The role of introns in evolution

What are the roles of 'classical' introns in the evolution of nuclear genes, and what was the origin of these introns? Exon shuffling has been important in the evolution of cell surface and extracellular proteins, but the evidence for it in respect of intracellular proteins is weak. Intron distributions imply that some introns have been removed while others have been inserted in the course of evolution: ancestral patterns of introns may thus have been obscured. Recent evidence on the self-splicing and reverse-splicing abilities of Group II introns supports the hypothesis that these could have been the ancestors of classical introns.     

Unravelling the proteome of degenerative human mitral valves

Degenerative mitral valve disease (DMVD), which includes the syndromes of mitral valve prolapse (MVP) and flail leaflet, is a common valvular condition which can be complicated by mitral regurgitation and adverse cardiovascular outcomes. Although several genetic and other studies of MVP in dog models have provided some information regarding the underlying disease mechanisms, the proteins and molecular events mediating human MVP pathogenesis have not been unraveled. In this study, we report the first large‐scale proteome profiling of mitral valve tissue resected from patients with MVP. A total of 1134 proteins were identified, some of which were validated using SWATH‐MS and western blotting. GO annotation of these proteins confirmed the validity of this proteome database in various cardiovascular processes. Among the list of proteins, we found several structural and extracellular matrix proteins, such as asporin, biglycan, decorin, lumican, mimecan, prolargin, versican, and vinculin, that have putative roles in the pathophysiology of MVP. These proteins could also be involved in the cardiac remodeling associated with mitral regurgitation. All MS data have been deposited in the ProteomeXchange with identifier PXD000774 ( http://proteomecentral.proteomexchange.org/dataset/PXD000774 ).     

Mechanosensitive Closed-Closed Transitions in Large Membrane Proteins: Osmoprotection and Tension Damping

Multiconformation membrane proteins are mechanosensitive (MS) if their conformations displace different bilayer areas. Might MS closed-closed transitions serve as tension buffers, that is, as membrane “spandex”? While bilayer expansion is effectively instantaneous, transitions of bilayer-embedded MS proteins are stochastic (thermally activated) so spandex kinetics would be critical. Here we model generic two-state (contracted/expanded) stochastic spandexes inspired by known bacterial osmovalves (MscL, MscS) then suggest experimental approaches to test for spandex-like behaviors in these proteins. Modeling shows: 1), spandex kinetics depend on the transition state location along an area reaction coordinate; 2), increasing membrane concentration of a spandex right-shifts its midpoint (= tension-Boltzmann); 3), spandexes with midpoints below the activating tension of an osmovalve could optimize osmovalve deployment (required: large midpoint, barrier near the expanded state); 4), spandexes could damp bilayer tension excursions (required: midpoint at target tension, and for speed, barrier halfway between the contracted and expanded states; the larger the spandex Δ-area, the more precise the maintenance of target tension; higher spandex concentrations damp larger amplitude strain fluctuations). One spandex species could not excel as both first line of defense for osmovalve partners and tension damper. Possible interactions among MS closed-closed and closed-open transitions are discussed for MscS- and MscL-like proteins.     

MALDI mass spectrometry in prostate cancer biomarker discovery

Matrix-assisted laser desorption/ionisation (MALDI) mass spectrometry (MS) is a highly versatile and sensitive analytical technique, which is known for its soft ionisation of biomolecules such as peptides and proteins. Generally, MALDI MS analysis requires little sample preparation, and in some cases like MS profiling it can be automated through the use of robotic liquid-handling systems. For more than a decade now, MALDI MS has been extensively utilised in the search for biomarkers that could aid clinicians in diagnosis, prognosis, and treatment decision making. This review examines the various MALDI-based MS techniques like MS imaging, MS profiling and proteomics in-depth analysis where MALDI MS follows fractionation and separation methods such as gel electrophoresis, and how these have contributed to prostate cancer biomarker research. This article is part of a Special Issue entitled: Biomarkers: A Proteomic Challenge.     

Assessing the prospects for a return of organisms in evolutionary biology

An argument has been raised from various perspectives against the Modern Synthesis (MS) in the past two decades: it has forgotten organisms. Niche construction theorists (Odling-Smee et al. 2003), developmental biologists like West-Eberhard (2003) and Evo-Devo elaborated various views which concur on a rehabilitation of the explanatory role of organisms, formerly neglected by an evolutionary science mostly centered on genes. This paper aims at assessing such criticisms by unraveling the specific arguments they use and evaluating how empirical findings may support them. In the first section, I review the usual critiques about the way MS treats organisms and show that the organisms-concerned critique is multifaceted, and I use the controversy about units of selection in order to show that purely conceptual and empirical arguments have been mixed up when organisms were concerned. In the second section, I consider successively the challenges raised to evolutionary MS by structuralist biologists and then the developmentalist challenge mostly raised by Evo-Devo. I distinguish what is purely conceptual among those criticisms and what mostly relies on recent empirical findings about genome activation, inheritance, and epigenetics. The last section discusses another program in MS, namely "evolutionary transitions" research, as enquiry into the emergence of organisms.     

The Genetic Code: What Is It Good For? An Analysis of the Effects of Selection Pressures on Genetic Codes

How did the ``universal' genetic code arise? Several hypotheses have been put forward, and the code has been analyzed extensively by authors looking for clues to selection pressures that might have acted during its evolution. But this approach has been ineffective. Although an impressive number of properties has been attributed to the universal code, it has been impossible to determine whether selection on any of these properties was important in the code's evolution or whether the observed properties arose as a consequence of selection on some other characteristic. Therefore we turned the question around and asked, what would a genetic code look like if it had evolved in response to various different selection pressures? To address this question, we constructed a genetic algorithm. We found first that selecting on a particular measure yields codes that are similar to each other. Second, we found that the universal code is far from minimized with respect to the effects of mutations (or translation errors) on the amino acid compositions of proteins. Finally, we found that the codes that most closely resembled real codes were those generated by selecting on aspects of the code's structure, not those generated by selecting to minimize the effects of amino acid substitutions on proteins. This suggests that the universal genetic code has been selected for a particular structure—a structure that confers an important flexibility on the evolution of genes and proteins—and that the particular assignments of amino acids to codons are secondary. Received: 29 December 1998 / Accepted: 8 July 1999     

Epithelia as bubble rafts: A new method for analysis of cell shape and intercellular adhesion in embryonic and other epithelia

Cell-cell interactions may often be characterized by a work of adhesion or equivalent surface tension between cells. This intercellular surface tension may vary from one part of an epithelium to another. Such gradients in surface tension may be important driving factors in morphogenesis. A computational method for the measurement of parameters of cell shape involved in surface tension has been developed. It permits detailed observational analysis of cell-cell interface curvature. The information thus obtained is utilized in an algorithm for estimating gradients of surface tension in epithelia. The use of these methods should permit cell biologists and embryologists to decide whether or not developing cells behave like bubbles, and if so, to what extent their adhesive forces contribute to various aspects of embryogenesis.     

蛇毒神经毒素类似物的免疫学分析

选取眼镜蛇蛇毒和银环蛇蛇毒中 1 0个差异较大的神经毒素类似物 ,通过麦芽糖融合表达系统使它们在大肠杆菌中得到高效可溶性表达 .通过 Amylose- Sepharose6B亲和树脂纯化这些融合蛋白 .分别制备这 1 0种融合蛋白多克隆抗体 .同时将这 1 0种神经毒素类似物进行硫氧还蛋白融合表达 ,表达产物基本上都是包涵体 .以硫氧还蛋白融合表达产物为抗原和上述 1 0种神经毒素类似物抗血清进行 Western blotting.结果显示 ,大多数神经毒素类似物之间没有免疫交叉反应 ,表明这些神经毒素类似物的抗原性差别较大 ,可能存在不同的分类和进化     

Voltage-induced gating of the mechanosensitive MscL ion channel reconstituted in a tethered lipid bilayer membrane

The mechanosensitive (MS) ion channel is gated by changes in bilayer deformation. It is functional without the presence of any other proteins and gating of the channel has been successfully achieved using conventional patch clamping techniques where a voltage has been applied together with a pressure over the membrane. Here, we have for the first time analyzed the large conducting (MscL) channel in a supported membrane using only an external electrical field. This was made possible using a newly developed technique utilizing a tethered lipid bilayer membrane (tBLM), which is part of an engineered microelectronic array chip. Single ion channel activity characteristic for MscL was obtained, albeit with lower conductivity. The ion channel was gated using solely a transmembrane potential of 300 mV. Computations demonstrate that this amount of membrane potential induces a membrane tension of 12 dyn/cm, equivalent to that calculated to gate the channel in patch clamp from pressure-induced stretching of the bilayer. These results strengthen the supposition that the MscL ion channel gates in response to stress in the lipid membrane rather than pressure across it. Furthermore, these findings illustrate the possibility of using the MscL as a release valve for engineered membrane devices; one step closer to mimicking the true function of the living cell.     

Proteomic analysis of photosystem I components from different plant species

In this study, the photosystem I (PSI) highly hydrophobic proteins present within stroma lamellae of the thylakoid membrane were separated by RP-HPLC and identified either by in-solution trypsin digestion peptide fragment fingerprinting or by the close correspondence between the intact mass measurements (IMMs) and those expected from the DNA sequence. Protein identification performed by MS/MS was as reliable as IMMs. Thus, IMM is an easy and valid method for identifying proteins that have no PTMs. This paper reports the M(r) for all PSI proteins in ten different species, including those whose genes have not yet been cloned. Lhca5 was revealed unequivocally in four species, corroborating that it is indeed a protein belonging to the light-harvesting antenna of PSI. In all species examined, the product of the Lhca6 gene has never been revealed. Concerning core proteins, Psa-O has been revealed in three species; isoforms of Psa-D and Psa-E have been found in both monocots and dicots. Small proteins like Psa-I and Psa-J are well separated and identified. RP-HPLC produces reliable fingerprints and reveals that the relative amounts of PSI proteins appear to be markedly different.     

The evolution of the development

The evolution of development required few new features not already present in the eukaryotic cell, as exemplified by the cell cycle. Moreover, the protozoa possess many features of spatial organization and regulation present in metazoan embryos.
The earliest multicellular organism could have been reproduced by a stem cell mechanism or by fission, the latter requiring cell-to-cell interactions that may have favoured cell-interactions and regulation. Regeneration can be considered as a meta-phenomenon related to asexual reproduction and retention of embryonic characters. The origin of embryonic structures like the gastrula may be accounted for in terms of Haeckel's 'Gastrea' theory. Mechanisms based on selection at the level of cell lineage are rejected.
It is not clear what selective forces act on development itself, as distinct from the requirement for reliably producing a functional orgainsm. There is, for example, a major problem why gastrulation should be so variable in related animals. Selection for rate of development in relation to energy utilization may play a role. If many variants are neutral this may facilitate the evolution of novelty.
In general terms there is a requirement for a continuity principle for the evolution of each form in development. Most groups pass through a phylotypic stage with considerable diversity before and after.     

Mechanical Activation of MscL Revealed by a Locally Distributed Tension Molecular Dynamics Approach

Membrane tension perceived by mechanosensitive (MS) proteins mediates cellular responses to mechanical stimuli and osmotic stresses, and it also guides multiple biological functions including cardiovascular control and development. In bacteria, MS channels function as tension-activated pores limiting excessive turgor pressure, with MS channel of large conductance (MscL) acting as an emergency release valve preventing cell lysis. Previous attempts to simulate gating transitions in MscL by either directly applying steering forces to the protein or by increasing the whole-system tension were not fully successful and often disrupted the integrity of the system. We present a novel, to our knowledge, locally distributed tension molecular dynamics (LDT-MD) simulation method that allows application of forces continuously distributed among lipids surrounding the channel using a specially constructed collective variable. We report reproducible and reversible transitions of MscL to the open state with measured parameters of lateral expansion and conductivity that exactly satisfy experimental values. The LDT-MD method enables exploration of the MscL-gating process with different pulling velocities and variable tension asymmetry between the inner and outer membrane leaflets. We use LDT-MD in combination with well-tempered metadynamics to reconstruct the tension-dependent free-energy landscape for the opening transition in MscL. The flexible definition of the LDT collective variable allows general application of our method to study mechanical activation of any membrane-embedded protein.     

Protein evolution is faster outside the cell

Some proteins are highly conserved across all species, whereas others diverge significantly even between closely related species. Attempts have been made to correlate the rate of protein evolution to amino acid composition, protein dispensability, and the number of protein-protein interactions, but in all cases, conflicting studies have shown that the theories are hard to confirm experimentally. The only correlation that is undisputed so far is that highly/broadly expressed proteins seem to evolve at a lower rate. Consequently, it has been suggested that correlations between evolution rate and factors like protein dispensability or the number of protein-protein interactions could be just secondary effects due to differences in expression. The purpose of this study was to analyze mammalian proteins/genes with known subcellular location for variations in evolution rates. We show that proteins that are exported (extracellular proteins) evolve faster than proteins that reside inside the cell (intracellular proteins). We find weak, but significant, correlations between evolution rates and expression levels, percentage of tissues in which the proteins are expressed (expression broadness), and the number of protein interaction partners. More important, we show that the observed difference in evolution rate between extra- and intracellular proteins is largely independent of expression levels, expression broadness, and the number of protein-protein interactions. We also find that the difference is not caused by an overrepresentation of immunological proteins or disulfide bridge-containing proteins among the extracellular data set. We conclude that the subcellular location of a mammalian protein has a larger effect on its evolution rate than any of the other factors studied in this paper, including expression levels/patterns. We observe a difference in evolution rates between extracellular and intracellular proteins for a yeast data set as well and again show that it is completely independent of expression levels.     

Why are proteins so robust to site mutations?

There have been repeated observations that proteins are surprisingly robust to site mutations, enduring significant numbers of substitutions with little change in structure, stability, or function. These results are almost paradoxical in light of what is known about random heteropolymers and the sensitivity of their properties to seemingly trivial mutations. To address this discrepancy, the preservation of biological protein properties in the presence of mutation has been interpreted as indicating the independence of selective pressure on such properties. Such results also lead to the prediction that de novo protein design should be relatively easy, in contrast to what is observed. Here, we use a computational model with lattice proteins to demonstrate how this robustness can result from population dynamics during the evolutionary process. As a result, sequence plasticity may be a characteristic of evolutionarily derived proteins and not necessarily a property of designed proteins. This suggests that this robustness must be re-interpreted in evolutionary terms, and has consequences for our understanding of both in vivo and in vitro protein evolution.     

Conservation of Surfactant Protein A: Evidence for a Single Origin for Vertebrate Pulmonary Surfactant

Surface tension is reduced at the air–liquid interface in the lung by a mixture of lipids and proteins termed pulmonary surfactant. This study is the first to provide evidence for the presence of a surfactant-specific protein (Surfactant Protein A—SP-A) in the gas-holding structures of representatives of all the major vertebrate groups. Western blot analysis demonstrated cross-reactivity between an antihuman SP-A antibody and material lavaged from lungs or swimbladders of members from all vertebrate groups. Immunocytochemistry localized this SP-A–like protein to the air spaces of lungs from the actinopterygiian fish and lungfish. Northern blot analysis indicated that regions of the mouse SP-A cDNA sequence are complementary to lung mRNA from all species examined. The presence of an SP-A–like protein and SP-A mRNA in members of all the major vertebrate groups implies that the surfactant system had a single evolutionary origin in the vertebrates. Moreover, the evolution of the surfactant system must have been a prerequisite for the evolution of airbreathing. The presence of SP-A in the goldfish swimbladder demonstrates a role for the surfactant system in an organ that is no longer used for airbreathing. Received: 5 March 1997 / Accepted: 14 June 1997     

Evidence of amino acid diversity-enhancing selection within humans and among primates at the candidate sperm-receptor gene PKDREJ

Sperm-egg interaction is a crucial step in fertilization, yet the identity of most interacting sperm-egg proteins that mediate this process remains elusive. Rapid evolution of some fertilization proteins has been observed in a number of species, including evidence of positive selection in the evolution of components of the mammalian egg coat. The rapid evolution of the egg-coat proteins could strongly select for changes on the sperm receptor, to maintain the interaction. Here, we present evidence that positive selection has driven the evolution of PKDREJ, a candidate sperm receptor of mammalian egg-coat proteins. We sequenced PKDREJ from a panel of 14 primates, including humans, and conducted a comparative maximum-likelihood analysis of nucleotide changes and found evidence of positive selection. An additional panel of 48 humans was surveyed for nucleotide polymorphisms at the PKDREJ locus. The regions predicted to have been subject to adaptive evolution among primates show several amino acid polymorphisms within humans. The distribution of polymorphisms suggests that balancing selection may maintain diverse PKDREJ alleles in some populations. It remains unknown whether there are functional differences associated with these diverse alleles, but their existence could have consequences for human fertility.     

14-3-3 proteins in neurodegeneration

Among the first reported functions of 14-3-3 proteins was the regulation of tyrosine hydroxylase (TH) activity suggesting a possible involvement of 14-3-3 proteins in Parkinson's disease. Since then the relevance of 14-3-3 proteins in the pathogenesis of chronic as well as acute neurodegenerative diseases, including Alzheimer's disease, polyglutamine diseases, amyotrophic lateral sclerosis and stroke has been recognized. The reported function of 14-3-3 proteins in this context are as diverse as the mechanism involved in neurodegeneration, reaching from basal cellular processes like apoptosis, over involvement in features common to many neurodegenerative diseases, like protein stabilization and aggregation, to very specific processes responsible for the selective vulnerability of cellular populations in single neurodegenerative diseases.Here, we review what is currently known of the function of 14-3-3 proteins in nervous tissue focussing on the properties of 14-3-3 proteins important in neurodegenerative disease pathogenesis.