litter size of the Mongolia ewes with the TT genotype was significantly higher (0.19 additional lambs, p < .05 and .31 additional lambs, p < .01, respectively) than those with the GT and GG genotypes of the c.1470G>T SNP. The silent c.1470G>T mutation is predicted to increase the stability of the mRNA secondary structure through reducing minimum free energy and is predicted to change the mRNA secondary structure of BMPR1B. Our findings may give potentially useful genetic markers for increasing litter size in sheep.

Association of a genetic polymorphism in the BMPR-1B gene, and non-genetic factors with the natural prolificacy of the Colombian-haired sheep.

INTRODUCTION: Colombian-haired sheep (OPC) is a creole breed with very good adaptation to the tropical conditions of our country. In sheep, it has been shown that the litter size (LS) is associated with ovulation rate, the number of fertilized eggs, and embryo survival. Also, LS is determined by genetic and environmental effects. In this sense, the receptor 1B of bone morphogenetic protein (BMPR-1B) has been described as a genetic factor. Therefore, the aim of the present work was to characterize and associate the SNP C864T in the BMPR-1B gene with LS in the specific OPC biotypes Ethiopian and Sudan. MATERIALS AND METHODS: Reproductive history (LS, number of calving in the mother, identification of the father, conception year, and conception period) of 200 OPC sheep was assessed. Additionally, sheep were genotyped by sequencing for the SNP C864T. An association between LS, reproductive history, and C864T variation was performed using a GLM fixed-effect model. RESULTS: The frequency of the T allele (0.75 ± 0.03) was higher than that of the C allele (P<0.05). The genotypic frequencies were 0.55 ± 0.06, 0.38 ± 0.04, and 0.07 ± 0.01, for TT, TC, and CC, respectively. An average value of He (0.37 ± 0.03) and HWE (P=0.97) was found. The LS found was 1.45 ± 0.15. This varied, between biotypes, with number of calving in the mother, with the father, and at the time of conception (P <0.05). CONCLUSION: The LS varied between genotypes (P<0.05). The CC genotype was the most prolific (1.81 ± 0.4), followed by the heterozygous (1.45 ± 0.04) and the TT homozygous (1.09 ± 0.04). However, we did not find a variation between biotypes within the genotypes (P>0.05). In conclusion, the polymorphism target in the exon 9 of the BMPR-1B gene and non-genetic factors affected significantly the litter size in the OPC.

Eukaryotic expression, Co-IP and MS identify BMPR-1B protein-protein interaction network.

BACKGROUND: BMPR-1B is part of the transforming growth factor ß super family and plays a pivotal role in ewe litter size. Functional loss of exon-8 mutations in the BMPR-1B gene (namely the FecB gene) can increase both the ewe ovulation rate and litter size. RESULTS: This study constructed a eukaryotic expression system, prepared a monoclonal antibody, and characterized BMPR-1B/FecB protein-protein interactions (PPIs). Using Co-immunoprecipitation coupled to mass spectrometry (Co-IP/MS), 23 proteins were identified that specifically interact with FecB in ovary extracts of ewes. Bioinformatics analysis of selected PPIs demonstrated that FecB associated with several other BMPs, primarily via signal transduction in the ovary. FecB and its associated interaction proteins enriched the reproduction process via BMP2 and BMP4 pathways. Signal transduction was identified via Smads proteins and TGF-beta signaling pathway by analyzing the biological processes and pathways. Moreover, other target proteins (GDF5, GDF9, RhoD, and HSP 10) that interact with FecB and that are related to ovulation and litter size in ewes were identified. CONCLUSIONS: In summary, this research identified a novel pathway and insight to explore the PPi network of BMPR-1B.

Smad4 Feedback Enhances BMPR1B Transcription in Ovine Granulosa Cells.

BMPR1B is a type 1B receptor of the canonical bone morphogenetic protein (BMP)/Sma- and mad-related protein (Smad) signaling pathway and is well known as the first major gene associated with sheep prolificacy. However, little is known about the transcriptional regulation of the ovine BMPR1B gene. In this study, we identified the ovine BMPR1B gene promoter and demonstrated that its transcription was regulated by Smad4. In sheep ovarian follicles, three transcriptional variants of BMPR1B gene with distinct transcription start sites were identified using 5' RACE assay while variants II and III were more strongly expressed. Luciferase assay showed that the region -405 to -200 nt is the PII promoter region of variant II. Interestingly, two putative Smad4-binding elements (SBEs) were detected in this region. Luciferase and ChIP assay revealed that Smad4 enhances PII promoter activity of the ovine BMPR1B gene by directly interacting with SBE1 motif. Furthermore, in the ovine granulosa cells, Smad4 regulated BMPRIB expression, and BMPRIB-mediated granulosa cells apoptosis. Overall, our findings not only characterized the 5' regulatory region of the ovine BMPR1B gene, but also uncovered a feedback regulatory mechanism of the canonical BMP/Smad signaling pathway and provided an insight into the transcriptional regulation of BMPR1B gene and sheep prolificacy.

Association of polymorphisms in bone morphogenetic protein receptor-1B gene exon-9 with litter size in Dorset, Mongolian, and Small Tail Han ewes.

OBJECTIVE: The present study was to investigate the association of polymorphisms in exon-9 of the bone morphogenetic protein receptor-1B (BMPR-1B) gene (C864T) with litter size in 240 Dorset, 232 Mongolian, and 124 Small Tail Han ewes. METHODS: Blood samples were collected from 596 ewes and genomic DNA was extracted using the phenol: chloroform extraction method. The 304-bp amplified polymerase chain reaction product was analyzed for polymorphism by single-strand conformation polymorphism method. The genotypic frequency and allele frequency of BMPR-1B gene exon-9 were computed after sequence alignment. The χ2 independence test was used to analyze the association of genotypic frequency and litter size traits with in each ewe breed, where the phenotype was directly treated as category. RESULTS: The results indicated two different banding patterns AA and AB for this fragment, with the most frequent genotype and allele of AA and A. Calculated Chi-square test for BMPR-1B gene exon-9 was found to be more than that of p value at the 5% level of significance, indicating that the population under study was in Hardy-Weinberg equilibrium for all ewes. The χ2 independence test analyses indicated litter size differences between genotypes was not the same for each breed. The 304-bp nucleotide sequence was subjected to BLAST analysis, and the C864T mutation significantly affected litter size in singletons, twins and multiples. The heterozygosity in exon-9 of BMPR-1B gene could increase litter size for all the studied ewes. CONCLUSION: Consequently, it appears that the polymorphism BMPR-1B gene exon-9 detected in this study may have potential use in marker assisted selection for litter size in Dorset, Mongolian, and Small Tail Han ewes.

BmpR1A is a major type 1 BMP receptor for BMP-Smad signaling during skull development.

Craniosynostosis is caused by premature fusion of one or more sutures in an infant skull, resulting in abnormal facial features. The molecular and cellular mechanisms by which genetic mutations cause craniosynostosis are incompletely characterized, and many of the causative genes for diverse types of syndromic craniosynostosis have not yet been identified. We previously demonstrated that augmentation of BMP signaling mediated by a constitutively active BMP type IA receptor (ca-BmpR1A) in neural crest cells (ca1A hereafter) causes craniosynostosis and superimposition of heterozygous null mutation of Bmpr1a rescues premature suture fusion (ca1A;1aH hereafter). In this study, we superimposed heterozygous null mutations of the other two BMP type I receptors, Bmpr1b and Acvr1 (ca1A;1bH and ca1A;AcH respectively hereafter) to further dissect involvement of BMP-Smad signaling. Unlike caA1;1aH, ca1A;1bH and ca1A;AcH did not restore the craniosynostosis phenotypes. In our in vivo study, Smad-dependent BMP signaling was decreased to normal levels in mut;1aH mice. However, BMP receptor-regulated Smads (R-Smads; pSmad1/5/9 hereafter) levels were comparable between ca1A, ca1A;1bH and ca1A;AcH mice, and elevated compared to control mice. Bmpr1a, Bmpr1b and Acvr1 null cells were used to examine potential mechanisms underlying the differences in ability of heterozygosity for Bmpr1a vs. Bmpr1b or Acvr1 to rescue the mut phenotype. pSmad1/5/9 level was undetectable in Bmpr1a homozygous null cells while pSmad1/5/9 levels did not decrease in Bmpr1b or Acvr1 homozygous null cells. Taken together, our study indicates that different levels of expression and subsequent activation of Smad signaling differentially contribute each BMP type I receptor to BMP-Smad signaling and craniofacial development. These results also suggest differential involvement of each type 1 receptor in pathogenesis of syndromic craniosynostoses.

A novel homozygous variant in BMPR1B underlies acromesomelic dysplasia Hunter-Thompson type.

Acromesomelic dysplasia is genetically heterogeneous group of skeletal disorders characterized by short stature and acromelia and mesomelia of limbs. Acromesomelic dysplasia segregates in an autosomal recessive pattern and is caused by biallelic sequence variants in three genes (NPR2, GDF5, and BMPR1B). A consanguineous family of Pakistani origin segregating a subtype of acromesomelic dysplasia called Hunter-Thompson was clinically and genetically evaluated. Genotyping of microsatellite markers and linkage analysis revealed a 7.78 Mb homozygous region on chromosome 4q22.3, which harbors BMPR1B. Sequence analysis of the gene revealed a novel homozygous missense variant (c.1190T > G, p.Met397Arg) that segregates with the disease phenotype within the family and produced a Logarithm of odds (LOD) score of 3.9 with the disease phenotype. This study reports on the first familial case of acromesomelic dysplasia Hunter-Thompson type. It is also the first report of BMPR1B underlying the etiology of acromesomelic dysplasia Hunter-Thompson type.

Polymorphism of GDF9 and BMPR1B genes and their association with litter size in Markhoz goats.

The main objective of this study was to investigate the polymorphism of GDF9 and BMPR1B genes and their relationship with litter size in Markhoz goats. The polymorphism of GDF9 and BMPR1B genes as well-documented genes regarding fecundity in sheep and goat was investigated using RFLP-PCR and a tetra-primer amplification refractory mutation system-PCR (T-ARMS-PCR) in Markhoz goats. The 164 blood samples were collected from the raised goats in Sanandaj Markhoz goat Performance Testing Station. The DNA extraction was carried out by salting-out procedure, and then, PCR was performed using four and two pairs of primers to detect polymorphism in GDF9 and BMPR1B genes, respectively. To disclose GDF9 loci polymorphism, PCR products were digested with SspI (G3288A), PvuII (G423A), MvaI (A959C) and MspI (G1189A) restriction enzymes. The results showed that these mutations are available in tested animals. Parity had no significant effect on litter size. Also, the effects of different genotypes of GDF9 and BMPR1B had no significant effect on litter size. Further studies with a high number of animals with minimum relatedness for testing the association of these SNPs and others in the fecundity genes with reproductive traits may be worthwhile.

MiR-125b Regulates the Osteogenic Differentiation of Human Mesenchymal Stem Cells by Targeting BMPR1b.

BACKGROUND/AIMS: Osteogenic differentiation of mesenchymal stem cells (MSCs) plays a crucial role in bone regeneration and bone reparation. This complex process is regulated precisely and firmly by specific factors. Recent studies have demonstrated that miR-125b regulates osteogenic differentiation, but little is known about the molecular mechanisms of this regulation. Furthermore, how miR-125b regulates the osteogenic differentiation of MSCs still needs elucidation. METHODS: In the present study, human bone marrow-derived mesenchymal stem cells (hBMSCs) were isolated and induced to osteoblasts with miR-125b inhibition or overexpression. qRT-PCR and western blot analysis were used to detect the expression of osteogenic marker genes and proteins. Alkaline phosphatase (ALP) and Alizarin Red (ARS) staining were performed to evaluate the osteoblast phenotype. TargetScan, PicTar and miRanda database were used to predict the target gene of miR-125b. Dual luciferase reporter assay and RNA interference were performed to verify the target gene. Micro-CT imaging and histochemical staining were used to investigate the bone defect repair capacity of miR-125b in vivo. RESULTS: We observed that miR-125b was expressed at a low level during the osteogenic differentiation of hBMSCs. Then, we found that osteogenic marker genes were negatively regulated by miR-125b during the course of osteogenic differentiation, suggesting that miR-125b down regulation plays an important role in the process of osteogenic differentiation. Bioinformatics approaches using miRNA target prediction algorithms indicated that the bone morphogenetic protein type Ib receptor (BMPR1b) is a potential target of miR-125b. The results of the dual luciferase reporter assay indicated that miR-125b binds to the 3'-UTR of the BMPR1b gene. We observed that knockdown of BMPR1b by siRNA inhibited the osteogenic differentiation of hBMSCs. Furthermore, by co-transfecting cells with an miR-125b inhibitor and si-BMPR1b, we found that the osteogenic capacity of the cells transfected with miR-125b inhibitor was blocked upon knockdown of BMPR1b. In vivo, demineralized bone matrix (DBM) was composited with hBMSCs as a scaffold to repair segmental femoral defects. By inhibiting the expression of miR-125b, hBMSCs showed a better capacity to repair bone defects. CONCLUSIONS: Taken together, our study demonstrated that miR-125b regulated the osteogenic differentiation of hBMSCs by targeting BMPR1b and that inhibiting miR-125b expression could enhance the capacity of bone defect repair in vivo.