Lessons (not) learned from mistakes about translation

Some popular ideas about translational regulation in eukaryotes have been recognized recently as mistakes. One example is the rejection of a long-standing idea about involvement of S6 kinase in translation of ribosomal proteins. Unfortunately, new proposals about how S6 kinase might regulate translation are based on evidence that is no better than the old. Recent findings have also forced rejection of some popular ideas about the function of sequences at the 3' end of viral mRNAs and rejection of some ideas about internal ribosome entry sequences (IRESs). One long-held belief was that tissue-specific translation via an IRES underlies the neurotropism of poliovirus and the attenuation of Sabin vaccine strains. Older experiments that appeared to support this belief and recent experiments that refute it are discussed. The hypothesis that dyskeratosis congenita is caused by a defect in IRES-mediated translation is probably another mistaken idea. The supporting evidence, such as it is, comes from a mouse model of the disease and is contradicted by studies carried out with cells from affected patients. The growing use of IRESs as tools to study other questions about translation is discussed and lamented. The inefficient function of IRESs (if they are IRESs) promotes misunderstandings. I explain again why it is not valid to invoke a special mechanism of initiation based on the finding that edeine (at very low concentrations) does not inhibit the translation of a putative IRES from cricket paralysis virus. I explain why new assays, devised to rule out splicing in tests with dicistronic vectors, are not valid and why experiments with IRESs are not a good way to investigate the mechanism whereby microRNAs inhibit translation.     

Protein name tagging guidelines: lessons learned

Interest in information extraction from the biomedical literature is motivated by the need to speed up the creation of structured databases representing the latest scientific knowledge about specific objects, such as proteins and genes. This paper addresses the issue of a lack of standard definition of the problem of protein name tagging. We describe the lessons learned in developing a set of guidelines and present the first set of inter-coder results, viewed as an upper bound on system performance. Problems coders face include: (a) the ambiguity of names that can refer to either genes or proteins; (b) the difficulty of getting the exact extents of long protein names; and (c) the complexity of the guidelines. These problems have been addressed in two ways: (a) defining the tagging targets as protein named entities used in the literature to describe proteins or protein-associated or -related objects, such as domains, pathways, expression or genes, and (b) using two types of tags, protein tags and long-form tags, with the latter being used to optionally extend the boundaries of the protein tag when the name boundary is difficult to determine. Inter-coder consistency across three annotators on protein tags on 300 MEDLINE abstracts is 0.868 F-measure. The guidelines and annotated datasets, along with automatic tools, are available for research use.     

Mistaken Identifiers: Gene name errors can be introduced inadvertently when using Excel in bioinformatics

Background  

When processing microarray data sets, we recently noticed that some gene names were being changed inadvertently to non-gene names.     

PKA: Lessons learned after twenty years

The first protein kinase structure, solved in 1991, revealed the fold that is shared by all members of the eukaryotic protein kinase superfamily and showed how the conserved sequence motifs cluster mostly around the active site. This structure of the PKA catalytic (C) subunit showed also how a single phosphate integrated the entire molecule. Since then the EPKs have become a major drug target, second only to the G-protein coupled receptors. Although PKA provided a mechanistic understanding of catalysis that continues to serve as a prototype for the family, by comparing many active and inactive kinases we subsequently discovered a hydrophobic spine architecture that is a characteristic feature of all active kinases. The ways in which the regulatory spine is dynamically assembled is the defining feature of each protein kinase. Protein kinases have thus evolved to be molecular switches, like the G-proteins, and unlike metabolic enzymes which have evolved to be efficient catalysis. PKA also shows how the dynamic tails surround the core and serve as essential regulatory elements. The phosphorylation sites in PKA, introduced both co- and post-translationally, are very stable. The resulting C-subunit is then packaged as an inhibited holoenzyme with cAMP-binding regulatory (R) subunits so that PKA activity is regulated exclusively by cAMP, not by the dynamic turnover of an activation loop phosphate. We could not understand activation and inhibition without seeing structures of R:C complexes; however, to appreciate the structural uniqueness of each R₂:C₂ holoenzyme required solving structures of tetrameric holoenzymes. It is these tetrameric holoenzymes that are localized to discrete sites in the cell, typically by A Kinase Anchoring Proteins where they create discrete foci for PKA signaling. Understanding these dynamic macromolecular complexes is the challenge that we now face. This article is part of a Special Issue entitled: Inhibitors of Protein Kinases (2012).     

Biofuel technologies: Lessons learned and pathways to decarbonization

This Opinion highlights several successful cases of biofuel technologies recently described by the IEA Bioenergy Intertask Report on Lessons Learned. The report discussed the potential of biofuels to contribute to a significant market supply, thus replacing fossil fuels and mitigating global warming, and it underscores the challenges in expanding biofuel production and replicating successful models between countries and regions. Based on the lessons learned from conventional, established technologies, the authors analyzed policies, feedstocks, products, technologies, economics, environmental concerns, social aspects, scalability, and ease of implementation and replication in different countries or regions. There are blending mandates in place around the world to foster the use of biofuels. Dependence on the availability and price fluctuations of crop feedstocks may limit biofuel production in certain circumstances. Legal restrictions on using food crops as feedstocks present obstacles to scaling up production. Temporary constraints related to feedstock costs and availability, as evidenced by changes and postponements of biofuel blending mandates in various countries (particularly during the COVID-19 pandemic) also pose challenges. Technological hurdles exist for advanced biofuels that implicate premium pricing. Still, 2G ethanol from sugarcane meets very strict feedstock requirements with a carbon footprint so low that only electric vehicles charged in Norway could have life-cycle GHG emissions at the same level as a 2G ethanol-fueled combustion engine car. The authors evaluate whether and how much electrification could contribute to advance the decarbonization efforts in different countries. Drawing from these observations, the authors express their viewpoints to assist researchers and policymakers in the energy sector in formulating viable approaches to combat the climate crisis.     

Lessons learned from gene transfer approaches

Recent technological advances allow the transfer of genes to the synovial lining of joints. As well as opening novel opportunities for therapy, these techniques provide valuable new tools for the study of synovitis and other aspects of the biology of joints in health and disease. This article reviews briefly the results of experiments in which selected genes have been transferred to the knee joints of healthy rabbits and rabbits with antigen-induced arthritis.     

Lessons learned from the dog genome

Extensive genetic resources and a high-quality genome sequence position the dog as an important model species for understanding genome evolution, population genetics and genes underlying complex phenotypic traits. Newly developed genomic resources have expanded our understanding of canine evolutionary history and dog origins. Domestication involved genetic contributions from multiple populations of gray wolves probably through backcrossing. More recently, the advent of controlled breeding practices has segregated genetic variability into distinct dog breeds that possess specific phenotypic traits. Consequently, genome-wide association and selective sweep scans now allow the discovery of genes underlying breed-specific characteristics. The dog is finally emerging as a novel resource for studying the genetic basis of complex traits, including behavior.     

Lessons learned from protein aggregation: toward technological and biomedical applications

The close relationship between protein aggregation and neurodegenerative diseases has been the driving force behind the renewed interest in a field where biophysics, neurobiology and nanotechnology converge in the study of the aggregate state. On one hand, knowledge of the molecular principles that govern the processes of protein aggregation has a direct impact on the design of new drugs for high-incidence pathologies that currently can only be treated palliatively. On the other hand, exploiting the benefits of protein aggregation in the design of new nanomaterials could have a strong impact on biotechnology. Here we review the contributions of our research group on novel neuroprotective strategies developed using a purely biophysical approach. First, we examine how doxycycline, a well-known and innocuous antibiotic, can reshape α-synuclein oligomers into non-toxic high-molecular-weight species with decreased ability to destabilize biological membranes, affect cell viability and form additional toxic species. This mechanism can be exploited to diminish the toxicity of α-synuclein oligomers in Parkinson’s disease. Second, we discuss a novel function in proteostasis for extracellular glyceraldehyde 3-phosphate dehydrogenase (GAPDH) in combination with a specific glycosaminoglycan (GAG) present in the extracellular matrix. GAPDH, by changing its quaternary structure from a tetramer to protofibrillar assembly, can kidnap toxic species of α-synuclein, and thereby interfere with the spreading of the disease. Finally, we review a brighter side of protein aggregation, that of exploiting the physicochemical advantages of amyloid aggregates as nanomaterials. For this, we designed a new generation of insoluble biocatalysts based on the binding of photo-immobilized enzymes onto hybrid protein:GAG amyloid nanofibrils. These new nanomaterials can be easily functionalized by attaching different enzymes through dityrosine covalent bonds.     

Lessons learned from the syndrome of oculopalatal tremor

Journal of Computational Neuroscience - The syndrome of oculopalatal tremor (OPT) featuring the olivo-cerebellar hypersychrony leads to disabling pendular nystagmus and palatal myoclonus. This rare...     

Lessons learned from nuclear transfer (cloning)

Somatic cell nuclear transfer (SCNT) has been accomplished in an ever-growing list of species. In each case, an enucleated oocyte has successfully reset the nucleus of a somatic cell such that the embryonic program could progress to the production of a live offspring. The overall efficiency of the process remains low due to a combination of biological and technical challenges, some of which are known and others remain to be elucidated. Comparative studies between livestock and laboratory species may help improve not only nuclear transfer efficiencies but also uncover basic underlying developmental principles.     

Lessons learned from type III effector transgenic plants

The Gram negative bacterial phytopathogen Pseudomonas syringae employs a molecular syringe termed the type III secretion system (TTSS) to deliver an array of type III secreted effector (TTSE) proteins into plant cells. The major function ascribed to type III effectors of P. syringae is their ability to suppress plant immunity. Because individual pathovars of P. syringae can possess over 30 TTSEs, functional redundancy can provide a hurdle to ascribing functions by TTSE-deletion or -overexpression in such TTSE-rich backgrounds. Approaches to overcome functional redundancy have included the deletion of multiple TTSEs from individual pathovars as well as engineering the plant commensal P. fluorescens strain to express the P. syringae TTSS and deliver P. syringae TTSEs. As we describe here, transgenic Arabidopsis plants expressing individual TTSEs have also been used to overcome problems of functional redundancy and provide invaluable insights into TTSE virulence functions.Key words: pathogen, virulence, effector, plant immunity, HopF2_Pto, RIN4