Ajith Harish

Mitochondrial genomes are retained by selective constraints on protein targeting

Mitochondria are energy-producing organelles in eukaryotic cells considered to be of bacterial origin. The mitochondrial genome has evolved under selection for minimization of gene content, yet it is not known why not all mitochondrial genes have been transferred to the nuclear genome. Here, we predict that hydrophobic membrane proteins encoded by the mitochondrial genomes would be recognized by the signal recognition particle and targeted to the endoplasmic reticulum if they were nuclear-encoded and translated in the cytoplasm. Expression of the mitochondrially encoded proteins Cytochrome oxidase subunit 1, Apocytochrome b, and ATP synthase subunit 6 in the cytoplasm of HeLa cells confirms export to the endoplasmic reticulum. To examine the extent to which the mitochondrial proteome is driven by selective constraints within the eukaryotic cell, we investigated the occurrence of mitochondrial protein domains in bacteria and eukaryotes. The accessory protein domains of the oxidative phosphorylation system are unique to mitochondria, indicating the evolution of new protein folds. Most of the identified domains in the accessory proteins of the ribosome are also found in eukaryotic proteins of other functions and locations. Overall, one-third of the protein domains identified in mitochondrial proteins are only rarely found in bacteria. We conclude that the mitochondrial genome has been maintained to ensure the correct localization of highly hydrophobic membrane proteins. Taken together, the results suggest that selective constraints on the eukaryotic cell have played a major role in modulating the evolution of the mitochondrial genome and proteome.

Mitochondria are not captive bacteria

Lynn Sagans conjecture (1967) that three of the fundamental organelles observed in eukaryote cells, specifically mitochondria, plastids and flagella were once free-living primitive (prokaryotic) cells was accepted after considerable opposition. Even though the idea was swiftly refuted for the specific case of origins of flagella in eukaryotes, the symbiosis model in general was accepted for decades as a realistic hypothesis to describe the endosymbiotic origins of eukaryotes. However, a systematic analysis of the origins of the mitochondrial proteome based on empirical genome evolution models now indicates that 97% of modern mitochondrial protein domains as well their homologues in bacteria and archaea were present in the universal common ancestor (UCA) of the modern tree of life (ToL). These protein domains are universal modular building blocks of modern genes and genomes, each of which is identified by a unique tertiary structure and a specific biochemical function as well as a characteristic sequence profile. Further, phylogeny reconstructed from genome-scale evolution models reveals that Eukaryotes and Akaryotes (archaea and bacteria) descend independently from UCA. That is to say, Eukaryotes and Akaryotes are both primordial lineages that evolved in parallel. Finally, there is no indication of massive inter-lineage exchange of coding sequences during the descent of the two lineages. Accordingly, we suggest that the evolution of the mitochondrial proteome was autogenic (endogenic) and not endosymbiotic (exogenic).

The phylogenomics of protein structures: The backstory.

In this introductory retrospective, evolution as viewed through gene trees is inspected through a lens compounded from its founding operational assumptions. The four assumptions of the gene tree culture that are singularly important to evolutionary interpretations are: a. that protein-coding sequences are molecular fossils; b. that gene trees are equivalent to species trees; c. that the tree of life is assumed to be rooted in a simple akaryote cell implying that akaryotes are primitive, and d. that the notion that all or most incongruities between alignment-based gene trees are due to horizontal gene transfer (HGT), which includes the endosymbiotic models postulated for the origins of eukaryotes. What has been unusual about these particular assumptions is that though each was taken on board explicitly, they are defended in the face of factual challenge by a stolid disregard for the conflicting observations. The factual challenges to the mainstream gene tree-inspired evolutionary view are numerous and most convincingly summarized as: Genome trees tell a very different story. Phylogeny inferred from genomic assortments of homologous protein structural-domains does not support any one of the four principle evolutionary interpretations of gene trees: a. 3D protein domain structures are the molecular fossils of evolution, while coding sequences are transients; b. Species trees are very different from gene trees; c. The ToL is rooted in a surprisingly complex universal common ancestor (UCA) that is distinct from any specific modern descendant and d. HGT including endosymbiosis is a negligible player in genome evolution from UCA to the present.

Empirical genome evolution models root the tree of life

A reliable phylogenetic reconstruction of the evolutionary history of contemporary species depends on a robust identification of the universal common ancestor (UCA) at the root of the Tree of Life (ToL). That root polarizes the tree so that the evolutionary succession of ancestors to descendants is discernable. In effect, the root determines the branching order and the direction of character evolution. Typically, conventional phylogenetic analyses implement time-reversible models of evolution for which character evolution is un-polarized. Such practices leave the root and the direction of character evolution undefined by the data used to construct such trees. In such cases, rooting relies on theoretic assumptions and/or the use of external data to interpret unrooted trees. The most common rooting method, the outgroup method is clearly inapplicable to the ToL, which has no outgroup. Both here and in the accompanying paper (Harish and Kurland, 2017) we have explored the theoretical and technical issues related to several rooting methods. We demonstrate (1) that Genome-level characters and evolution models are necessary for species phylogeny reconstructions. By the same token, standard practices exploiting sequence-based methods that implement gene-scale substitution models do not root species trees; (2) Modeling evolution of complex genomic characters and processes that are non-reversible and non-stationary is required to reconstruct the polarized evolution of the ToL; (3) Rooting experiments and Bayesian model selection tests overwhelmingly support the earlier finding that akaryotes and eukaryotes are sister clades that descend independently from UCA (Harish and Kurland, 2013); (4) Consistent ancestral state reconstructions from independent genome samplings confirm the previous finding that UCA features three fourths of the unique protein domain-superfamilies encoded by extant genomes.

Did viruses evolve as a distinct supergroup from common ancestors of cells

The evolutionary origins of viruses according to marker gene phylogenies, as well as their relationships to the ancestors of host cells remains unclear. In a recent article Nasir and Caetano-Anollés reported that their genome-scale phylogenetic analyses based on genomic composition of protein structural-domains identify an ancient origin of the “viral supergroup” (Nasir et al. 2015. A phylogenomic data-driven exploration of viral origins and evolution. Sci Adv. 1(8):e1500527.). It suggests that viruses and host cells evolved independently from a universal common ancestor. Examination of their data and phylogenetic methods indicates that systematic errors likely affected the results. Reanalysis of the data with additional tests shows that small-genome attraction artifacts distort their phylogenomic analyses, particularly the location of the root of the phylogenetic tree of life that is central to their conclusions. These new results indicate that their suggestion of a distinct ancestry of the viral supergroup is not well supported by the evidence.

Structural biology and genome evolution: An introduction.

For fifty years, reconstruction of a relatively small number of “significant” gene trees has driven the preconceptions and perceptions of molecular evolution. More recently sequencing of thousands of genomes along with 3D structure determination of one hundred thousand proteins has refocused and clarified the outlines of molecular evolution. In particular, phylogenetic analyses of genome scale data based on distributions of modular protein domains challenge conventional beliefs about the evolution of organisms, and their proteomes. In 1992 a note entitled “One thousand families for the molecular biologist” [1] by Cyrus Chothia sparked a radical advance in the study of molecular evolution. His calculations suggested, “The large majority of proteins come from no more than one thousand families”. Further, he recognized the proteins in such structural families as homologs because, as he reasoned, they share “sequences, functional and/or genetic similarities that strongly imply that they are descended from a common ancestor” [1]. Then in 1998, Chothia and company, made the discovery that pairwise sequence comparisons such as BLAST searches commonly used by gene tree enthusiasts seriously underestimate identifications of significantly diverged homologs [2,3]. As Park et al. say, “ Sequence Comparisons Using Multiple Sequences Detect Three Times as Many Remote Homologues as PairwiseMethods” [3]. Thus, without a comprehensive grasp of the distributions of homologous protein characters encoded by genomes, divergent evolution of sequences was often mistaken for convergent evolution of structures [4,5]. Similarly, the realization that missense mutations significantly affect protein folding and function encouraged fundamental concerns about the generality of the neutral theory of molecular evolution [6,7]. It has taken a few decades of large-scale genome sequencing as well as many tens of thousands of 3D structural determinations of proteins to realize the full reach of Chothias structural insights. It is now recognized that most proteins are constructed from one or more compact domains that are in turn associated with shorter terminal regions that we refer to as linkers [8]. Linkers are essential to the associations of proteins with other macromolecules and subcellular structures, while most enzymatic activities reside in the tertiary structures of domains that constitute the major fraction of protein mass [8]. In isolation, linkers and some fibrous proteins are identified with fluid secondary structures, but they present no stable tertiary folds. Stable domains are gathered in large groups with a common 3D structure, for example as superfamilies, of which there are a mere two thousand or so in nature [9]. Such 3D structures or tertiary folds are identifiable in the hundreds of thousands of proteins whose structures have been

What is an archaeon and are the Archaea really unique

The recognition of the group Archaea as a major branch of the tree of life (ToL) prompted a new view of the evolution of biodiversity. The genomic representation of archaeal biodiversity has since significantly increased. In addition, advances in phylogenetic modeling of multi-locus datasets have resolved many recalcitrant branches of the ToL. Despite the technical advances and an expanded taxonomic representation, two important aspects of the origins and evolution of the Archaea remain controversial, even as we celebrate the 40th anniversary of the monumental discovery. These issues concern (i) the uniqueness (monophyly) of the Archaea, and (ii) the evolutionary relationships of the Archaea to the Bacteria and the Eukarya; both of these are relevant to the deep structure of the ToL. To explore the causes for this persistent ambiguity, I examine multiple datasets and different phylogenetic approaches that support contradicting conclusions. I find that the uncertainty is primarily due to a scarcity of information in standard datasets—universal core-genes datasets—to reliably resolve the conflicts. These conflicts can be resolved efficiently by comparing patterns of variation in the distribution of functional genomic signatures, which are less diffused unlike patterns of primary sequence variation. Relatively lower heterogeneity in distribution patterns minimizes uncertainties and supports statistically robust phylogenetic inferences, especially of the earliest divergences of life. This case study further highlights the limitations of primary sequence data in resolving difficult phylogenetic problems, and raises questions about evolutionary inferences drawn from the analyses of sequence alignments of a small set of core genes. In particular, the findings of this study corroborate the growing consensus that reversible substitution mutations may not be optimal phylogenetic markers for resolving early divergences in the ToL, nor for determining the polarity of evolutionary transitions across the ToL.

Mayr Versus Woese: Akaryotes and Eukaryotes

In 1998, on the brink of a great public effort that by now has delivered the sequences of thousands of genomes and has annotated these genomes by translating tens of thousands of 3D protein domain structures from their coding sequences, Ernst Mayr and Carl Woese engaged in a debate. At issue were the virtues of phenotypic contra genotypic approaches to phylogeny and taxonomy. Though not conclusive, this confrontation in retrospect illustrates the defects of both their perspectives and simultaneously illuminates the strengths of the approach to phylogenetic systematics that was favored by Willi Hennig. Hennig’s cladism lends itself well to a rigorous exploitation of genome sequence data in which both the genotypic and phenotypic modes replace the technically questionable gene tree approach to deep phylogeny championed by Woese. Diverse phylogenomic data now suggest that though Mayr’s phenetic arguments were incomplete, his division of organisms into two major taxonomic groups, the akaryotes (formerly the prokaryotes) and eukaryotes, is probably correct. Thus, in a phylogeny based on genome repertoires of protein domains, the universal common ancestor of the three superkingdoms descends in two primary lineages, Akaryote and Eukaryote.

Reply to Caetano-Anollés et al. comment on “Empirical genome evolution models root the tree of life”

We recently analyzed the robustness of competing evolution models developed to identify the root of the Tree of Life: 1) An empirical Sankoff parsimony (ESP) model (Harish and Kurland, 2017), which is a nonstationary and directional evolution model; and 2) An a priori ancestor (APA) model (Kim and Caetano-Anollés, 2011) that is a stationary and reversible evolution model. Both Bayesian model selection tests as well as maximum parsimony analyses demonstrate that the ESP model is, overwhelmingly, the better model. Moreover, we showed that the APA model is not only sensitive to artifacts, but also that the underlying assumptions are neither empirically grounded nor biologically realistic.