Clinical significance of APOB inactivation in hepatocellular carcinoma.

Recent findings from The Cancer Genome Atlas project have provided a comprehensive map of genomic alterations that occur in hepatocellular carcinoma (HCC), including unexpected mutations in apolipoprotein B (APOB). We aimed to determine the clinical significance of this non-oncogenetic mutation in HCC. An Apob gene signature was derived from genes that differed between control mice and mice treated with siRNA specific for Apob (1.5-fold difference; P < 0.005). Human gene expression data were collected from four independent HCC cohorts (n = 941). A prediction model was constructed using Bayesian compound covariate prediction, and the robustness of the APOB gene signature was validated in HCC cohorts. The correlation of the APOB signature with previously validated gene signatures was performed, and network analysis was conducted using ingenuity pathway analysis. APOB inactivation was associated with poor prognosis when the APOB gene signature was applied in all human HCC cohorts. Poor prognosis with APOB inactivation was consistently observed through cross-validation with previously reported gene signatures (NCIP A, HS, high-recurrence SNUR, and high RS subtypes). Knowledge-based gene network analysis using genes that differed between low-APOB and high-APOB groups in all four cohorts revealed that low-APOB activity was associated with upregulation of oncogenic and metastatic regulators, such as HGF, MTIF, ERBB2, FOXM1, and CD44, and inhibition of tumor suppressors, such as TP53 and PTEN. In conclusion, APOB inactivation is associated with poor outcome in patients with HCC, and APOB may play a role in regulating multiple genes involved in HCC development.

Wnt signaling is regulated by endoplasmic reticulum retention.

Precise regulation of Wnt signaling is important in many contexts, as in development of the vertebrate forebrain, where excessive or ectopic Wnt signaling leads to severe brain defects. Mutation of the widely expressed oto gene causes loss of the anterior forebrain during mouse embryogenesis. Here we report that oto is the mouse ortholog of the gpi deacylase gene pgap1, and that the endoplasmic reticulum (ER)-resident Oto protein has a novel and deacylase-independent function during Wnt maturation. Oto increases the hydrophobicities of Wnt3a and Wnt1 by promoting the addition of glycophosphatidylinositol (gpi)-like anchors to these Wnts, which results in their retention in the ER. We also report that oto-deficient embryos exhibit prematurely robust Wnt activity in the Wnt1 domain of the early neural plate. We examine the effect of low oto expression on Wnt1 in vitro by knocking down endogenous oto expression in 293 and M14 melanoma cells using shRNA. Knockdown of oto results in increased Wnt1 secretion which is correlated with greatly enhanced canonical Wnt activity. These data indicate that oto deficiency increases Wnt signaling in vivo and in vitro. Finally, we address the mechanism of Oto-mediated Wnt retention under oto-abundant conditions, by cotransfecting Wnt1 with gpi-specific phospholipase D (GPI-PLD). The presence of GPI-PLD in the secretory pathway results in increased secretion of soluble Wnt1, suggesting that the gpi-like anchor lipids on Wnt1 mediate its retention in the ER. These data now provide a mechanistic framework for understanding the forebrain defects in oto mice, and support a role for Oto-mediated Wnt regulation during early brain development. Our work highlights a critical role for ER retention in regulating Wnt signaling in the mouse embryo, and gives insight into the notoriously inefficient secretion of Wnts.

Functional architecture of atrophins.

Vertebrate genomes harbor two Atrophin genes, Atrophin-1 (Atn1) and Atrophin-2 (Atn2). The Atn1 locus produces a single polypeptide, whereas two different protein products are expressed from the Atn2 (also known as Rere) locus. A long, or full-length, form contains an amino-terminal MTA-2-homologous domain followed by an Atrophin-1-related domain. A short form, expressed via an internal promoter, consists solely of the Atrophin domain. Atrophin-1 can be co-immunoprecipitated along with Atrophin-2, suggesting that the Atrophins ordinarily function together. Mutations that disrupt the expression of the long form of Atrophin-2 disrupt early embryonic development. To determine the requirement for Atrophin-1 during development we generated a null allele. Somewhat surprisingly we found that Atrophin-1 function is dispensable. To gain a better understanding of the requirement for Atrophin function during development, an analysis of the functional domains of the three different gene products was carried out. Taken together, these data suggest that Atrophins function as bifunctional transcriptional regulators. The long form of Atrophin-2 has a transcriptional repression activity that is not found in the other Atrophin polypeptides and that is required for normal embryogenesis. Atrophin-1 and the short form of Atrophin-2, on the other hand, can act as potent and evolutionarily conserved transcriptional activators.