Joha

Nature Methods 11, 713 (2014). doi:10.1038/nmeth.3006

Nature Methods. doi:10.1038/nmeth.3014

Authors: Pieter Mestdagh, Nicole Hartmann, Lukas Baeriswyl, Ditte Andreasen, Nathalie Bernard, Caifu Chen, David Cheo, Petula D'Andrade, Mike DeMayo, Lucas Dennis, Stefaan Derveaux, Yun Feng, Stephanie Fulmer-Smentek, Bernhard Gerstmayer, Julia Gouffon, Chris Grimley, Eric Lader, Kathy Y Lee, Shujun Luo, Peter Mouritzen, Aishwarya Narayanan, Sunali Patel, Sabine Peiffer, Silvia Rüberg, Gary Schroth, Dave Schuster, Jonathan M Shaffer, Elliot J Shelton, Scott Silveria, Umberto Ulmanella, Vamsi Veeramachaneni, Frank Staedtler, Thomas Peters, Toumy Guettouche & Jo Vandesompele

Publication date: 12 June 2014
Source:Cell Reports, Volume 7, Issue 5
Author(s): Natalia Gromak , Martin Dienstbier , Sara Macias , Mireya Plass , Eduardo Eyras , Javier F. Cáceres , Nicholas J. Proudfoot

Nature Structural & Molecular Biology 21, 599 (2014). doi:10.1038/nsmb.2837

Authors: Stefanie Jonas, Mary Christie, Daniel Peter, Dipankar Bhandari, Belinda Loh, Eric Huntzinger, Oliver Weichenrieder & Elisa Izaurralde

Nature Structural & Molecular Biology 21, 585 (2014). doi:10.1038/nsmb.2842

Authors: Grace X Y Zheng, Brian T Do, Dan E Webster, Paul A Khavari & Howard Y Chang

Nature Structural & Molecular Biology 21, 633 (2014). doi:10.1038/nsmb.2844

Authors: Jessica A Brown, David Bulkley, Jimin Wang, Max L Valenstein, Therese A Yario, Thomas A Steitz & Joan A Steitz

Nature Chemical Biology 10, 574 (2014). doi:10.1038/nchembio.1532

Authors: Toni Pfaffeneder, Fabio Spada, Mirko Wagner, Caterina Brandmayr, Silvia K Laube, David Eisen, Matthias Truss, Jessica Steinbacher, Benjamin Hackner, Olga Kotljarova, David Schuermann, Stylianos Michalakis, Olesea Kosmatchev, Stefan Schiesser, Barbara Steigenberger, Nada Raddaoui, Gengo Kashiwazaki, Udo Müller, Cornelia G Spruijt, Michiel Vermeulen, Heinrich Leonhardt, Primo Schär, Markus Müller & Thomas Carell

Nature Chemical Biology 10, 512 (2014). doi:10.1038/nchembio.1556

Authors: Kevin M Dean & Amy E Palmer

by David B. Searls

A recent proliferation of Massive Open Online Courses (MOOCs) and other web-based educational resources has greatly increased the potential for effective self-study in many fields. This article introduces a catalog of several hundred free video courses of potential interest to those wishing to expand their knowledge of bioinformatics and computational biology. The courses are organized into eleven subject areas modeled on university departments and are accompanied by commentary and career advice.

Publication date: 8 May 2014
Source:Cell, Volume 157, Issue 4
Author(s): Israel S. Fernández , Xiao-Chen Bai , Garib Murshudov , Sjors H.W. Scheres , V. Ramakrishnan
The cricket paralysis virus internal ribosome entry site (CrPV-IRES) is a folded structure in a viral mRNA that allows initiation of translation in the absence of any host initiation factors. By using recent advances in single-particle electron cryomicroscopy, we have solved the structure of CrPV-IRES bound to the ribosome of the yeast Kluyveromyces lactis in both the canonical and rotated states at overall resolutions of 3.7 and 3.8 Å, respectively. In both states, the pseudoknot PKI of the CrPV-IRES mimics a tRNA/mRNA interaction in the decoding center of the A site of the 40S ribosomal subunit. The structure and accompanying factor-binding data show that CrPV-IRES binding mimics a pretranslocation rather than initiation state of the ribosome. Translocation of the IRES by elongation factor 2 (eEF2) is required to bring the first codon of the mRNA into the A site and to allow the start of translation.

Graphical abstract

Teaser

A high-resolution structure of the cricket paralysis virus IRES bound to the eukaryotic ribosome reveals a surprising mechanism of translation initiation that requires a translocation step.

Nature Chemical Biology 10, 408 (2014). doi:10.1038/nchembio.1538

Author: Terry L. Sheppard

The faithful transmission of DNA methylation patterns through cell divisions is essential for the daughter cells to retain a proper cell identity. To achieve a comprehensive assessment of methylation fidelity, we implemented a genome-scale hairpin bisulfite sequencing approach to generate methylation data for DNA double strands simultaneously. We show here that methylation fidelity is increasing globally during differentiation of mouse embryonic stem cells (mESCs), and is particularly high in the promoter regions of actively expressed genes and positively correlated with active histone modification marks and binding of transcriptional factors. The majority of intermediately (40%-60%) methylated CpG dinucleotides are hemi-methylated and with low methylation fidelity, particularly in the differentiating mESCs. However, while 5-hmC and 5-mC tend to coexist, there is no significant correlation between 5hmC levels and methylation fidelity. Our findings may shed new light on our understanding of the origins of methylation variations and the mechanisms underlying DNA methylation transmission.

Nature Methods. doi:10.1038/nmeth.2970

Authors: Jacob M Tome, Abdullah Ozer, John M Pagano, Dan Gheba, Gary P Schroth & John T Lis

Guidelines for investigating causality of sequence variants in human disease

Nature 508, 7497 (2014). doi:10.1038/nature13127

Authors: D. G. MacArthur, T. A. Manolio, D. P. Dimmock, H. L. Rehm, J. Shendure, G. R. Abecasis, D. R. Adams, R. B. Altman, S. E. Antonarakis, E. A. Ashley, J. C. Barrett, L. G. Biesecker, D. F. Conrad, G. M. Cooper, N. J. Cox, M. J. Daly, M. B. Gerstein, D. B. Goldstein, J. N. Hirschhorn, S. M. Leal, L. A. Pennacchio, J. A. Stamatoyannopoulos, S. R. Sunyaev, D. Valle, B. F. Voight, W. Winckler & C. Gunter

The discovery of rare genetic variants is accelerating, and clear guidelines for distinguishing disease-causing sequence variants from the many potentially functional variants present in any human genome are urgently needed. Without rigorous standards we risk an acceleration of false-positive reports of causality, which would impede the translation of genomic research findings into the clinical diagnostic setting and hinder biological understanding of disease. Here we discuss the key challenges of assessing sequence variants in human disease, integrating both gene-level and variant-level support for causality. We propose guidelines for summarizing confidence in variant pathogenicity and highlight several areas that require further resource development.

Medical genomics: Gather and use genetic data in health care

Nature 508, 7497 (2014). doi:10.1038/508451a

Author: Geoffrey Ginsburg

Research into how genetic variants can guide successful treatments must become part of routine medical practice and records, says Geoffrey Ginsburg.

Long intergenic noncoding RNAs (lincRNAs) play diverse regulatory roles in human development and disease, but little is known about their evolutionary history and constraint. Here, we characterize human lincRNA expression patterns in nine tissues across six mammalian species and multiple individuals. Of the 1898 human lincRNAs expressed in these tissues, we find orthologous transcripts for 80% in chimpanzee, 63% in rhesus, 39% in cow, 38% in mouse, and 35% in rat. Mammalian-expressed lincRNAs show remarkably strong conservation of tissue specificity, suggesting that it is selectively maintained. In contrast, abundant splice-site turnover suggests that exact splice sites are not critical. Relative to evolutionarily young lincRNAs, mammalian-expressed lincRNAs show higher primary sequence conservation in their promoters and exons, increased proximity to protein-coding genes enriched for tissue-specific functions, fewer repeat elements, and more frequent single-exon transcripts. Remarkably, we find that ~20% of human lincRNAs are not expressed beyond chimpanzee and are undetectable even in rhesus. These hominid-specific lincRNAs are more tissue specific, enriched for testis, and faster evolving within the human lineage.

CAGE (cap analysis gene expression) and RNA-seq are two major technologies used to identify transcript abundances as well as structures. They measure expression by sequencing from either the 5' end of capped molecules (CAGE) or tags randomly distributed along the length of a transcript (RNA-seq). Library protocols for clonally amplified (Illumina, SOLiD, 454 Life Sciences [Roche], Ion Torrent), second-generation sequencing platforms typically employ PCR preamplification prior to clonal amplification, while third-generation, single-molecule sequencers can sequence unamplified libraries. Although these transcriptome profiling platforms have been demonstrated to be individually reproducible, no systematic comparison has been carried out between them. Here we compare CAGE, using both second- and third-generation sequencers, and RNA-seq, using a second-generation sequencer based on a panel of RNA mixtures from two human cell lines to examine power in the discrimination of biological states, detection of differentially expressed genes, linearity of measurements, and quantification reproducibility. We found that the quantified levels of gene expression are largely comparable across platforms and conclude that CAGE and RNA-seq are complementary technologies that can be used to improve incomplete gene models. We also found systematic bias in the second- and third-generation platforms, which is likely due to steps such as linker ligation, cleavage by restriction enzymes, and PCR amplification. This study provides a perspective on the performance of these platforms, which will be a baseline in the design of further experiments to tackle complex transcriptomes uncovered in a wide range of cell types.

by Margarete Boos, Johannes Pritz, Simon Lange, Michael Belz

How is movement of individuals coordinated as a group? This is a fundamental question of social behaviour, encompassing phenomena such as bird flocking, fish schooling, and the innumerable activities in human groups that require people to synchronise their actions. We have developed an experimental paradigm, the HoneyComb computer-based multi-client game, to empirically investigate human movement coordination and leadership. Using economic games as a model, we set monetary incentives to motivate players on a virtual playfield to reach goals via players' movements. We asked whether (I) humans coordinate their movements when information is limited to an individual group member's observation of adjacent group member motion, (II) whether an informed group minority can lead an uninformed group majority to the minority's goal, and if so, (III) how this minority exerts its influence. We showed that in a human group – on the basis of movement alone – a minority can successfully lead a majority. Minorities lead successfully when (a) their members choose similar initial steps towards their goal field and (b) they are among the first in the whole group to make a move. Using our approach, we empirically demonstrate that the rules of swarming behaviour apply to humans. Even complex human behaviour, such as leadership and directed group movement, follow simple rules that are based on visual perception of local movement.

Nature Biotechnology. doi:10.1038/nbt.2860

Authors: Thomas S Elliott, Fiona M Townsley, Ambra Bianco, Russell J Ernst, Amit Sachdeva, Simon J Elsässer, Lloyd Davis, Kathrin Lang, Rudolf Pisa, Sebastian Greiss, Kathryn S Lilley & Jason W Chin

Jeffrey M. Perkel

Blocking two crucial repair factors prevents DNA repair during mitosis, saving the cell from catastrophic chromosome fusions. Authors: Alexandre Orthwein, Amélie Fradet-Turcotte, Sylvie M. Noordermeer, Marella D. Canny, Catherine M. Brun, Jonathan Strecker, Cristina Escribano-Diaz, Daniel Durocher

Nature Structural & Molecular Biology 21, 423 (2014). doi:10.1038/nsmb.2799

Authors: Xinyi Lu, Friedrich Sachs, LeeAnn Ramsay, Pierre-Étienne Jacques, Jonathan Göke, Guillaume Bourque & Huck-Hui Ng

Human endogenous retrovirus subfamily H (HERVH) is a class of transposable elements expressed preferentially in human embryonic stem cells (hESCs). Here, we report that the long terminal repeats of HERVH function as enhancers and that HERVH is a nuclear long noncoding RNA required to maintain hESC identity. Furthermore, HERVH is associated with OCT4, coactivators and Mediator subunits. Together, these results uncover a new role of species-specific transposable elements in hESCs.

Nature Biotechnology 32, 341 (2014). doi:10.1038/nbt.2850

Authors: Nathan Boley, Marcus H Stoiber, Benjamin W Booth, Kenneth H Wan, Roger A Hoskins, Peter J Bickel, Susan E Celniker & James B Brown

Nature advance online publication 09 April 2014. doi:10.1038/nature13166

Authors: Yuexin Zhou, Shiyou Zhu, Changzu Cai, Pengfei Yuan, Chunmei Li, Yanyi Huang & Wensheng Wei

Targeted genome editing technologies are powerful tools for studying biology and disease, and have a broad range of research applications. In contrast to the rapid development of toolkits to manipulate individual genes, large-scale screening methods based on the complete loss of gene expression are only now beginning to be developed. Here we report the development of a focused CRISPR/Cas-based (clustered regularly interspaced short palindromic repeats/CRISPR-associated) lentiviral library in human cells and a method of gene identification based on functional screening and high-throughput sequencing analysis. Using knockout library screens, we successfully identified the host genes essential for the intoxication of cells by anthrax and diphtheria toxins, which were confirmed by functional validation. The broad application of this powerful genetic screening strategy will not only facilitate the rapid identification of genes important for bacterial toxicity but will also enable the discovery of genes that participate in other biological processes.

Nature Methods. doi:10.1038/nmeth.2910

Authors: Sandy Klemm, Stefan Semrau, Kay Wiebrands, Dylan Mooijman, Dina A Faddah, Rudolf Jaenisch & Alexander van Oudenaarden

We have developed a quantitative technique for sorting cells on the basis of endogenous RNA abundance, with a molecular resolution of 10–20 transcripts. We demonstrate efficient and unbiased RNA extraction from transcriptionally sorted cells and report a high-fidelity transcriptome measurement of mouse induced pluripotent stem cells (iPSCs) isolated from a heterogeneous reprogramming culture. This method is broadly applicable to profiling transcriptionally distinct cellular states without requiring antibodies or transgenic fluorescent proteins.

Nature Structural & Molecular Biology. doi:10.1038/nsmb.2799

Authors: Xinyi Lu, Friedrich Sachs, LeeAnn Ramsay, Pierre-Étienne Jacques, Jonathan Göke, Guillaume Bourque & Huck-Hui Ng

Human endogenous retrovirus subfamily H (HERVH) is a class of transposable elements expressed preferentially in human embryonic stem cells (hESCs). Here, we report that the long terminal repeats of HERVH function as enhancers and that HERVH is a nuclear long noncoding RNA required to maintain hESC identity. Furthermore, HERVH is associated with OCT4, coactivators and Mediator subunits. Together, these results uncover a new role of species-specific transposable elements in hESCs.

Nature Structural & Molecular Biology. doi:10.1038/nsmb.2801

Authors: Germano Cecere, Sebastian Hoersch, Sean O'Keeffe, Ravi Sachidanandam & Alla Grishok

Nature advance online publication 30 March 2014. doi:10.1038/nature13148

Authors: Joseph D. Mancias, Xiaoxu Wang, Steven P. Gygi, J. Wade Harper & Alec C. Kimmelman

Autophagy, the process by which proteins and organelles are sequestered in double-membrane structures called autophagosomes and delivered to lysosomes for degradation, is critical in diseases such as cancer and neurodegeneration. Much of our understanding of this process has emerged from analysis of bulk cytoplasmic autophagy, but our understanding of how specific cargo, including organelles, proteins or intracellular pathogens, are targeted for selective autophagy is limited. Here we use quantitative proteomics to identify a cohort of novel and known autophagosome-enriched proteins in human cells, including cargo receptors. Like known cargo receptors, nuclear receptor coactivator 4 (NCOA4) was highly enriched in autophagosomes, and associated with ATG8 proteins that recruit cargo–receptor complexes into autophagosomes. Unbiased identification of NCOA4-associated proteins revealed ferritin heavy and light chains, components of an iron-filled cage structure that protects cells from reactive iron species but is degraded via autophagy to release iron through an unknown mechanism. We found that delivery of ferritin to lysosomes required NCOA4, and an inability of NCOA4-deficient cells to degrade ferritin led to decreased bioavailable intracellular iron. This work identifies NCOA4 as a selective cargo receptor for autophagic turnover of ferritin (ferritinophagy), which is critical for iron homeostasis, and provides a resource for further dissection of autophagosomal cargo–receptor connectivity.

CAGE (cap analysis gene expression) and RNA-seq are two major technologies used to identify transcript abundances as well as structures. They measure expression by sequencing from either the 5' end of capped molecules (CAGE) or tags randomly distributed along the length of a transcript (RNA-seq). Library protocols for clonally amplified (Illumina, SOLiD, 454 Life Sciences [Roche], Ion Torrent), second-generation sequencing platforms typically employ PCR preamplification prior to clonal amplification, while third-generation, single-molecule sequencers can sequence unamplified libraries. Although these transcriptome profiling platforms have been demonstrated to be individually reproducible, no systematic comparison has been carried out between them. Here we compare CAGE, using both second- and third-generation sequencers, and RNA-seq, using a second-generation sequencer based on a panel of RNA mixtures from two human cell lines to examine power in the discrimination of biological states, detection of differentially expressed genes, linearity of measurements, and quantification reproducibility. We found that the quantified levels of gene expression are largely comparable across platforms and conclude that CAGE and RNA-seq are complementary technologies that can be used to improve incomplete gene models. We also found systematic bias in the second- and third-generation platforms, which is likely due to steps such as linker ligation, cleavage by restriction enzymes, and PCR amplification. This study provides a perspective on the performance of these platforms, which will be a baseline in the design of further experiments to tackle complex transcriptomes uncovered in a wide range of cell types.

Genomic researchers are increasingly faced with difficult decisions about whether, under what circumstances, and how to return research results and significant incidental findings to study participants. Many have argued that there is an ethical, maybe even a legal obligation to disclose significant findings under some circumstances. At the international level, over the last decade there has begun to emerge a clear legal obligation to return significant findings discovered during the course of research. However, there is no explicit legal duty to disclose in the United States. This creates legal uncertainty that may lead to unmanaged variation in practice and poor quality care. This paper discusses liability risks associated with the disclosure of significant research findings for investigators in the United States.