A burst of genomic innovation at the origin of placental mammals mediated embryo implantation.

The origin of embryo implantation in mammals ~148 million years ago was a dramatic shift in reproductive strategy, yet the molecular changes that established mammal implantation are largely unknown. Although progesterone receptor signalling predates the origin of mammals and is highly conserved in, and critical for, successful mammal pregnancy, it alone cannot explain the origin and subsequent diversity of implantation strategies throughout the placental mammal radiation. MiRNAs are known to be flexible and dynamic regulators with a well-established role in the pathophysiology of mammal placenta. We propose that a dynamic core microRNA (miRNA) network originated early in placental mammal evolution, responds to conserved mammal pregnancy cues (e.g. progesterone), and facilitates species-specific responses. Here we identify 13 miRNA gene families that arose at the origin of placental mammals and were subsequently retained in all descendent lineages. The expression of these miRNAs in response to early pregnancy molecules is regulated in a species-specific manner in endometrial epithelia of species with extreme implantation strategies (i.e. bovine and human). Furthermore, this set of miRNAs preferentially target proteins under positive selective pressure on the ancestral eutherian lineage. Discovery of this core embryo implantation toolkit and specifically adapted proteins helps explain the origin and evolution of implantation in mammals.

The role of CAPG in molecular communication between the embryo and the uterine endometrium: Is its function conserved in species with different implantation strategies?

During the preimplantation period of pregnancy in eutherian mammals, transcriptional and proteomic changes in the uterine endometrium are required to facilitate receptivity to an implanting blastocyst. These changes are mediated, in part, by proteins produced by the developing conceptus (inner cell mass and extraembryonic membranes). We hypothesized that this common process in early pregnancy in eutheria may be facilitated by highly conserved conceptus-derived proteins such as macrophage capping protein (CAPG). We propose that CAPG may share functionality in modifying the transcriptome of the endometrial epithelial cells to facilitate receptivity to implantation in species with different implantation strategies. A recombinant bovine form of CAPG (91% sequence identity between bovine and human) was produced and bovine endometrial epithelial (bEECs) and stromal (bESCs) and human endometrial epithelial cells (hEECs) were cultured for 24 hours with and without recombinant bovine CAPG (rbCAPG). RNA sequencing and quantitative real-time PCR analysis were used to assess the transcriptional response to rbCAPG (Control, vehicle, CAPG 10, 100, 1000 ng/mL: n = 3 biological replicates per treatment per species). Treatment of bEECs with CAPG resulted in alterations in the abundance of 1052 transcripts (629 increased and 423 decreased) compared to vehicle controls. Treatment of hEECs with bovine CAPG increased expression of transcripts previously known to interact with CAPG in different systems (CAPZB, CAPZA2, ADD1, and ADK) compared with vehicle controls (P < .05). In conclusion, we have demonstrated that CAPG, a highly conserved protein in eutherian mammals, elicits a transcriptional response in the endometrial epithelium in species with different implantation strategies that may contribute to pregnancy success.

Functional characterisation of a novel ovarian cancer cell line, NUOC-1.

BACKGROUND: Cell lines provide a powerful model to study cancer and here we describe a new spontaneously immortalised epithelial ovarian cancer cell line (NUOC-1) derived from the ascites collected at a time of primary debulking surgery for a mixed endometrioid / clear cell / High Grade Serous (HGS) histology. RESULTS: This spontaneously immortalised cell line was found to maintain morphology and epithelial markers throughout long-term culture. NUOC-1 cells grow as an adherent monolayer with a doubling time of 58 hours. The cells are TP53 wildtype, positive for PTEN, HER2 and HER3 expression but negative for oestrogen, progesterone and androgen receptor expression. NUOC-1 cells are competent in homologous recombination and non-homologous end joining, but base excision repair defective. Karyotype analysis demonstrated a complex tetraploid karyotype. SNP array analysis of parent and derived subpopulations (NUOC-1-A1 and NUOC-1-A2) cells demonstrated heterogeneous cell populations with numerous copy number alterations and a pro-amplification phenotype. The characteristics of this new cell line lends it to be an excellent model for investigation of a number of the identified targets. MATERIALS AND METHODS: The cell line has been characterised for growth, drug sensitivity, expression of common ovarian markers and mutations, clonogenic potential and ability to form xenografts in SCID mice. Copy number changes and clonal evolution were assessed by SNP arrays.

Novel missense mutations in a conserved loop between ERCC6 (CSB) helicase motifs V and VI: Insights into Cockayne syndrome.

Cockayne syndrome is caused by biallelic ERCC8 (CSA) or ERCC6 (CSB) mutations and is characterized by growth restriction, microcephaly, developmental delay, and premature pathological aging. Typically affected patients also have dermal photosensitivity. Although Cockayne syndrome is considered a DNA repair disorder, patients with UV-sensitive syndrome, with ERCC8 (CSA) or ERCC6 (CSB) mutations have indistinguishable DNA repair defects, but none of the extradermal features of Cockayne syndrome. We report novel missense mutations affecting a conserved loop in the ERCC6 (CSB) protein, associated with the Cockayne syndrome phenotype. Indeed, the amino acid sequence of this loop is more highly conserved than the adjacent helicase motifs V and VI, suggesting that this is a crucial structural component of the SWI/SNF family of proteins, to which ERCC6 (CSB) belongs. These comprise two RecA-like domains, separated by an interdomain linker, which interact through helicase motif VI. As the observed mutations are likely to act through destabilizing the tertiary protein structure, this prompted us to re-evaluate ERCC6 (CSB) mutation data in relation to the structure of SWI/SNF proteins. Our analysis suggests that antimorphic mutations cause Cockayne syndrome and that biallelic interdomain linker deletions produce more severe phenotypes. Based on our observations, we propose that further investigation of the pathogenic mechanisms underlying Cockayne syndrome should focus on the effect of antimorphic rather than null ERCC6 (CSB) mutations.

The Cockayne Syndrome Natural History (CoSyNH) study: clinical findings in 102 individuals and recommendations for care.

PURPOSE: Cockayne syndrome (CS) is a rare, autosomal-recessive disorder characterized by microcephaly, impaired postnatal growth, and premature pathological aging. It has historically been considered a DNA repair disorder; fibroblasts from classic patients often exhibit impaired transcription-coupled nucleotide excision repair. Previous studies have largely been restricted to case reports and small series, and no guidelines for care have been established. METHODS: One hundred two study participants were identified through a network of collaborating clinicians and the Amy and Friends CS support groups. Families with a diagnosis of CS could also self-recruit. Comprehensive clinical information for analysis was obtained directly from families and their clinicians. RESULTS AND CONCLUSION: We present the most complete evaluation of Cockayne syndrome to date, including detailed information on the prevalence and onset of clinical features, achievement of neurodevelopmental milestones, and patient management. We confirm that the most valuable prognostic factor in CS is the presence of early cataracts. Using this evidence, we have created simple guidelines for the care of individuals with CS. We aim to assist clinicians in the recognition, diagnosis, and management of this condition and to enable families to understand what problems they may encounter as CS progresses.Genet Med 18 5, 483-493.

A homoplasmic mtDNA variant can influence the phenotype of the pathogenic m.7472Cins MTTS1 mutation: are two mutations better than one?

Mutations in mitochondrial tRNA (mt-tRNA) genes are well recognized as a common cause of human disease, exhibiting a significant degree of clinical heterogeneity. While these differences are explicable, in part, by differences in the innate pathogenicity of the mutation, its distribution and abundance, other factors, including nuclear genetic background, mitochondrial DNA (mtDNA) haplotype and additional mtDNA mutations may influence the expression of mt-tRNA mutations. We describe the clinical, biochemical and molecular findings in a family with progressive myopathy, deafness and diabetes and striking respiratory chain abnormalities due to a well-characterized heteroplasmic mt-tRNA mutation in the mt-tRNA(Ser(UCN)) (MTTS1) gene. In addition to the m.7472Cins mutation, all individuals were homoplasmic for another variant, m.7472A > C, affecting the adjacent nucleotide in the mt-tRNA(Ser(UCN)) structure. In addition to available patient tissues, we have analysed transmitochondrial cybrid clones harbouring homoplasmic levels of m.7472A > C and varying levels of the m.7472Cins mutation in an attempt to clarify the precise role of the m.7472A > C transversion in the underlying respiratory chain abnormality. Evidence from both in vivo and in vitro studies demonstrate that the m.7472A > C is able to modify the expression of the m.7472Cins mutation and would suggest that it is not a neutral variant but appears to cause a biochemical defect by itself, confirming that homoplasmic mtDNA variants can modulate the phenotypic expression of pathogenic, heteroplasmic mtDNA mutations.