The native subcellular location (also referred to as localization or cellular compartment) of a protein is the one in which it acts most frequently; it is one aspect of protein function. Do ten eukaryotic model organisms differ in their location spectrum, i.e., the fraction of its proteome in each of seven major cellular compartments? As experimental annotations of locations remain biased and incomplete, we need prediction methods to answer this question. After systematic bias corrections, the complete but faulty prediction methods appeared to be more appropriate to compare location spectra between species than the incomplete more accurate experimental data. This work compared the location spectra for ten eukaryotes: Homo sapiens (human), Gorilla gorilla (gorilla), Pan troglodytes (chimpanzee), Mus musculus (mouse), Rattus norvegicus (rat), Drosophila melanogaster (fruit/vinegar fly), Anopheles gambiae (African malaria mosquito), Caenorhabitis elegans (nematode), Saccharomyces cerevisiae (baker’s yeast), and Schizosaccharomyces pombe (fission yeast). The two largest classes were predicted to be the nucleus and the cytoplasm together accounting for 47–62% of all proteins, while 7–21% of the proteins were predicted in the plasma membrane and 4–15% to be secreted. Overall, the predicted location spectra were largely similar. However, in detail, the differences sufficed to plot trees (UPGMA) and 2D (PCA) maps relating the ten organisms using a simple Euclidean distance in seven states (location classes). The relations based on the simple predicted location spectra captured aspects of cross-species comparisons usually revealed only by much more detailed evolutionary comparisons. Most interestingly, known phylogenetic relations were reproduced better by paralog-only than by ortholog-only trees.

Preventing and controlling epidemics caused by vector-borne viruses are particularly challenging due to their diverse pool of hosts and highly adaptive nature. Many vector-borne viruses belong to the Flavivirus genus, whose members vary greatly in host range and specificity. Members of the Flavivirus genus can be categorized to four main groups: insect-specific viruses that are maintained solely in arthropod populations, mosquito-borne viruses and tick-borne viruses that are transmitted to vertebrate hosts by mosquitoes or ticks via blood feeding, and those with no-known vector. The mosquito-borne group encompasses the yellow fever, dengue, and West Nile viruses, all of which are globally spread and cause severe morbidity in humans. The Flavivirus genus is genetically diverse, and its members are subject to different host-specific and vector-specific selective constraints, which do not always align. Thus, understanding the underlying genetic differences that led to the diversity in host range within this genus is an important aspect in deciphering the mechanisms that drive host compatibility and can aid in the constant arms-race against viral threats. Here, we review the phylogenetic relationships between members of the genus, their infection bottlenecks, and phenotypic and genomic differences. We further discuss methods that utilize these differences for prediction of host shifts inflaviviruses and can contribute to viral surveillance efforts.

We use large-scale mutagenesis data and computer simulations to quantify the mutational robustness of protein-coding genes by taking into account constraints arising from protein function and the genetic code. Analyses of the distribution of amino acid substitutions from 18 mutagenesis studies revealed an average of 45% of neutral variants; while mutagenesis data of 12 proteins artificially designed under no other constraints but stability, reach an average of 60%. Simulations using a lattice protein model allow us to contrast these estimates to the expected mutational robustness of protein families by generating unbiased samples of foldable sequences, which we find to have 30% of neutral variants. In agreement with mutagenesis data of designed proteins, the model shows that maximally robust protein families might access up to twice the amount of neutral variants observed in the unbiased samples (i.e. 60%). A biophysical model of protein-ligand binding suggests that constraints associated to molecular function have only a moderate impact on robustness of approximately 5 to 10% of neutral variants; and that the direction of this effect depends on the relation between functional performance and thermodynamic stability. Although the genetic code constraints the access of a gene's nucleotide sequence to only 30% of the full distribution of amino acid mutations, it provides an extra 15 to 20% of neutral variants to the estimations above, such that the expected, observed, and maximal robustness of protein-coding genes are approximately 50, 65, and 75%, respectively. We discuss our results in the light of three main hypothesis put forward to explain the existence of mutationally robust genes.

A fundamental paradox motivates the study of plant mitochondrial genomics: the mutation rate is very low (lower than in the nucleus) but the rearrangement rate is high. A landmark paper published in Journal of Molecular Evolution in 1988 established these facts and revealed the paradox. Jeffrey Palmer and Laura Herbon did a prodigious amount of work in the pre-genome sequencing era to identify both the high frequency of rearrangements between closely related species, and the low frequency of mutations, observations that have now been confirmed many times by sequencing. This paper was also the first to use molecular data on rearrangements as a phylogenetic trait to build a parsimonious tree. The work was a technical tour-de-force, its findings are still at the heart of plant mitochondrial genomics, and the underlying molecular mechanisms that produce this paradox are still not completely understood.

Due to their integral roles in oxidative phosphorylation, mitochondrially encoded proteins represent common targets of selection in response to altitudinal hypoxia across high-altitude taxa. While previous studies revealed evidence of positive selection on mitochondrial genomes of high-altitude Phrynocephalus lizards, their conclusions were restricted by out-of-date phylogenies and limited taxonomic sampling. Using topologies derived from both nuclear and mitochondrial DNA phylogenies, we re-assessed the evidence of positive selection on the mitochondrial genomes of high-altitude Phrynocephalus. We sampled representative species from all four main lineages and sequenced the mitochondrial genome of P. maculatus, a putative sister taxon to the high-altitude group. Positive selection was assessed through two widely used branch-site tests: the branch-site model in PAML and BUSTED in HyPhy. No evidence of positive selection on mitochondrial genes was detected on branches leading to two most recent common ancestors of high-altitude species; however, we recovered evidence of positive selection on COX1 on the P. forsythii branch, which represents a reversal from high- to low-elevation environments. A positively selected site therein marked a threonine to valine substitution at position 419. We suggest this bout of selection occurred as the ancestors of P. forsythii re-colonized lower altitude environments north of the Tibetan Plateau. Despite their role in oxidative phosphorylation, we posit that mitochondrial genes are unlikely to have represented historical targets of selection for high-altitude adaptation in Phrynocephalus. Consequently, future studies should address the roles of nuclear genes and differential gene expression.

Which variables determine the constraints on gene sequence evolution is one of the most central questions in molecular evolution. In the fission yeast Schizosaccharomyces pombe, an important model organism, the variables influencing the rate of sequence evolution have yet to be determined. Previous studies in other single celled organisms have generally found gene expression levels to be most significant, with numerous other variables such as gene length and functional importance identified as having a smaller impact. Using publicly available data, we used partial least squares regression, principal components regression, and partial correlations to determine the variables most strongly associated with sequence evolution constraints. We identify centrality in the protein–protein interactions network, amino acid composition, and cellular location as the most important determinants of sequence conservation. However, each factor only explains a small amount of variance, and there are numerous variables having a significant or heterogeneous influence. Our models explain more than half of the variance in dN, raising the possibility that future refined models could quantify the role of stochastics in evolutionary rate variation.

Molluscan shells are composed of calcium carbonates, with small amounts of extracellular matrices secreted from mantle epithelial cells. Many types of shell matrix proteins (SMPs) have been identified from molluscan shells or mantle cells. The pen shell Atrina pectinata (Pinnidae) has two different shell microstructures, the nacreous and prismatic layers. Nacreous and prismatic layer-specific matrix proteins have been reported in Pteriidae bivalves, but remain unclear in Pinnidae. We performed transcriptome analysis using the mantle cells of A. pectinata to screen the candidate transcripts involved in its prismatic layer formation. We found Asprich and nine highly conserved prismatic layer-specific SMPs encoding transcript in P. fucata, P. margaritifera, and P. maxima (Tyrosinase, Chitinase, EGF-like proteins, Fibronectin, valine-rich proteins, and prismatic uncharacterized shell protein 2 [PUSP2]) using molecular phylogenetic analysis or multiple alignment. We confirmed these genes were expressed in the epithelial cells of the mantle edge (outer surface of the outer fold) and the mantle pallium. Phylogenetic character mapping of these SMPs was used to infer a possible evolutionary scenario of them in Pteriomorphia. EGF-like proteins, Fibronectin, and valine-rich proteins encoding genes each evolved in the linage leading to four Pteriomorphia (Mytilidae, Pinnidae, Ostreidae, and Pteriidae), PUSP2 evolved in the linage leading to three Pteriomorphia families (Pinnidae, Ostreidae, and Pteriidae), and chitinase was independently evolved as SMPs in Mytilidae and in other Pteriomorphia (Pinnidae, Ostreidae, and Pteriidae). Our results provide a new dataset for A. pectinata SMP annotation, and a basis for understanding the evolution of prismatic layer formation in bivalves.

Relative to other vertebrates, birds have unusually high blood glucose levels. In humans, the hyperglycemia observed in birds would be associated with diabetes mellitus and the non-enzymatic glycation of proteins, which leads to the accumulation of advanced glycation products and to a plethora of microvascular pathologies. How do birds avoid the negative effects of hyperglycemia? Anthony-Regnitz et al. (J Mol Evol 88: 653-661, 2020) discovered that birds might have evolved glycation-resistant proteins. Serum albumin is an important multifunctional protein susceptible to glycation. Anthony-Regnitz et al. (J Mol Evol 88: 653-661, 2020) found that chicken albumin is resistant to glycation relative to bovine serum albumin. Protein glycation takes place primarily in lysine residues, which are less abundant in chicken than in bovine serum albumin. A multispecies comparison of serum albumin sequences revealed lower numbers of lysine residues in birds than in mammals. Benign hyperglycemia is a shared derived trait of birds and glycation resistance mechanisms appear to have accompanied its evolution. The evolution of benign hyperglycemia in birds coincided with a genomic upheaval that included the loss of important genes, including the one that codes for GLUT4, the transporter responsible for insulin-dependent glucose transport in other vertebrates' insulin-sensitive cells. This loss seems to have resulted in the remodeling of the insulin-signaling pathway in bird tissues. Avian hyperglycemia has been considered a mystery for a long time. Although we remain ignorant of its origins and its repercussions for the physiology of birds, the discovery of resistance to glycation in bird serum albumin offers a path forward to solve this mystery.

Protein glycation and formation of advanced glycation end products is associated with several diseases resulting from high blood glucose concentrations. Plasma albumin is directly exposed to circulating glucose concentrations and is therefore at greater risk of glycation than hemoglobin. As plasma glucose concentrations in birds are 1.5-2 times higher than mammals of similar mass, avian albumin may be particularly at risk of glycation. Thus, the goal of the present study was to compare the in vitro formation of glycated albumin in chicken serum albumin (CSA) and bovine serum albumin (BSA) exposed to a range of glucose concentrations over a 16-week period. The level of glycation for CSA and BSA was quantified using boronate affinity columns to separate glycated albumin from non-glycated albumin and calculating the difference in protein concentration of each sample. The results indicate that CSA is glycated to a lesser degree than BSA when the albumins are exposed to increasing concentrations of glucose (38.8-500mM). This was most apparent at week sixteen (500mM glucose) where BSA expressed a higher degree of glycation (37.8 ± 0.76%) compared to CSA (19.7 ± 1.06%, P < 0.05). Additionally, percent glycation at week sixteen was significantly higher than the glucose-free solutions for both BSA and CSA, indicating that glycation is glucose-dependent. Analyses of the protein structures suggest that the relative resistance of CSA to glycation may be due to fewer lysine residues and variations in protein folding that shield more lysine residues from the plasma. Moreover, comparisons of reconstructed ancestral albumin sequences show that the ancestor of birds had 6-8 fewer lysine residues compared to that of mammals.

The function of a protein is primarily determined by its structure and amino acid sequence. Many biological questions of interest rely on being able to accurately determine the group of structures to which domains of a protein belong; this can be done through alignment and comparison of protein structures. Dozens of different methods for Protein Structure Alignment (PSA) have been proposed that use a wide range of techniques. The aim of this study is to determine the ability of PSA methods to identify pairs of protein domains known to share differing levels of structural similarity, and to assess their utility for clustering domains from several different folds into known groups. We present the results of a comprehensive investigation into eighteen PSA methods, to our knowledge the largest piece of independent research on this topic. Overall, SP-AlignNS (non-sequential) was found to be the best method for classification, and among the best performing methods for clustering. Methods (where possible) were split into the algorithm used to find the optimal alignment and the score used to assess similarity. This allowed us to largely separate the algorithm from the score it maximizes and thus, to assess their effectiveness independently of each other. Surprisingly, we found that some hybrids of mismatched scores and algorithms performed better than either of the native methods at classification and, in some cases, clustering as well. It is hoped that this investigation and the accompanying discussion will be useful for researchers selecting or designing methods to align protein structures.