Loss of Ing3 Expression Results in Growth Retardation and Embryonic Death

The ING3 candidate tumour suppressor belongs to a family of histone modifying proteins involved in regulating cell proliferation, senescence, apoptosis, chromatin remodeling, and DNA repair. It is a stoichiometric member of the minimal NuA4 histone acetyl transferase (HAT) complex consisting of EAF6, EPC1, ING3, and TIP60. This complex is responsible for the transcription of an essential cascade of genes involved in embryonic development and in tumour suppression. ING3 has been linked to head and neck and hepatocellular cancers, although its status as a tumour suppressor has not been well established. Recent studies suggest a pro-metastasis role in prostate cancer progression. Here, we describe a transgenic mouse strain with insertional mutation of an UbC-mCherry expression cassette into the endogenous Ing3 locus, resulting in the disruption of ING3 protein expression. Homozygous mutants are embryonically lethal, display growth retardation, and severe developmental disorders. At embryonic day (E) 10.5, the last time point viable homozygous embryos were found, they were approximately half the size of heterozygous mice that develop normally. µCT analysis revealed a developmental defect in neural tube closure, resulting in the failure of formation of closed primary brain vesicles in homozygous mid-gestation embryos. This is consistent with high ING3 expression levels in the embryonic brains of heterozygous and wild type mice and its lack in homozygous mutant embryos that show a lack of ectodermal differentiation. Our data provide direct evidence that ING3 is an essential factor for normal embryonic development and that it plays a fundamental role in prenatal brain formation.

The spectraplakin Dystonin antagonizes YAP activity and suppresses tumourigenesis

Aberrant expression of the Spectraplakin Dystonin (DST) has been observed in various cancers, including those of the breast. However, little is known about its role in carcinogenesis. In this report, we demonstrate that Dystonin is a candidate tumour suppressor in breast cancer and provide an underlying molecular mechanism. We show that in MCF10A cells, Dystonin is necessary to restrain cell growth, anchorage-independent growth, self-renewal properties and resistance to doxorubicin. Strikingly, while Dystonin maintains focal adhesion integrity, promotes cell spreading and cell-substratum adhesion, it prevents Zyxin accumulation, stabilizes LATS and restricts YAP activation. Moreover, treating DST-depleted MCF10A cells with the YAP inhibitor Verteporfin prevents their growth. In vivo, the Drosophila Dystonin Short stop also restricts tissue growth by limiting Yorkie activity. As the two Dystonin isoforms BPAG1eA and BPAG1e are necessary to inhibit the acquisition of transformed features and are both downregulated in breast tumour samples and in MCF10A cells with conditional induction of the Src proto-oncogene, they could function as the predominant Dystonin tumour suppressor variants in breast epithelial cells. Thus, their loss could deem as promising prognostic biomarkers for breast cancer.

Inferring the metabolism of human orphan metabolites from their metabolic network context affirms human gluconokinase activity

Metabolic network reconstructions define metabolic information within a target organism and can therefore be used to address incomplete metabolic information. In the present study we used a computational approach to identify human metabolites whose metabolism is incomplete on the basis of their detection in humans but exclusion from the human metabolic network reconstruction RECON 1. Candidate solutions, composed of metabolic reactions capable of explaining the metabolism of these compounds, were then identified computationally from a global biochemical reaction database. Solutions were characterized with respect to how metabolites were incorporated into RECON 1 and their biological relevance. Through detailed case studies we show that biologically plausible non-intuitive hypotheses regarding the metabolism of these compounds can be proposed in a semi-automated manner, in an approach that is similar to de novo network reconstruction. We subsequently experimentally validated one of the proposed hypotheses and report that C9orf103, previously identified as a candidate tumour suppressor gene, encodes a functional human gluconokinase. The results of the present study demonstrate how semi-automatic gap filling can be used to refine and extend metabolic reconstructions, thereby increasing their biological scope. Furthermore, we illustrate how incomplete human metabolic knowledge can be coupled with gene annotation in order to prioritize and confirm gene functions.