Genetics

In contrast to the human standard for mind established by Alan Turing, I search for a "minimal mind," which is present in animals and even lower-level organisms. Mind is a tool for the classi fi cation and modeling of objects. Its origin marks an evolutionary transition from protosemiotic agents, whose signs directly control actions, to eusemiotic agents, whose signs correspond to ideal objects. The hallmark of mind is a holistic perception of objects, which is not reducible to individual features or signals. Mind can support true intentionality of agents because goals become represented by classes or states of objects. Basic components of mind appear in the evolution of protosemiotic agents; thus, the emergence of mind was inevitable. The classi fi cation capacity of mind may have originated from the ability of organisms to classify states of their own body. Within primary modeling systems, ideal objects are not connected with each other and often tailored for speci fi c functions, whereas in the secondary modeling system, ideal objects are independent from functions and become interconnected via arbitrarily established links. Testing of models can be described by commuting diagrams that integrate measurements, model predictions, object tracking, and actions. Language, which is the tertiary modeling system, supports ef fi cient communication of models between individuals.

Background: Target genes of a transcription factor (TF) Pou5f1 (Oct3/4 or Oct4), which is essential for pluripotency maintenance and self-renewal of embryonic stem (ES) cells, have previously been identified based on their response to Pou5f1 manipulation and occurrence of Chromatin-immunoprecipitation (ChIP)-binding sites in promoters. However, many responding genes with binding sites may not be direct targets because response may be mediated by other genes and ChIP-binding site may not be functional in terms of transcription regulation.

POU transcription factor Pou5f1 (Oct3/4) is required to maintain ES cells in an undifferentiated state. Here we show that global expression profiling of Oct3/4-manipulated ES cells delineates the downstream target genes of Oct3/4. Combined with data from genome-wide chromatin-immunoprecipitation (ChIP) assays, this analysis identifies not only primary downstream targets of Oct3/4, but also secondary or tertiary targets. Furthermore, the analysis also reveals that downstream target genes are regulated either positively or negatively by Oct3/4. Identification of a group of genes that show both activation and repression depending on Oct3/4 expression levels provides a possible mechanism for the requirement of appropriate Oct3/4 expression to maintain undifferentiated ES cells. As a proof-of-principle study, one of the downstream genes, Tcl1, has been analyzed in detail. We show that Oct3/4 binds to the promoter region of Tcl1 and activates its transcription. We also show that Tcl1 is involved in the regulation of proliferation, but not differentiation, in ES cells. These findings suggest that the global expression profiling of gene-manipulated ES cells can help to delineate the structure and dynamics of gene regulatory networks.

Decreasing oocyte competence with maternal aging is a major factor in human infertility. To investigate the age-dependent molecular changes in a mouse model, we compared the expression profiles of metaphase II oocytes collected from 5-to 6-week-old mice with those collected from 42-to 45-week-old mice using the NIA 22K 60-mer oligo microarray. Among approximately 11 000 genes whose transcripts were detected in oocytes, about 5% (530) showed statistically significant expression changes, excluding the possibility of global decline in transcript abundance. Consistent with the generally accepted view of aging, the differentially expressed genes included ones involved in mitochondrial function and oxidative stress. However, the expression of other genes involved in chromatin structure, DNA methylation, genome stability and RNA helicases was also altered, suggesting the existence of additional mechanisms for aging. Among the transcripts decreased with aging, we identified and characterized a group of new oocyte-specific genes, members of the human NACHT, leucine-rich repeat and PYD-containing (NALP) gene family. These results have implications for aging research as well as for clinical ooplasmic donation to rejuvenate aging oocytes.

Specific neuronal types derived from embryonic stem cells (ESCs) can facilitate mechanistic studies and potentially aid in regenerative medicine. Existing induction methods, however, mostly rely on the effects of the combined action of multiple added growth factors, which generally tend to result in mixed populations of neurons. Here, we report that overexpression of specific transcription factors (TFs) in ESCs can rather guide the differentiation of ESCs towards specific neuron lineages. Analysis of data on gene expression changes 2 d after induction of each of 185 TFs implicated candidate TFs for further ESC differentiation studies. Induction of 23 TFs (out of 49 TFs tested) for 6 d facilitated neural differentiation of ESCs as inferred from increased proportion of cells with neural progenitor marker PSA-NCAM. We identified early activation of the Notch signaling pathway as a common feature of most potent inducers of neural differentiation. The majority of neuron-like cells generated by induction of Ascl1,

Mouse embryonic stem cells (ESCs) can differentiate into a wide range – and possibly all cell types in vitro, and thus provide an ideal platform to study systematically the action of transcription factors (TFs) in cell differentiation. Previously, we have generated and analyzed 137 TF-inducible mouse ESC lines. As an extension of this " NIA Mouse ESC Bank, " we generated and characterized 48 additional mouse ESC lines, in which single TFs in each line could be induced in a doxycycline-controllable manner. Together, with the previous ESC lines, the bank now comprises 185 TF-manipulable ESC lines (>10% of all mouse TFs). Global gene expression (transcriptome) profiling revealed that the induction of individual TFs in mouse ESCs for 48 hours shifts their transcriptomes toward specific differentiation fates (e.g., neural lineages by Myt1 Isl1, and St18; mesodermal lineages by Pitx1, Pitx2, Barhl2, and Lmx1a; white blood cells by Myb, Etv2, and Tbx6, and ovary by Pitx1, Pitx2, and Dmrtc2). These data also provide and lists of inferred target genes of each TF and possible functions of these TFs. The results demonstrate the utility of mouse ESC lines and their transcriptome data for understanding the mechanism of cell differentiation and the function of TFs. Pluripotent stem cells, such as embryonic stem cells (ESCs) are able to differentiate into many different cell types in vitro 1. Because the ESCs are also immortal and can maintain their pluripotency, they can be inexhaustible research tools for investigating cell differentiation processes. Previously, we had established a " NIA Mouse ESC Bank " of 137 ESC lines, each of which carries a transcription factor (TF) that can be induced in a doxycycline-controllable manner 2,3. We also carried out global gene expression profiling of these ESC lines 48 hours after the induction of TFs and demonstrated that these transcriptome data indicate the direction of cell differentiation 2,3. In particular, we have validated the cell differentiation into neural lineages, skeletal muscles, hepatocytes, and blood cells 4. To increase the number of manipulated TFs and the coverage of cell types, we have generated ESC lines with 48 new transgenic TFs. With a total of 185 TFs, the " NIA Mouse ESC Bank " covers about 10% of all TFs encoded in the mouse genome 5. We have measured the global gene expression profiles of these new ESC lines 48 hours after overexpressing each TF and compared the changes to tissue-specific gene expression profiles and functionally annotated gene sets. Results Generation of TF-inducible mouse ESC lines. To generate TF-inducible ESC lines, we used the procedure reported previously 2,3. The exogenous copy (transgene) of a TF integrated into the ubiquitously expressing Rosa26 locus 2,6 is repressed by doxycycline (Dox), which is added to the culture medium., TF can then be activated by Dox removal (Fig. 1a). A Venus reporter included into the expression vector was used to visualize cells with the transgene induced (Fig. 1a,b). All tested clones showed at least 70% Venus-positive cells after the removal of Dox (Fig. 1b). The majority of the forty-eight new manipulated genes were TFs selected from a set of high-priority genes involved in important functions in mouse ES cells and their differentiation 7 .

Biological evolution is often viewed narrowly as a change of morphology or allele frequency in a sequence of generations. Here I pursue an alternative informational concept of evolution, as preservation, advance, and emergence of functional information in natural agents. Functional information is a network of signs (e.g., memory, transient messengers, and external signs) that are used by agents to preserve and regulate their functions. Functional information is preserved in evolution via complex interplay of copying and construction processes: the digital components are copied, whereas interpreting subagents together with scaffolds, tools, and resources, are constructed. Some of these processes are simple and invariant, whereas others are complex and contextual. Advance of functional information includes improvement and modification of already existing functions. Although the genome information may change passively and randomly, the interpretation is active and guided by the logic of agent behavior and embryonic development. Emergence of new functions is based on the reinterpretation of already existing information, when old tools, resources, and control algorithms are adopted for novel functions. Evolution of functional information progressed from protosemiosis, where signs correspond directly to actions, to eusemiosis, where agents associate signs with objects. Language is the most advanced form of eusemiosis, where the knowledge of objects and models is communicated between agents.

The life-system concept is a promising theoretical base for the system's approach to population ecology. A revision of the principles of this concept is proposed. To explain population dynamics I examine how quantitative characteristics of its pattern depend on the structure of the life-system. Two aspects of regulation in populations are distinguished: m- and v-stability. The first concerns the stability of the mean value of population density, while the second concerns the stability of density variance. They are characterized by coefficients of m- and v-stability. The contribution of ecological processes and their interactions to these coefficients represents their regu- lative role in population dynamics. The importance of process interactions is demon- strated using a simulation model of the common pine sawfly Diprion pini L. The life-system approach is useful for population management as well. Principles of safety and optimization provide a powerful network for selecting the best control strategy taking into account environmental uncertainty and long-term consequences of ap- plied measures.

The origin of life means the emergence of heritable and evolvable self-reproduction. However the mechanisms of primordial heredity were different from those in contemporary cells. Here I argue that primordial life had no nucleic acids; instead heritable signs were represented by isolated catalytically active self-reproducing molecules, similar to extant coenzymes, which presumably colonized surfaces of oil droplets in water. The model further assumes that coenzyme-like molecules (CLMs) changed surface properties of oil droplets (e.g., by oxidizing terminal carbons), and in this way created and sustained favorable conditions for their own self-reproduction. Such niche-dependent self-reproduction is a necessary condition for cooperation between different kinds of CLMs because they have to coexist in the same oil droplets and either succeed or perish together. Additional kinds of hereditary molecules were acquired via coa-lescence of oil droplets carrying different kinds of CLMs or via modification of already existing CLMs. Eventually, polymerization of CLMs became controlled by other polymers used as templates; and this kind of template-based synthesis eventually resulted in the emergence of RNA-like replicons. Apparently , oil droplets transformed into the outer membrane of cells via engulfing water, stabilization of the surface, and osmoregulation. In result, the metabolism was internalized allowing cells to accumulate free-floating resources (e.g., animoacids, ATP), which was a necessary condition for the development of protein synthesis. Thus, life originated from simple but already functional molecules, and its gradual evolution towards higher complexity was driven by cooperation and natural selection. Published by Elsevier Ireland Ltd.

Natural selection is traditionally viewed as a leading factor of evolution, whereas variation is assumed to be random and non-directional. Any order in variation is attributed to epigenetic or developmental constraints that can hinder the action of natural selection. In contrast I consider the positive role of epigenetic mechanisms in evolution because they provide organisms with opportunities for rapid adaptive change. Because the term " constraint " has negative connotations, I use the term " regulated variation " to emphasize the adaptive nature of phenotypic variation, which helps populations and species to survive and evolve in changing environments. The capacity to produce regulated variation is a phenotypic property, which is not described in the genome. Instead, the genome acts as a switchboard, where mostly random mutations switch " on " or " off " preexisting functional capacities of organism components. Thus, there are two channels of heredity: informational (genomic) and structure-functional (phenotypic). Functional capacities of organisms most likely emerged in a chain of modifications and combinations of more simple ancestral functions. The role of DNA has been to keep records of these changes (without describing the result) so that they can be reproduced in the following generations. Evolutionary opportunities include adjustments of individual functions, multitasking, connection between various components of an organism , and interaction between organisms. The adaptive nature of regulated variation can be explained by the differential success of lineages in macro-evolution. Lineages with more advantageous patterns of regulated variation are likely to produce more species and secure more resources (i.e., long-term lineage selection).

A network of transcription factors (TFs) determines cell identity, but identity can be altered by overexpressing a combination of TFs. However , choosing and verifying combinations of TFs for specific cell differentiation have been daunting due to the large number of possible combinations of ~2,000 TFs. Here, we report the identification of individual TFs for lineage-specific cell differentiation based on the correlation matrix of global gene expression profiles. The overexpression of identified or Ascl1—can direct efficient, specific, and rapid differentiation into myocytes, hepatocytes, blood cells, and neurons. Furthermore, transfection of synthetic mRNAs of TFs generates their appropriate target cells. These results demonstrate both the utility of this approach to identify potent TFs for cell differentiation, and the unanticipated capacity of single TFs directly guides differentiation to specific lineage fates.