Intermolecular Interactions Drive Protein Adaptive and Coadaptive Evolution at Both Species and Population Levels

Proteins are the building blocks for almost all the functions in cells. Understanding the molecular evolution of proteins and the forces that shape protein evolution is essential in understanding the basis of function and evolution. Previous studies have shown that adaptation frequently occurs at the protein surface, such as in genes involved in host–pathogen interactions. However, it remains unclear whether adaptive sites are distributed randomly or at regions associated with particular structural or functional characteristics across the genome, since many proteins lack structural or functional annotations. Here, we seek to tackle this question by combining large-scale bioinformatic prediction, structural analysis, phylogenetic inference, and population genomic analysis of Drosophila protein-coding genes. We found that protein sequence adaptation is more relevant to function-related rather than structure-related properties. Interestingly, intermolecular interactions contribute significantly to protein adaptation. We further showed that intermolecular interactions, such as physical interactions, may play a role in the coadaptation of fast-adaptive proteins. We found that strongly differentiated amino acids across geographic regions in protein-coding genes are mostly adaptive, which may contribute to the long-term adaptive evolution. This strongly indicates that a number of adaptive sites tend to be repeatedly mutated and selected throughout evolution in the past, present, and maybe future. Our results highlight the important roles of intermolecular interactions and coadaptation in the adaptive evolution of proteins both at the species and population levels.     

Modeling the age distribution of gene duplications in vertebrate genome using mixture density

Gene duplication is a fundamental source of genetic novelty in vertebrate evolution. In this study, we hypothesized that both continuous small-scale and discrete large-scale duplication play crucial roles in vertebrate genome. On the basis of the hypothesis, we developed mixture density to model the age distribution of gene duplications. The results of formal statistical inference suggest that the contribution of both duplication modes can be confirmed by the model, and one or two successive rounds of large-scale duplication are placed at the early origin of vertebrates. The half life of a duplicate becomes much longer in the long run than in the short run, which implies its functional evolution from redundancy to conservation. In addition, the model reveals disparate impact of the duplication modes, which appears to be correlated with macroevolution.     

Assessing functional redundancy in chronically trawled benthic communities

Fishing disturbance on ecosystems leads to changes in community structure and composition, which may have drastic implications for ecosystem functional performance. Functional redundancy, defined as species sharing similar functional roles, is a community property that plays an important role in preventing functional changes in ecosystems under pressure. In this study, we suggest that functional redundancy may be achieved through trait abundance (i.e. large amounts of a trait, hereafter “common traits”), or through trait richness (i.e. large numbers of distinct taxa exhibiting the same trait, hereafter “widespread traits”). We assessed the variability of both measures obtained from epifaunal and infaunal communities in soft-bottom trawling grounds. Sampling sites were located in four Mediterranean areas that were subjected to different levels of trawling effort. Common and widespread traits measures were based on the analysis of biological traits linked to key soft-bottoms functions such as nutrient cycling, bentho-pelagic coupling and habitat provision. The role of rare species in both measures was also assessed and we observed that, in our study sites, rare species generally exhibited the same traits as the most abundant species. Common and widespread traits measures provided complementary information on benthic functional redundancy. Thus, we suggest that a combination of the two measures should be used to appropriately assess benthic functional redundancy in trawling grounds. As redundancy is a component of ecosystem resilience, functional redundancy evaluation is important to assess the overall integrity of ecosystems.     

Context characterization of amino acid homorepeats using evolution,position, and order

Amino acid repeats, or homorepeats, are low complexity protein motifs consisting of tandem repetitions of a single amino acid. Their presence and relative number vary in different proteomes, and some studies have tried to address this variation, proteome by proteome. In this work, we present a full characterization of amino acid homorepeats across evolution. We studied the presence and differential usage of each possible homorepeat in proteomes from various taxonomic groups, using clusters of very similar proteins to eliminate redundancy. The position of each amino acid repeat within proteins, and the order of co‐occurring amino acid repeats were also addressed. As a result, we present evidence about the unevenly evolution of homorepeats, as well as the functional implications of their relative position in proteins. We discuss some of these cases in their taxonomic context. Collectively, our results show evolutionary and positional signals that suggest that homorepeats have biological function, likely creating unspecific protein interactions or modulating specific interactions in a context dependent manner. In conclusion, our work supports the functional importance of homorepeats and establishes a basis for the study of other low complexity repeats. Proteins 2017; 85:709–719. © 2016 Wiley Periodicals, Inc.     

Blocking the large extracellular loop (LEL) domain of FcTetraspanin-3 could inhibit the infection of white spot syndrome virus (WSSV) in Chinese shrimp, Fenneropenaeus chinensis

Tetraspanins belong to the transmembrane 4 superfamily (TM(4)SF), which span the cell membrane 4 times and act as bridges or connectors. Increasing evidences have shown that tetraspanins play important role in virus infection. The large extracellular loop (LEL) of a tetraspanin is considered as a possible target of some virus. Tetraspanins are widely found in invertebrates, but the functional roles of most invertebrate tetraspanins have remained unknown. Recently, a tetraspanin, called FcTetraspanin-3, was cloned from the cDNA library of Chinese shrimp, Fenneropenaeus chinensis. The FcTetraspanin-3 constitutive expression in all examined tissues and the expression of the gene were highly induced in hepatopancreas, lymphoid organ and intestine by white spot syndrome virus (WSSV) challenge. In this study, we expressed and purified the recombinant peptide containing the LEL domain of FcTetraspanin-3, and produced the anti-LEL polyclone antibody. The expression of FcTetraspanin-3 was observed by real-time PCR and Western blot. Also, the localization of FcTetraspanin-3-positive cells in intestine and hepatopancreas were revealed by immunofluorescence. The results of anti-LEL antibody blocking experiments shown that the antibody can significantly reduce the mortality of shrimp challenged by WSSV. Additionally, dsRNA interference was utilized to examine the functional role of FcTetraspanin-3 in response to WSSV infection, and a sensible decrease of the viral copy number in the tetraspanin knockdown shrimp. These results suggested the blocking of LEL domain of FcTetraspanin-3 could inhibit the infection of WSSV. FcTetraspanin-3 might play an important role in response to WSSV infection, and the LEL domain of FcTetraspanin-3 might mediate the entry of WSSV.     

Unequal genetic redundancies in Arabidopsis--a neglected phenomenon?

Genetic redundancy is a common phenomenon in Arabidopsis and is thought to be responsible for the absence of phenotypes in the majority of single loss-of-function mutants. In this review, we highlight an increasing number of examples in which redundancy between homologous genes is limited or absent despite functional equivalence of the respective proteins. In particular, we focus on cases of unequal redundancy, where the absence of a mutant phenotype in loss-of-function mutants of one gene contrasts with a strong phenotype in mutants of its homolog. In the double mutants, this phenotype is strongly enhanced. Possible explanations for such scenarios are discussed. We propose that the study of unequally redundant gene pairs offers a unique opportunity to understand global patterns of functional genome evolution.     

Neutral biogeography of phylogenetically structured interaction networks

Little is known about how biogeographic processes affect the dynamics of species interactions in space and time, although it is widely accepted that they drive community assemblage. In functional interactions, such as pollination and seed dispersal, species that share common ancestry tend to retain a common number of interactions and interact with similar sets of species, a pattern more commonly observed for animals than plants. On the one hand, the most coherent explanation for the phylogenetic structure of pollination and seed dispersal networks is that species retain ecological traits over evolution, which would cause the conservation of interaction partners. On the other hand, fundamental processes of biodiversity, such as dispersal and evolutionary rates seem to have important roles shaping the observed phylogenetic structure of mutualistic networks, but no model has been created to study the effect of these processes in the phylogenetic structure of mutualistic interactions. Here, we developed a stochastic simulation model to study the evolution of two interacting groups of species, which evolve independently over the same geographical domain. In our model, individuals of the same interaction group share ecological traits, whereas individuals of different trophic groups are ecologically distinct. We show that even in the absence of ecological differences between individuals, and disregarding any conservation of phenotypical and phenological traits between species, the interplay of dispersal and speciation is still a major driver of complex phylogenetic structure of functional interactions, such as pollination and seed dispersal.     

Regulation of clathrin coat assembly by Eps15 homology domain-mediated interactions during endocytosis

Clathrin-mediated endocytosis involves a coordinated series of molecular events regulated by interactions among a variety of proteins and lipids through specific domains. One such domain is the Eps15 homology (EH) domain, a highly conserved protein-protein interaction domain present in a number of proteins distributed from yeast to mammals. Several lines of evidence suggest that the yeast EH domain-containing proteins Pan1p, End3p, and Ede1p play important roles during endocytosis. Although genetic and cell-biological studies of these proteins suggested a role for the EH domains in clathrin-mediated endocytosis, it was unclear how they regulate clathrin coat assembly. To explore the role of the EH domain in yeast endocytosis, we mutated those of Pan1p, End3p, or Ede1p, respectively, and examined the effects of single, double, or triple mutation on clathrin coat assembly. We found that mutations of the EH domain caused a defect of cargo internalization and a delay of clathrin coat assembly but had no effect on assembly of the actin patch. We also demonstrated functional redundancy among the EH domains of Pan1p, End3p, and Ede1p for endocytosis. Of interest, the dynamics of several endocytic proteins were differentially affected by various EH domain mutations, suggesting functional diversity of each EH domain.     

A gene for speed? The evolution and function of α‐actinin‐3

The alpha-actinins are an ancient family of actin-binding proteins that play structural and regulatory roles in cytoskeletal organisation and muscle contraction. alpha-actinin-3 is the most-highly specialised of the four mammalian alpha-actinins, with its expression restricted largely to fast glycolytic fibres in skeletal muscle. Intriguingly, a significant proportion ( approximately 18%) of the human population is totally deficient in alpha-actinin-3 due to homozygosity for a premature stop codon polymorphism (R577X) in the ACTN3 gene. Recent work in our laboratory has revealed a strong association between R577X genotype and performance in a variety of athletic endeavours. We are currently exploring the function and evolutionary history of the ACTN3 gene and other alpha-actinin family members. The alpha-actinin family provides a fascinating case study in molecular evolution, illustrating phenomena such as functional redundancy in duplicate genes, the evolution of protein function, and the action of natural selection during recent human evolution.     

Individual-based model highlights the importance of trade-offs for virus-host population dynamics and long-term co-existence

Viruses play diverse and important roles in ecosystems. In recent years, trade-offs between host and virus traits have gained increasing attention in viral ecology and evolution. However, microbial organism traits, and viral population parameters in particular, are challenging to monitor. Mathematical and individual-based models are useful tools for predicting virus-host dynamics. We have developed an individual-based evolutionary model to study ecological interactions and evolution between bacteria and viruses, with emphasis on the impacts of trade-offs between competitive and defensive host traits on bacteria-phage population dynamics and trait diversification. Host dynamics are validated with lab results for different initial virus to host ratios (VHR). We show that trade-off based, as opposed to random bacteria-virus interactions, result in biologically plausible evolutionary outcomes, thus highlighting the importance of trade-offs in shaping biodiversity. The effects of nutrient concentration and other environmental and organismal parameters on the virus-host dynamics are also investigated. Despite its simplicity, our model serves as a powerful tool to study bacteria-phage interactions and mechanisms for evolutionary diversification under various environmental conditions.     

Tetraspanins as regulators of protein trafficking

Small transmembrane proteins of the tetraspanin superfamily are believed to function as the main structural blocks of specialized membrane microdomains (referred to as tetraspanin-enriched microdomains, TERM or TEM). Through a multitude of homotypic and heterotypic interactions, tetraspanins regulate lateral clustering and, consequently, signalling involving adhesion and growth factor receptors as well as costimulatory proteins. The presence of major histocompatibility complex (MHC) I and MHCII molecules in TERM led to suggestion of tetraspanins' involvement in antigen presentation. In addition, certain tetraspanins function as viral co-receptors and may be important for viral egress from infected cells. It has recently become apparent that in addition to their purely structural function as organizers of TERM, tetraspanins also regulate various aspects of trafficking and biosynthetic processing of associated receptors. Here, we review recent studies, which specifically focus on this issue.     

Functional Roles of Centridini Oil Bees and Malpighiaceae Oil Flowers in Biome‐wide Pollination Networks

Interactions between oil‐collecting bees and oil‐producing flowers are a very specialized mutualism, whose natural history is well known at the organism and population levels. In this study, we assessed these interactions at the biome level with a network approach, and hypothesized that widespread bee and plant species would occupy different ecological functional roles (Eltonian niches) in different biomes. Furthermore, we expected the most important functional roles in each network to be occupied more frequently by Byrsonima oil flowers and Centris oil bees, which share the longest coevolutionary history in the Neotropics. By compiling data from 40 articles on oil flower interactions within the Malpighiaceae family, we built six networks for different Brazilian biomes. We assessed the ecological functional role of each species in pollination networks of oil flowers through the metric known as ‘network functional role’. Although 90 percent of the species occupied peripheral roles in each network, some were found to occupy highly central roles. Oil flowers of the genera Byrsonima and Banisteriopsis and oil bees of the genera Centris and Epicharis were the most important species in all networks, as they made a disproportionally high number of interactions (hubs), or helped bind together different modules (connectors). Our findings suggest that functional roles vary geographically and seem to be affected by local conditions in different biomes. Furthermore, coevolutionary history seems to play an important role in determining functional roles in oil flower networks, although other factors are probably also important, especially the degree of specialization in this kind of interaction.     

Microbial biofilms in nature: unlocking their potential for agricultural applications

Soil environments are dynamic and the plant rhizosphere harbours a phenomenal diversity of micro-organisms which exchange signals and beneficial nutrients. Bipartite beneficial or symbiotic interactions with host roots, such as mycorrhizae and various bacteria, are relatively well characterized. In addition, a tripartite interaction also exists between plant roots, arbuscular mycorrhizal fungi (AMF) and associated bacteria. Bacterial biofilms exist as a sheet of bacterial cells in association with AMF structures, embedded within a self-produced exopolysaccharide matrix. Such biofilms may play important functional roles within these tripartite interactions. However, the details about such interactions in the rhizosphere and their relevant functional relationships have not been elucidated. This review explores the current understanding of naturally occurring microbial biofilms, and their interaction with biotic surfaces, especially AMF. The possible roles played by bacterial biofilms and the potential for their application for a more productive and sustainable agriculture is discussed in this review.     

Tetraspanins regulate the protrusive activities of cell membrane

Tetraspanins have gained increased attention due to their functional versatility. But the universal cellular mechanism that governs such versatility remains unknown. Herein we present the evidence that tetraspanins CD81 and CD82 regulate the formation and/or development of cell membrane protrusions. We analyzed the ultrastructure of the cells in which a tetraspanin is either overexpressed or ablated using transmission electron microscopy. The numbers of microvilli on the cell surface were counted, and the radii of microvillar tips and the lengths of microvilli were measured. We found that tetraspanin CD81 promotes the microvillus formation and/or extension while tetraspanin CD82 inhibits these events. In addition, CD81 enhances the outward bending of the plasma membrane while CD82 inhibits it. We also found that CD81 and CD82 proteins are localized at microvilli using immunofluorescence. CD82 regulates microvillus morphogenesis likely by altering the plasma membrane curvature and/or the cortical actin cytoskeletal organization. We predict that membrane protrusions embody a common morphological phenotype and cellular mechanism for, at least some if not all, tetraspanins. The differential effects of tetraspanins on microvilli likely lead to the functional diversification of tetraspanins and appear to correlate with their functional propensity.     

Evolution of Specificity in Protein-Protein Interactions

Hub proteins are proteins that maintain promiscuous molecular recognition. Because they are reported to play essential roles in cellular control, there has been a special interest in the study of their structural and functional properties, yet the mechanisms by which they evolve to maintain functional interactions are poorly understood. By combining biophysical simulations of coarse-grained proteins and analysis of proteins-complex crystallographic structures, we seek to elucidate those mechanisms. We focus on two types of hub proteins: Multi hubs, which interact with their partners through different interfaces, and Singlish hubs, which do so through a single interface. We show that loss of structural stability is required for the evolution of protein-protein-interaction (PPI) networks, and it is more profound in Singlish hub systems. In addition, different ratios of hydrophobic to electrostatic interfacial amino acids are shown to support distinct network topologies (i.e., Singlish and Multi systems), and therefore underlie a fundamental design principle of PPI in a crowded environment. We argue that the physical nature of hydrophobic and electrostatic interactions, in particular, their favoring of either same-type interactions (hydrophobic-hydrophobic), or opposite-type interactions (negatively-positively charged) plays a key role in maintaining the network topology while allowing the protein amino acid sequence to evolve.     

Evolution of Specificity in Protein-Protein Interactions

Hub proteins are proteins that maintain promiscuous molecular recognition. Because they are reported to play essential roles in cellular control, there has been a special interest in the study of their structural and functional properties, yet the mechanisms by which they evolve to maintain functional interactions are poorly understood. By combining biophysical simulations of coarse-grained proteins and analysis of proteins-complex crystallographic structures, we seek to elucidate those mechanisms. We focus on two types of hub proteins: Multi hubs, which interact with their partners through different interfaces, and Singlish hubs, which do so through a single interface. We show that loss of structural stability is required for the evolution of protein-protein-interaction (PPI) networks, and it is more profound in Singlish hub systems. In addition, different ratios of hydrophobic to electrostatic interfacial amino acids are shown to support distinct network topologies (i.e., Singlish and Multi systems), and therefore underlie a fundamental design principle of PPI in a crowded environment. We argue that the physical nature of hydrophobic and electrostatic interactions, in particular, their favoring of either same-type interactions (hydrophobic-hydrophobic), or opposite-type interactions (negatively-positively charged) plays a key role in maintaining the network topology while allowing the protein amino acid sequence to evolve.     

Investigation of protein-RNA interactions by mass spectrometry--Techniques and applications

Protein-RNA complexes play many important roles in diverse cellular functions. They are involved in a wide variety of different processes in growth and differentiation at the various stages of the cell cycle. As their function and catalytic activity are directly coupled to the structural arrangement of their components--proteins and ribonucleic acids--the investigation of protein-RNA interactions is of great functional and structural importance. Here we discuss the most prominent examples of protein-RNA complexes and describe some frequently used purification strategies. We present various techniques and applications of mass spectrometry to study protein-RNA complexes. We discuss the analysis of intact complexes as well as proteomics-based and crosslinking-based approaches in which proteins are cleaved into smaller peptides. This article is part of a Special Section entitled: Understanding genome regulation and genetic diversity by mass spectrometry.