Bringing bioinformatics to schools with the 4273pi project

Over the last few decades, the nature of life sciences research has changed enormously, generating a need for a workforce with a variety of computational skills such as those required to store, manage, and analyse the large biological datasets produced by next-generation sequencing. Those with such expertise are increasingly in demand for employment in both research and industry. Despite this, bioinformatics education has failed to keep pace with advances in research. At secondary school level, computing is often taught in isolation from other sciences, and its importance in biological research is not fully realised, leaving pupils unprepared for the computational component of Higher Education and, subsequently, research in the life sciences. The 4273pi Bioinformatics at School project (https://4273pi.org) aims to address this issue by designing and delivering curriculum-linked, hands-on bioinformatics workshops for secondary school biology pupils, with an emphasis on equitable access. So far, we have reached over 180 schools across Scotland through visits or teacher events, and our open education resources are used internationally. Here, we describe our project, our aims and motivations, and the practical lessons we have learned from implementing a successful bioinformatics education project over the last 5 years.     

A nonsynonymous SNP within PCDH15 is associated with lipid traits in familial combined hyperlipidemia

Familial combined hyperlipidemia (FCHL) is a common lipid disorder characterized by the presence of multiple lipoprotein phenotypes that increase the risk of premature coronary heart disease. In a previous study, we identified an intragenic microsatellite marker within the protocadherin 15 (PCDH15) gene to be associated with high triglycerides (TGs) in Finnish dyslipidemic families. In this study we analyzed all four known nonsynonymous SNPs within PCDH15 in 1,268 individuals from Finnish and Dutch multigenerational families with FCHL. Association analyses of quantitative traits for SNPs were performed using the QTDT test. The nonsynonymous SNP rs10825269 resulted in a P = 0.0006 for the quantitative TG trait. Additional evidence for association was observed with the same SNP for apolipoprotein B levels (apo-B) (P = 0.0001) and total cholesterol (TC) levels (P = 0.001). None of the other three SNPs tested showed a significant association with any lipid-related trait. We investigated the expression of PCDH15 in different human tissues and observed that PCDH15 is expressed in several tissues including liver and pancreas. In addition, we measured the plasma lipid levels in mice with loss-of-function mutations in Pcdh15 (Pcdh15^av-Tg and Pcdh15^av-3J) to investigate possible abnormalities in their lipid profile. We observed a significant difference in plasma TG and TC concentrations for the Pcdh15^av-3J carriers when compared with the wild type (P = 0.013 and P = 0.044, respectively). Our study suggests that PCDH15 is associated with lipid abnormalities.     

Phosphorylation regulates the Star-PAP-PIPKIα interaction and directs specificity toward mRNA targets

Star-PAP is a nuclear non-canonical poly(A) polymerase (PAP) that shows specificity toward mRNA targets. Star-PAP activity is stimulated by lipid messenger phosphatidyl inositol 4,5 bisphoshate (PI4,5P₂) and is regulated by the associated Type I phosphatidylinositol-4-phosphate 5-kinase that synthesizes PI4,5P₂ as well as protein kinases. These associated kinases act as coactivators of Star-PAP that regulates its activity and specificity toward mRNAs, yet the mechanism of control of these interactions are not defined. We identified a phosphorylated residue (serine 6, S6) on Star-PAP in the zinc finger region, the domain required for PIPKIα interaction. We show that S6 is phosphorylated by CKIα within the nucleus which is required for Star-PAP nuclear retention and interaction with PIPKIα. Unlike the CKIα mediated phosphorylation at the catalytic domain, Star-PAP S6 phosphorylation is insensitive to oxidative stress suggesting a signal mediated regulation of CKIα activity. S6 phosphorylation together with coactivator PIPKIα controlled select subset of Star-PAP target messages by regulating Star-PAP-mRNA association. Our results establish a novel role for phosphorylation in determining Star-PAP target mRNA specificity and regulation of 3′-end processing.     

Effects of cinnamaldehyde on the glucose transport activity of GLUT1

There is accumulating evidence that cinnamon extracts contain components that enhance insulin action. However, little is know about the effects of cinnamon on non-insulin stimulated glucose uptake. Therefore, the effects of cinnamaldehyde on the glucose transport activity of GLUT1 in L929 fibroblast cells were examined under both basal conditions and conditions where glucose uptake is activated by glucose deprivation. The data reveal that cinnamaldehyde has a dual action on the glucose transport activity of GLUT1. Under basal conditions it stimulates glucose uptake and reaches a 3.5 fold maximum stimulation at 2.0 mM. However, cinnamaldehyde also inhibits the activation of glucose uptake by glucose deprivation in a dose dependent manner. Experiments with cinnamaldehyde analogs reveal that these activities are dependent on the α,β-unsaturated aldehyde structural motif in cinnamaldehyde. The inhibitory, but not the stimulatory activity of cinnamaldehyde was maintained after a wash-recovery period. Pretreatment of cinnamaldehyde with thiol-containing compounds, such as β-mercaptoethanol or cysteine, blocked the inhibitory activity of cinnamaldehyde. These results suggest that cinnamaldehyde inhibits the activation of GLUT1 by forming a covalent link to target cysteine residue/s. This dual activity of cinnamaldehyde on the transport activity of GLUT1 suggests that cinnamaldehyde is not a major contributor to the anti-diabetic properties of cinnamon.     

Sequencing three crocodilian genomes to illuminate the evolution of archosaurs and amniotes

The International Crocodilian Genomes Working Group (ICGWG) will sequence and assemble the American alligator (Alligator mississippiensis), saltwater crocodile (Crocodylus porosus) and Indian gharial (Gavialis gangeticus) genomes. The status of these projects and our planned analyses are described.     

Cyclops: new modular software suite for cryo-EM

Cyclops is a new computer program designed as a graphical front-end that allows easy control and interaction with tasks and programs for 3D reconstruction of biological complexes using cryo-electron microscopy. Cyclops' current plug-ins are designed for automated particle picking and include two new algorithms, automated carbon masking and quaternion based rotation space sampling, which are also presented here. Additional plug-ins are in the pipeline. Cyclops allows straightforward organization and visualization of all data and tasks and allows both interactive and batch-wise processing. Furthermore, it was designed for straightforward implementation in grid architectures. As a front-end to a collection of programs it provides a common interface to these programs, thus enhancing the usability of the suite and the productivity of the user.     

Rank–rank hypergeometric overlap: identification of statistically significant overlap between gene-expression signatures

Comparing independent high-throughput gene-expression experiments can generate hypotheses about which gene-expression programs are shared between particular biological processes. Current techniques to compare expression profiles typically involve choosing a fixed differential expression threshold to summarize results, potentially reducing sensitivity to small but concordant changes. We present a threshold-free algorithm called Rank–rank Hypergeometric Overlap (RRHO). This algorithm steps through two gene lists ranked by the degree of differential expression observed in two profiling experiments, successively measuring the statistical significance of the number of overlapping genes. The output is a graphical map that shows the strength, pattern and bounds of correlation between two expression profiles. To demonstrate RRHO sensitivity and dynamic range, we identified shared expression networks in cancer microarray profiles driving tumor progression, stem cell properties and response to targeted kinase inhibition. We demonstrate how RRHO can be used to determine which model system or drug treatment best reflects a particular biological or disease response. The threshold-free and graphical aspects of RRHO complement other rank-based approaches such as Gene Set Enrichment Analysis (GSEA), for which RRHO is a 2D analog. Rank–rank overlap analysis is a sensitive, robust and web-accessible method for detecting and visualizing overlap trends between two complete, continuous gene-expression profiles. A web-based implementation of RRHO can be accessed at http://systems.crump.ucla.edu/rankrank/.     

Epithelial cell kinetics in the descending colon of the rat

Epithelial cell kinetics were investigated in the descending colon of the rat. The number of cells per crypt was found to be approximately 625, with 33 cells per cell column and 19 cell columns per crypt circumference. The growth fraction of the colonic crypt was 0.42, and proliferating cells were situated largely in the lower half of the crypt. The cell cycle time was 50.5 h, with values for the G1, S and G2 phases of 40.0, 7.6 and 2.9 h respectively. Cell migration studies showed that it took 60-72 h for a cell to migrate from the upper border of the proliferative cell compartment in the crypt to the luminal surface of the colon. Data were also obtained from continuous labelling with tritiated thymidine and from studying the circadian rhythm of proliferative activity, which suggest that the cells in the bottom of the crypt may constitute a separate, more slowly cycling (stem)cell compartment.