Social Networking of Quasi-Species Consortia drive Virolution via Persistence.

The emergence of cooperative quasi-species consortia (QS-C) thinking from the more accepted quasispecies equations of Manfred Eigen, provides a conceptual foundation from which concerted action of RNA agents can now be understood. As group membership becomes a basic criteria for the emergence of living systems, we also start to understand why the history and context of social RNA networks become crucial for survival and function. History and context of social RNA networks also lead to the emergence of a natural genetic code. Indeed, this QS-C thinking can also provide us with a transition point between the chemical world of RNA replicators and the living world of RNA agents that actively differentiate self from non-self and generate group identity with membership roles. Importantly the social force of a consortia to solve complex, multilevel problems also depend on using opposing and minority functions. The consortial action of social networks of RNA stem-loops subsequently lead to the evolution of cellular organisms representing a tree of life.

That is life: communicating RNA networks from viruses and cells in continuous interaction.

All the conserved detailed results of evolution stored in DNA must be read, transcribed, and translated via an RNA-mediated process. This is required for the development and growth of each individual cell. Thus, all known living organisms fundamentally depend on these RNA-mediated processes. In most cases, they are interconnected with other RNAs and their associated protein complexes and function in a strictly coordinated hierarchy of temporal and spatial steps (i.e., an RNA network). Clearly, all cellular life as we know it could not function without these key agents of DNA replication, namely rRNA, tRNA, and mRNA. Thus, any definition of life that lacks RNA functions and their networks misses an essential requirement for RNA agents that inherently regulate and coordinate (communicate to) cells, tissues, organs, and organisms. The precellular evolution of RNAs occurred at the core of the emergence of cellular life and the question remained of how both precellular and cellular levels are interconnected historically and functionally. RNA networks and RNA communication can interconnect these levels. With the reemergence of virology in evolution, it became clear that communicating viruses and subviral infectious genetic parasites are bridging these two levels by invading, integrating, coadapting, exapting, and recombining constituent parts in host genomes for cellular requirements in gene regulation and coordination aims. Therefore, a 21st century understanding of life is of an inherently social process based on communicating RNA networks, in which viruses and cells continuously interact.

Persistent virus and addiction modules: an engine of symbiosis.

The giant DNA viruses are highly prevalent and have a particular affinity for the lytic infection of unicellular eukaryotic host. The giant viruses can also be infected by inhibitory virophage which can provide lysis protection to their host. The combined protective and destructive action of such viruses can define a general model (PD) of virus-mediated host survival. Here, I present a general model for role such viruses play in the evolution of host symbiosis. By considering how virus mixtures can participate in addiction modules, I provide a functional explanation for persistence of virus derived genetic 'junk' in their host genomic habitats.

Viruses and the placenta: the essential virus first view.

A virus first perspective is presented as an alternative hypothesis to explain the role of various endogenized retroviruses in the origin of the mammalian placenta. It is argued that virus-host persistence is a key determinant of host survival and the various ERVs involved have directly affected virus-host persistence.

When Competing Viruses Unify: Evolution, Conservation, and Plasticity of Genetic Identities.

In the early 1970s, Manfred Eigen and colleagues developed the quasispecies model (qs) for the population-based origin of RNAs representing the early genetic code. The Eigen idea is basically that a halo of mutants is generated by error-prone replication around the master fittest type which will behave similarly as a biological population. But almost from the start, very interesting and unexpected observations were made regarding competition versus co-operation which suggested more complex interactions. It thus became increasingly clear that although viruses functioned similar to biological species, their behavior was much more complex than the original theory could explain, especially adaptation without changing the consensus involving minority populations. With respect to the origin of natural codes, meaning, and code-use in interactions (communication), it also became clear that individual fittest type-based mechanisms were likewise unable to explain the origin of natural codes such as the genetic code with their context- and consortia-dependence (pragmatic nature). This, instead, required the participation of groups of agents competent in the code and able to edit code because natural codes do not code themselves. Three lines of inquiry, experimental virology, quasispecies theory, and the study of natural codes converged to indicate that consortia of co-operative RNA agents such as viruses must be involved in the fitness of RNA and its involvement in communication, i.e., code-competent interactions. We called this co-operative form quasispecies consortia (qs-c). They are the essential agents that constitute the possibility of evolution of biological group identity. Finally, the basic interactional motifs for the emergence of group identity, communication, and co-operation-together with its opposing functions-are explained by the "Gangen" hypothesis.

Force for ancient and recent life: viral and stem-loop RNA consortia promote life.

Lytic viruses were thought to kill the most numerous host (i.e., kill the winner). But persisting viruses/defectives can also protect against viruses, especially in a ubiquitous virosphere. In 1991, Yarmolinsky et al. discovered the addiction modules of P1 phage, in which opposing toxic and protective functions stabilize persistence. Subsequently, I proposed that lytic and persisting cryptic virus also provide addiction modules that promote group identity. In eukaryotes (and the RNA world), a distinct RNA virus-host relationship exists. Retrovirurses/retroposons are major contributors to eukaryotic genomes. Eukaryotic complexity appears to be mostly mediated by regulatory complexity involving noncoding retroposon-derived RNA. RNA viruses evolve via quasispecies, which contain cooperating, minority, and even opposing RNA types. Quasispecies can also demonstrate group preclusion (e.g., hepatitis C). Stem-loop RNA domains are found in long terminal repeats (and viral RNA) and mediate viral regulation/identity. Thus, stem-loop RNAs may be ancestral regulators. I consider the RNA (ribozyme) world scenario from the perspective of addiction modules and cooperating quasispecies (i.e., subfunctional agents that establish group identity). Such an RNA collective resembles a "gang" but requires the simultaneous emergence of endonuclease, ligase, cooperative catalysis, group identity, and history markers (RNA). I call such a collective a gangen (pathway to gang) needed for life to emerge.

Rethinking quasispecies theory: From fittest type to cooperative consortia.

Recent investigations surprisingly indicate that single RNA "stem-loops" operate solely by chemical laws that act without selective forces, and in contrast, self-ligated consortia of RNA stem-loops operate by biological selection. To understand consortial RNA selection, the concept of single quasi-species and its mutant spectra as drivers of RNA variation and evolution is rethought here. Instead, we evaluate the current RNA world scenario in which consortia of cooperating RNA stem-loops (not individuals) are the basic players. We thus redefine quasispecies as RNA quasispecies consortia (qs-c) and argue that it has essential behavioral motifs that are relevant to the inherent variation, evolution and diversity in biology. We propose that qs-c is an especially innovative force. We apply qs-c thinking to RNA stem-loops and evaluate how it yields altered bulges and loops in the stem-loop regions, not as errors, but as a natural capability to generate diversity. This basic competence-not error-opens a variety of combinatorial possibilities which may alter and create new biological interactions, identities and newly emerged self identity (immunity) functions. Thus RNA stem-loops typically operate as cooperative modules, like members of social groups. From such qs-c of stem-loop groups we can trace a variety of RNA secondary structures such as ribozymes, viroids, viruses, mobile genetic elements as abundant infection derived agents that provide the stem-loop societies of small and long non-coding RNAs.

The DNA Habitat and its RNA Inhabitants: At the Dawn of RNA Sociology.

Most molecular biological concepts derive from physical chemical assumptions about the genetic code that are basically more than 40 years old. Additionally, systems biology, another quantitative approach, investigates the sum of interrelations to obtain a more holistic picture of nucleotide sequence order. Recent empirical data on genetic code compositions and rearrangements by mobile genetic elements and noncoding RNAs, together with results of virus research and their role in evolution, does not really fit into these concepts and compel a reexamination. In this review, we try to find an alternate hypothesis. It seems plausible now that if we look at the abundance of regulatory RNAs and persistent viruses in host genomes, we will find more and more evidence that the key players that edit the genetic codes of host genomes are consortia of RNA agents and viruses that drive evolutionary novelty and regulation of cellular processes in all steps of development. This agent-based approach may lead to a qualitative RNA sociology that investigates and identifies relevant behavioral motifs of cooperative RNA consortia. In addition to molecular biological perspectives, this may lead to a better understanding of genetic code evolution and dynamics.

Phycodnavirus potassium ion channel proteins question the virus molecular piracy hypothesis.

Phycodnaviruses are large dsDNA, algal-infecting viruses that encode many genes with homologs in prokaryotes and eukaryotes. Among the viral gene products are the smallest proteins known to form functional K(+) channels. To determine if these viral K(+) channels are the product of molecular piracy from their hosts, we compared the sequences of the K(+) channel pore modules from seven phycodnaviruses to the K(+) channels from Chlorella variabilis and Ectocarpus siliculosus, whose genomes have recently been sequenced. C. variabilis is the host for two of the viruses PBCV-1 and NY-2A and E. siliculosus is the host for the virus EsV-1. Systematic phylogenetic analyses consistently indicate that the viral K(+) channels are not related to any lineage of the host channel homologs and that they are more closely related to each other than to their host homologs. A consensus sequence of the viral channels resembles a protein of unknown function from a proteobacterium. However, the bacterial protein lacks the consensus motif of all K(+) channels and it does not form a functional channel in yeast, suggesting that the viral channels did not come from a proteobacterium. Collectively, our results indicate that the viruses did not acquire their K(+) channel-encoding genes from their current algal hosts by gene transfer; thus alternative explanations are required. One possibility is that the viral genes arose from ancient organisms, which served as their hosts before the viruses developed their current host specificity. Alternatively the viral proteins could be the origin of K(+) channels in algae and perhaps even all cellular organisms.

Viral ancestors of antiviral systems.

All life must survive their corresponding viruses. Thus antiviral systems are essential in all living organisms. Remnants of virus derived information are also found in all life forms but have historically been considered mostly as junk DNA. However, such virus derived information can strongly affect host susceptibility to viruses. In this review, I evaluate the role viruses have had in the origin and evolution of host antiviral systems. From Archaea through bacteria and from simple to complex eukaryotes I trace the viral components that became essential elements of antiviral immunity. I conclude with a reexamination of the 'Big Bang' theory for the emergence of the adaptive immune system in vertebrates by horizontal transfer and note how viruses could have and did provide crucial and coordinated features.

Viruses are essential agents within the roots and stem of the tree of life.

In contrast with former definitions of life limited to membrane-bound cellular life forms which feed, grow, metabolise and replicate (i) a role of viruses as genetic symbionts, (ii) along with peripheral phenomena such as cryptobiosis and (iii) the horizontal nature of genetic information acquisition and processing broaden our view of the tree of life. Some researchers insist on the traditional textbook conviction of what is part of the community of life. In a recent review [Moreira, D., Lopez-Garcia, P., 2009. Ten reasons to exclude viruses from the tree of life. Nat. Rev. Microbiol. 7, 306-311.] they assemble four main arguments which should exclude viruses from the tree of life because of their inability to self-sustain and self-replicate, their polyphyly, the cellular origin of their cell-like genes and the volatility of their genomes. In this article we will show that these features are not coherent with current knowledge about viruses but that viral agents play key roles within the roots and stem of the tree of life.

The source of self: genetic parasites and the origin of adaptive immunity.

Stable colonization of the host by viruses (genetic parasites) can alter the systems of host identity and provide immunity against related viruses. To attain the needed stability, some viruses of prokaryotes (P1 phage) use a strategy called an addiction module. The linked protective and destructive gene functions of an addiction module insures both virus persistence but will also destroy cells that interrupt this module and thereby prevent infection by competitors. Previously, I have generalized this concept to also include persistent and lytic states of virus infection, which can be considered as a virus addiction module. Such states often involve defective viruses. In this report, I examine the origin of the adaptive immune system from the perspective of a virus addiction module. The likely role of both endogenous and exogenous retroviruses, DNA viruses, and their defective elements is considered in the origin of all the basal components of adaptive immunity (T-cell receptor, RAG-mediated gene rearrangement, clonal lymphocyte proliferation, antigen surface presentation, apoptosis, and education of immune cells). It is concluded that colonization by viruses and their defectives provides a more coherent explanation for the origin of adaptive immunity.

Persistence pays: how viruses promote host group survival.

Recently, we have realized that viruses numerically dominate all life. Although viruses are known to affect host survival in populations, this has not been previously evaluated in the context of host group selection. Group selection per se is not a currently accepted idea and its apparent occurrence is explained by statistical gene frequency models of kin selection. Viruses were not considered in such models. Prevalent views associate viruses and disease. Yet many viruses establish species-specific persistent, inapparent infections that are stable on an evolutionary time scale. Such persistent infections can have large effects on relative reproductive fitness of competing host populations. In this essay, I present arguments on how persistent infections can promote population survival. Mouse hepatitis virus is used as well studied examplar to re-evaluate the theoretical basis of the mouse haystack model of M Smith. This virus-centric re-examination concludes that viruses can indeed affect and promote relative group selection.

Profiling the humoral immune response to infection by using proteome microarrays: high-throughput vaccine and diagnostic antigen discovery.

Despite the increasing availability of genome sequences from many human pathogens, the production of complete proteomes remains at a bottleneck. To address this need, a high-throughput PCR recombination cloning and expression platform has been developed that allows hundreds of genes to be batch-processed by using ordinary laboratory procedures without robotics. The method relies on high-throughput amplification of each predicted ORF by using gene specific primers, followed by in vivo homologous recombination into a T7 expression vector. The proteins are expressed in an Escherichia coli-based cell-free in vitro transcription/translation system, and the crude reactions containing expressed proteins are printed directly onto nitrocellulose microarrays without purification. The protein microarrays are useful for determining the complete antigen-specific humoral immune-response profile from vaccinated or infected humans and animals. The system was verified by cloning, expressing, and printing a vaccinia virus proteome consisting of 185 individual viral proteins. The chips were used to determine Ab profiles in serum from vaccinia virus-immunized humans, primates, and mice. Human serum has high titers of anti-E. coli Abs that require blocking to unmask vaccinia-specific responses. Naive humans exhibit reactivity against a subset of 13 antigens that were not associated with vaccinia immunization. Naive mice and primates lacked this background reactivity. The specific profiles between the three species differed, although a common subset of antigens was reactive after vaccinia immunization. These results verify this platform as a rapid way to comprehensively scan humoral immunity from vaccinated or infected humans and animals.

Structural proteomics of the poxvirus family.

Recent concerns over the potential use of variola virus-commonly known as smallpox-and other orthopox viruses as weapons of bioterrorism have increased research efforts towards creating new antiviral drugs and safer more effective vaccines. Here we introduce a new resource for structural information of poxvirus proteins: the poxvirus proteomics database (PPDB). In the PPDB, we leverage recently developed bioinformatics structure prediction tools on a genomic scale and provide results in a publicly accessible format. The current version of the system contains both experimentally determined and predicted information about protein structural features, such as secondary structure and relative solvent accessibility, as well as tertiary structure and homology information. The system is automated to read the primary sequences from the database, produce the new information for each sequence, and update the database monthly and as new tools are incorporated. The PPDB contains detailed information on the open reading frames (ORFs) in the Copenhagen strain of the vaccinia virus genome. The contents of the PPDB can be accessed through a simple web interface. Inclusion of additional poxvirus genomes in the PPDB is in progress. The PPDB has an upward scalable informatics infrastructure that can readily be applied to viral, bacterial, as well as eukaryotic genomes.

Detection of human polyomaviruses and papillomaviruses in prostatic tissue reveals the prostate as a habitat for multiple viral infections.

BACKGROUND: To determine whether human polyomavirus (hPy) genomes are present in prostate tissues, we have carried out a polymerase chain reaction (PCR) screening in two sets of prostate samples, archival and fresh frozen, as well as performing in situ hybridization (ISH). The frozen prostate samples as well as the urine from the same patients were also screened for human papillomavirus (HPV) sequences. METHODS: Highly sensitive nested-PCR assays were used. The detection of subpopulations of JC virus (JCV) -transcriptional control regions (TCRs) was also evaluated by Southern analysis and by direct DNA sequencing. An in situ hybridization technique was also used to detect JCV DNA in prostatic tissue. RESULTS: The paraffin-embedded archival samples gave variable, unsatisfactory results. Results from the fresh frozen samples, however, were consistent and were frequently positive for JCV and less frequent for BK virus DNA. ISH confirmed the presence of JCV DNA in prostatic glandular epithelium. The TCR region of JCV from prostate tissue and urine from prostate cancer patients showed the presence of both archetypal and rearranged TCRs, including several new sequence variants. HPV DNA was also frequently detected and in some cases also mixed with hPy DNA from frozen tissue and urine. CONCLUSION: The use of fresh frozen samples proved to be essential for consistent and reproducible detection of HPV and hPy viral DNAs. The presence of JCV DNA by ISH and the occurrence of a subpopulation of JCV TCR regions suggests that the prostate is a site for virus replication. The prostate is a complex habitat where mixed infections with oncogenic DNA viruses frequently occur and opens the discussion to the potential role of these viruses in the cancer of the prostate.

Evidence of diversifying selection in human papillomavirus type 16 E6 but not E7 oncogenes.

Human papillomavirus type 16 is a common sexually transmitted pathogen capable of giving rise to cervical intraepithelial neoplasia and invasive carcinoma through the expression and activity of two adjacent oncogenes: E6 and E7. Naturally occurring amino acid variation is commonly observed in the E6 protein but to a much lesser extent in E7. In order to investigate the evolutionary mechanisms involved in the generation and maintenance of this variation, we examine 42 distinct E6-E7 haplotypes using codon-based genealogical techniques. These techniques involve estimation of the ratio of nonsynonymous to synonymous substitutions (dn/ds) and allow testing for directional (positive) natural selection. Positive selection was detected for four codon sites within the E6 oncogene but not in any E7 codons. The amino acid compositions and locations of selected sites are described. Possible sources of natural selection including antiviral immune pressure and polymorphism of host cellular proteins are discussed.