High-throughput discovery of genetic determinants of circadian misalignment.

Circadian systems provide a fitness advantage to organisms by allowing them to adapt to daily changes of environmental cues, such as light/dark cycles. The molecular mechanism underlying the circadian clock has been well characterized. However, how internal circadian clocks are entrained with regular daily light/dark cycles remains unclear. By collecting and analyzing indirect calorimetry (IC) data from more than 2000 wild-type mice available from the International Mouse Phenotyping Consortium (IMPC), we show that the onset time and peak phase of activity and food intake rhythms are reliable parameters for screening defects of circadian misalignment. We developed a machine learning algorithm to quantify these two parameters in our misalignment screen (SyncScreener) with existing datasets and used it to screen 750 mutant mouse lines from five IMPC phenotyping centres. Mutants of five genes (Slc7a11, Rhbdl1, Spop, Ctc1 and Oxtr) were found to be associated with altered patterns of activity or food intake. By further studying the Slc7a11tm1a/tm1a mice, we confirmed its advanced activity phase phenotype in response to a simulated jetlag and skeleton photoperiod stimuli. Disruption of Slc7a11 affected the intercellular communication in the suprachiasmatic nucleus, suggesting a defect in synchronization of clock neurons. Our study has established a systematic phenotype analysis approach that can be used to uncover the mechanism of circadian entrainment in mice.

Increased Expressions of Matrix Metalloproteinases (MMPs) in Prostate Cancer Tissues of Men with Type 2 Diabetes.

Type 2 diabetes (T2D) is associated with worse prognosis of prostate cancer (PCa). The molecular mechanisms behind this association are still not fully understood. The aim of this study was to identify key factors, which contribute to the more aggressive PCa phenotype in patients with concurrent T2D. Therefore, we investigated benign and PCa tissue of PCa patients with and without diabetes using real time qPCR. Compared to patients without diabetes, patients with T2D showed a decreased E-cadherin/N-cadherin (CDH1/CDH2) ratio in prostate tissue, indicating a switch of epithelial-mesenchymal transition (EMT), which is a pivotal process in carcinogenesis. In addition, the gene expression levels of matrix metalloproteinases (MMPs) and CC chemokine ligands (CCLs) were higher in prostate samples of T2D patients. Next, prostate adenocarcinoma PC3 cells were treated with increasing glucose concentrations to replicate hyperglycemia in vitro. In these cells, high glucose induced expressions of MMPs and CCLs, which showed significant positive associations with the proliferation marker proliferating cell nuclear antigen (PCNA). These results indicate that in prostate tissue of men with T2D, hyperglycemia may induce EMT, increase MMP and CCL gene expressions, which in turn activate invasion and inflammatory processes accelerating the progression of PCa.

A truncating Aspm allele leads to a complex cognitive phenotype and region-specific reductions in parvalbuminergic neurons.

Neurodevelopmental disorders are heterogeneous and identifying shared genetic aetiologies and converging signalling pathways affected could improve disease diagnosis and treatment. Truncating mutations of the abnormal spindle-like microcephaly associated (ASPM) gene cause autosomal recessive primary microcephaly (MCPH) in humans. ASPM is a positive regulator of Wnt/ß-Catenin signalling and controls symmetric to asymmetric cell division. This process balances neural progenitor proliferation with differentiation during embryogenesis, the malfunction of which could interfere with normal brain development. ASPM mutations may play a role also in other neurodevelopmental disorders, nevertheless, we lack the details of how or to what extent. We therefore assessed neurodevelopmental disease and circuit endophenotypes in mice with a truncating Aspm1-7 mutation. Aspm1-7 mice exhibited impaired short- and long-term object recognition memory and markedly enhanced place learning in the IntelliCage®. This behaviour pattern is reminiscent of a cognitive phenotype seen in mouse models and patients with a rare form of autism spectrum disorder (ASD) as well as in mouse models of altered Wnt signalling. These alterations were accompanied by ventriculomegaly, corpus callosum dysgenesis and decreased parvalbumin (PV)+ interneuron numbers in the hippocampal Cornu Ammonis (CA) region and thalamic reticular nucleus (TRN). PV+ cell number correlated to object recognition (CA and TRN) and place learning (TRN). This opens the possibility that, as well as causing MCPH, mutant ASPM potentially contributes to other neurodevelopmental disorders such as ASD through altered parvalbuminergic interneuron development affecting cognitive behaviour. These findings provide important information for understanding the genetic overlap and improved treatment of neurodevelopmental disorders associated with ASPM.

Prevalence of sexual dimorphism in mammalian phenotypic traits.

The role of sex in biomedical studies has often been overlooked, despite evidence of sexually dimorphic effects in some biological studies. Here, we used high-throughput phenotype data from 14,250 wildtype and 40,192 mutant mice (representing 2,186 knockout lines), analysed for up to 234 traits, and found a large proportion of mammalian traits both in wildtype and mutants are influenced by sex. This result has implications for interpreting disease phenotypes in animal models and humans.

Toxicity modelling of Plk1-targeted therapies in genetically engineered mice and cultured primary mammalian cells.

High attrition rates of novel anti-cancer drugs highlight the need for improved models to predict toxicity. Although polo-like kinase 1 (Plk1) inhibitors are attractive candidates for drug development, the role of Plk1 in primary cells remains widely unexplored. Therefore, we evaluated the utility of an RNA interference-based model to assess responses to an inducible knockdown (iKD) of Plk1 in adult mice. Here we show that Plk1 silencing can be achieved in several organs, although adverse events are rare. We compared responses in Plk1-iKD mice with those in primary cells kept under controlled culture conditions. In contrast to the addiction of many cancer cell lines to the non-oncogene Plk1, the primary cells' proliferation, spindle assembly and apoptosis exhibit only a low dependency on Plk1. Responses to Plk1-depletion, both in cultured primary cells and in our iKD-mouse model, correspond well and thus provide the basis for using validated iKD mice in predicting responses to therapeutic interventions.

Combination of in silico and in situ hybridisation approaches to identify potential Dll1 associated miRNAs during mouse embryogenesis.

MicroRNAs (miRNAs) have regulatory functions during vertebrate embryogenesis. They are short approximately 21bp long endogenously expressed single-stranded RNAs, which preferentially bind to complementary sequences in the 3' untranslated regions (UTR) of mRNAs and typically down-regulate the respective target mRNAs by translational repression or enhanced mRNA degradation. The Notch ligand Delta-like 1 (Dll1) is expressed in a highly dynamic pattern and has pleiotropic functions during embryogenesis and in adult tissues. Here, we report an interspecies in silico analysis to identify 16 miRNAs, which potentially bind to the mouse, human and chicken Dll1 3'UTRs. To analyze whether these miRNAs could regulate Dll1 gene expression during somitogenesis and neurogenesis, we performed a systematic whole mount in situ hybridisation screen, followed by radioactive in situ hybridisation on sections, using LNA modified DNA probes in mouse embryos. We find that 7 miRNAs (miR-34a, miR-103, miR-107, miR-130a, miR-130b, miR-449a and miR-449c) are expressed in developing somites, limbs, restricted regions of the brain and neural tube between 9.5 dpc and 12.5 dpc. This suggests that these miRNAs could possibly target the Dll1 3'UTR in these regions. The other miRNAs are not expressed or below the detection limit and thus are unlikely to regulate Dll1 at the analyzed embryonic stages.