The localization of amyloid precursor protein to ependymal cilia in vertebrates and its role in ciliogenesis and brain development in zebrafish

Amyloid precursor protein (APP) is expressed in many tissues in human, mice and in zebrafish. In zebrafish, there are two orthologues, Appa and Appb. Interestingly, some cellular processes associated with APP overlap with cilia-mediated functions. Whereas the localization of APP to primary cilia of in vitro-cultured cells has been reported, we addressed the presence of APP in motile and in non-motile sensory cilia and its potential implication for ciliogenesis using zebrafish, mouse, and human samples. We report that Appa and Appb are expressed by ciliated cells and become localized at the membrane of cilia in the olfactory epithelium, otic vesicle and in the brain ventricles of zebrafish embryos. App in ependymal cilia persisted in adult zebrafish and was also detected in mouse and human brain. Finally, we found morphologically abnormal ependymal cilia and smaller brain ventricles in appa-/-appb-/- mutant zebrafish. Our findings demonstrate an evolutionary conserved localisation of APP to cilia and suggest a role of App in ciliogenesis and cilia-related functions

Cell stemness is maintained upon concurrent expression of RB and the mitochondrial ribosomal protein S18-2

Stemness encompasses the capability of a cell for self-renewal and differentiation. The stem cell maintains a balance between proliferation, quiescence, and regeneration via interactions with the microenvironment. Previously, we showed that ectopic expression of the mitochondrial ribosomal protein S18-2 (MRPS18-2) led to immortalization of primary fibroblasts, accompanied by induction of an embryonic stem cell (ESC) phenotype. Moreover, we demonstrated interaction between S18-2 and the retinoblastoma-associated protein (RB) and hypothesized that the simultaneous expression of RB and S18-2 is essential for maintaining cell stemness. Here, we experimentally investigated the role of S18-2 in cell stemness and differentiation. Concurrent expression of RB and S18-2 resulted in immortalization of Rb1−/− primary mouse embryonic fibroblasts and in aggressive tumor growth in severe combined immunodeficiency mice. These cells, which express both RB and S18-2 at high levels, exhibited the potential to differentiate into various lineages in vitro, including osteogenic, chondrogenic, and adipogenic lineages. Mechanistically, S18-2 formed a multimeric protein complex with prohibitin and the ring finger protein 2 (RNF2). This molecular complex increased the monoubiquitination of histone H2ALys119, a characteristic trait of ESCs, by enhanced E3-ligase activity of RNF2. Furthermore, we found enrichment of KLF4 at the S18-2 promoter region and that the S18-2 expression is positively correlated with KLF4 levels. Importantly, knockdown of S18-2 in zebrafish larvae led to embryonic lethality. Collectively, our findings suggest an important role for S18-2 in cell stemness and differentiation and potentially also in cancerogenesis

β-Amyloid precursor protein-b is essential for Mauthner cell development in the zebrafish in a Notch-dependent manner

Amyloid precursor protein (APP) is a transmembrane glycoprotein that has been the subject of intense research because of its implication in Alzheimer's disease. However, the physiological function of APP in the development and maintenance of the central nervous system remains largely unknown. We have previously shown that the APP homologue in zebrafish (Danio rerio), Appb, is required for motor neuron patterning and formation. Here we study the function of Appb during neurogenesis in the zebrafish hindbrain. Partial knockdown of Appb using antisense morpholino oligonucleotides blocked the formation of the Mauthner neurons, uni- or bilaterally, with an aberrant behavior as a consequence of this cellular change. The Appb morphants had decreased neurogenesis, increased notch signaling and notch1a expression at the expense of deltaA/D expression. The Mauthner cell development could be restored either by a general decrease in Notch signaling through γ-secretase inhibition or by a partial knock down of Notch1a. Together, this demonstrates the importance of Appb in neurogenesis and for the first time shows the essential requirement of Appb in the formation of a specific cell type, the Mauthner cell, in the hindbrain during development. Our results suggest that Appb-regulated neurogenesis is mediated through balancing the Notch1a signaling pathway and provide new insights into the development of the Mauthner cell

Fluoranthene and phenantrene, two predominant PAHs in heat-prepared food, do not influence the frequency of micronucleated mouse erythrocytes induced by other PAHs

Polycyclic aromatic hydrocarbons (PAHs) occur in complex mixtures present in the human environment. Because of the carcinogenic properties of some of these PAHs, they raise concerns regarding health and food safety. Because the occurrence of benzo[a]pyrene, chrysene, benz[a]anthracene, and benzo[b]fluoranthene (PAH4) are considered markers for other genotoxic PAHs in foodstuffs, the European Union has put a maximum level of PAH4 in some foodstuffs. Fluoranthene (Flu) and phenanthrene (Phe), two other PAHs, are not classified as genotoxic and are abundant at rather high concentrations in food. Inasmuch as PAH4, Flu, and Phe are metabolized by the same cytochrome P450 pathway system, it is important to clarify whether Phe and Flu influence the genotoxicity of PAH4. We have analyzed the genotoxic response of Phe and Flu, separately and together, as well as in combination with different low doses of PAH4. In all experiments we used the flow cytometer-based micronucleus test in vivo. Phe and Flu, when administered separately, did not show any dose-related effect on the frequency of micronucleated polychromatic erythrocytes (fMNPCE). Nor did a mixture of Phe and Flu change the fMNPCEs. Phe and Flu did not significantly change the fMNPCE of PAH4-exposed FVB and BALB/c mice

The HEATOX project

HEATOX is the acronym for a European Union-funded project entitled Heat-Generated Food Toxicants: Identification, Characterization, and Risk Minimization. Acrylamide will be the main experimental focus, but identification of unknown toxicants in heated carbohydrate-rich foods will also be attempted. The project includes research on formation chemistry, food technology, analytical methods, hazard characterization, and exposure assessment. The results will finally be used in risk assessment and risk management advice

Internal Doses of Glycidol in Children and Estimation of Associated Cancer Risk

The general population is exposed to the genotoxic carcinogen glycidol via food containing refined edible oils where glycidol is present in the form of fatty acid esters. In this study, internal (in vivo) doses of glycidol were determined in a cohort of 50 children and in a reference group of 12 adults (non-smokers and smokers). The lifetime in vivo doses and intakes of glycidol were calculated from the levels of the hemoglobin (Hb) adduct N-(2,3-dihydroxypropyl)valine in blood samples from the subjects, demonstrating a fivefold variation between the children. The estimated mean intake (1.4 μg/kg/day) was about two times higher, compared to the estimated intake for children by the European Food Safety Authority. The data from adults indicate that the non-smoking and smoking subjects are exposed to about the same or higher levels compared to the children, respectively. The estimated lifetime cancer risk (200/105) was calculated by a multiplicative risk model from the lifetime in vivo doses of glycidol in the children, and exceeds what is considered to be an acceptable cancer risk. The results emphasize the importance to further clarify exposure to glycidol and other possible precursors that could give a contribution to the observed adduct levels