"Chromatomics" the analysis of the chromatome

Chromatin is a highly complex mixture of proteins and DNA that is involved in the regulation and coordination of gene expression within the eukaryotic nucleus. Changes in chromatin structure can convey heritable changes of gene activity in response to external stimuli without altering the primary DNA sequence. This epigenetic inheritance of particular traits very likely plays a major role during evolutionary processes. It is however, still ill-defined how this non DNA-mediated inheritance is accomplished at a molecular level. The advent of new methods to systematically study genome-wide changes in chromatin condensation, DNA methylation levels, RNA synthesis and the association of specific proteins or protein modifications now allows a thorough investigation of changes in chromatin structure and function in response to environmental alterations. We would like to review some of these global approaches and to introduce the term "chromatomics" for the systematic analysis of the DNA, RNA and protein content of the genetic material in the eukaryotic nucleus.     

Evolution of the "autophagamiR"

MicroRNAs (miRs) are increasingly important diagnostic and prognostic markers in cancer but have not been defined in medullary thyroid carcinoma (MTC). MiR microarray profiling was performed on 19 primary MTC tumors, validated with qPCR in 45 cases and correlated with clinical outcomes. MiRs-183 and 375 were overexpressed and miR-9* underexpressed in sporadic vs. hereditary MTC (SMTC; HMTC). MiR-183 and 375 overexpression predicted lateral nodal metastases, residual disease, distant metastases and mortality. MiR-183 knockdown in an MTC cell line (TT cells) reduced cellular proliferation in association with elevated LC3B expression. This is suggestive of increased autophagic flux and potential cell death via autophagy induction. MiRs may subsequently be shown to serve as efficacious therapeutic strategies in MTC with a mechanism based upon autophagy.     

The distributions of "new" and "old" Alu sequences in the human genome: the solution of a "mystery"

The distribution in the human genome of the largest family of mobile elements, the Alu sequences, has been investigated for the past 30 years, and the vast majority of Alu sequences were shown to have the highest density in GC-rich isochores. Ten years ago, it was discovered, however, that the small "youngest" (most recently transposed) Alu families had a strikingly different distribution compared with the "old" families. This raised the question as to how this change took place in evolution. We solved what was considered to be a "mystery" by 1) revisiting our previous results on the integration and stability of retroviral sequences, and 2) assessing the densities of acceptor sites TTTT/AA in isochore families. We could conclude 1) that the open state of chromatin structure plays a crucial role in allowing not only the initial integration of retroviral sequences but also that of the youngest Alu sequences, and 2) that the distribution of old Alus can be explained as due to Alu sequences being unstable in the GC-poor isochores but stable in the compositionally matching GC-rich isochores, again in line with what happens in the case of retroviral sequences.     

"Light" and "dark" pinealocytes of the porcine pineal gland

Both qualitative and quantitative comparative studies of "dark" and "light" pinealocytes of the porcine pineal gland have been carried out. These cells differ from each other in their electronic density of cytoplasm, shape of nucleus, the structure of membrane bound dense bodies and the number of microtubules and smooth endoplasmic reticulum. The membrane bound dense bodies--characteristic structures of pig pinealocytes as well dense core vesicles occur in both types of cells. The relative volume of the majority of the cells' organellae apart from the Golgi apparatus, also do not show any significant difference. The results obtained support a functional basis for pinealocyte differentiation in the porcine pineal gland.     

Taking the "I" out of IRB--and putting "community" in

If one looks back on the history of American research ethics, a bold pattern emerges. Since World War II, about every twenty years or so a breach of the social contract between investigators and human research subjects galvanizes public and professional interest in the ethical foundations and oversight mechanisms governing research with humans.     

"Dark" and "pale" cells of the amygdala complex in dynamics of the estrous cycle

Data on cytological peculiarities of dorsomedial nucleus neurones of the amygdala complex, one of the main zones of sexual dimorphism, in dynamics of estrous cycle are reported. We show that structural and functional characteristics of "dark" and "pale" cells may change depending on the concentrations of gonadal steroids in estrous and metaestrous stages. This specifies the previous hypothesis about mutual reorganization of these cells.     

An explantion for the determination of "self" versus "non-self" proteins

Deamidation of proteins, which has been linked to turnover, results in the production of a different protein—different in sequence and shape. It is proposed that this is the protein which is normally encountered by the immune system and is therefore viewed as “self”. If the protein in the form in which it exists in situ is released through disease, or introduced by artificial means, it would then be recognized as “non-self.” This is offered as a hypothesis to explain autoimmune response to the basic protein of myelin.     

Characteristics of activity of "fast" and "slow" Purkinje cells of the cerebellum

In the guinea pig cerebellar cortex, three types of Purkinje cells were identified according to the properties of complex spikes: fast, intermediate, and slow cells. Fast Purkinje cells have following properties as compared with slow Purkinje cells: (i) salient components with short intervals in complex impulses (on the average, five components with a period of about 2 ms versus two components with a period of about 4 ms); (ii) a short duration of simple spikes (in the average, 2.13 +/- 0.53 ms versus 3.9 +/- 0.65 ms) and a quick restoration of their amplitude after preceding simple spikes (in the mean, 2.83 +/- 0.75 ms versus 11.0 +/- 2.82 ms); and (iii) a more pronounced rebound in the auto-correlation histogram of simple spikes (3.09 +/- 2.12 versus 1.45 +/- 0.36) and a short-latency excitation of simple spikes after complex spikes (2.81 +/- 1.64 versus 1.26 +/- 0.52). A decrease of interspike intervals in simple spike activity of all Purkinje cells was revealed (5.25 +/- 2.71 ms versus 9.71 +/- 3.48 ms in activity fragments without complex spikes). It is supposed that the properties of complex spikes depend on the type of Purkinje cells and may be one of the basic factors determining the interactions between the inputs of climbing and parallel fibers in Purkinje cells.     

The nature of and the interrelations between the concepts of "life","disease" and "aging"

An attempt is made to consider disease and aging following from the concepts of the essence of life. The proposed definition of life represents a modified Engels' (1878) definition. Proceeding from the analysis of possible mechanisms of different disturbances in the life process leading to a decreased probability of the organism existence it is concluded that disease develops either as a result of hereditary changes in the genome or due-to disorders in its realization under certain unfavorable conditions. Aging is determined by the properties of the genome itself and develops in connection with age increase.     

An analysis of the meaning of the concepts of "commitment" and "determination" in the study of patterns of development

The use of the term "commitment" by different authors was compared and the term itself was compared with the term "determination". Different authors understand the term "commitment" in different ways. It is proposed to preserve the terms "competence", "determination" (labile and stable) and "differentiation" in studies of normal development at stages preceding the appearance of organ rudiments in order to facilitate the use of the knowledge acquired by experimental embryology and to decipher these concepts at the molecular level. The meaning of the term "commitment" should be made more precise when describing the experimental results and also when assessing the results obtained by various authors and published in numerous papers and reviews.     

The crystal structure of the "open" and the "closed" conformation of the flexible loop of trypanosomal triosephosphate isomerase

Triosephosphate isomerase has an important loop near the active site which can exist in a "closed" and in an "open" conformation. Here we describe the structural properties of this "flexible" loop observed in two different structures of trypanosomal triosephosphate isomerase. Trypanosomal triosephosphate isomerase, crystallized in the presence of 2.4 M ammonium sulfate, packs as an asymmetric dimer of 54,000 Da in the crystallographic asymmetric unit. Due to different crystal contacts, peptide 167-180 (the flexible loop of subunit-1) is an open conformation, whereas in subunit-2, this peptide (residues 467-480) is in a closed conformation. In the closed conformation, a hydrogen bond exists between the tip of the loop and a well-defined sulfate ion which is bound to the active site of subunit-2. Such an active site sulfate is not present in subunit-1 due to crystal contacts. When the native (2.4 M ammonium sulfate) crystals are transferred to a sulfate-free mother liquor, the flexible loop of subunit-2 adopts the open conformation. From a closed starting model, this open conformation was discovered through molecular dynamics refinement without manual intervention, despite involving C alpha shifts of up to 7 A. The tip of the loop, residues 472, 473, 474, and 475, moves as a rigid body. Our analysis shows that in this crystal form the flexible loop of subunit-2 faces a solvent channel. Therefore the open and the closed conformations of this flexible loop are virtually unaffected by crystal contacts. The actual observed conformation depends only on the absence or presence of a suitable ligand in the active site.     

Cytogerontology at the beginning of the third millenium: from "correlative" to "gist" models

For the most part, research in the area of cytogerontology, i.e., investigation of the mechanisms of aging in the experiments on cultured cells, is carried out using the "Hayflick's model". More than forty years have passed since the appearance of that model, and during this period of time, very much data were obtained on its basis. These data contributed significantly to our knowledge of the behavior of both animal and human cultured cells. Specifically, we already know of the mechanisms underlying the aging in vitro. On the other hand, in my opinion, little has changed in our knowledge of the aging of the whole organism. In all likelihood, this can be explained by that the Hayflick's model is, like many others used in the experimental gerontology, correlative, i.e. based on a number of detected correlations. In the case of Hayflick's model, these are correlations between the mitotic potential of cells (cell population doubling potential) and some "gerontological" parameters and indices: species life-span, donor age, evidence of progeroid syndromes, etc., as well as various changes of normal (diploid) cells during long-term cultivation and during aging of the organism. It is, however, well known that very frequently a good correlation has nothing to do with the essence (gist) of the phenomenon. For example, we do know that the amount of gray hair correlates quite well with the age of an individual but is in no way related to the mechanisms of his/her aging and probability of death. In this case, the absence of cause-effect relationships is evident, which are, at the same time, indispensable for the development of gist models. These models, as distinct from the correlative ones, are based on a certain concept of aging. In the case of Hayflick's model, such a concept is absent: we cannot explain, using the "Hayflick's limit", why our organism ages. This conclusion was convincingly confirmed by the discovery of telomere mechanism which determines the aging of cells in vitro. That discovery initiated the appearance of theories attempting to explain the process of aging in vivo also on its basis. However, it has become clear that the mechanisms of aging of the entire organism, located, apparently, in its postmitotic cells, such as neurons or cardiomyocytes, cannot be explained in the framework of this approach. Hence, we believe that it is essential to develop "gist" models of aging using cultured cells. The mechanisms of cell aging in such models should be similar to the mechanisms of cell aging in the entire organism. Our "stationary phase aging" model could be one of such models, which is based on the assumption of the leading role of cell proliferation restriction in the processes of aging. We assume that the accumulation of "senile" damage is caused by the restriction of cell proliferation either due to the formation of differentiated cell populations during development (in vivo) or to the existence of saturation density phenomenon (in vitro). Cell proliferation changes themselves do not induce aging, they only lead to the accumulation of macromolecular defects, which, in turn, lead to the deterioration of tissues, organs, and, eventually, of the entire organism, increasing the probability of its death. Within the framework of our model, we define cell aging as the accumulation in a cell population of various types of damage identical to the damage arising in senescing multicellular organism. And, finally, it is essential to determine how the cell is dying and what the death of the cell is. These definitions will help to draw real parallels between the "genuine" aging of cells (i.e., increasing probability of their death with "age") and the aging of multicellular organisms.     

Ultrastructural characteristics of the "light" and "dark" B-cells in the pancreatic islets of rats]

The ultrastructure of B-cells in the rat pancreatic islets has been studied under various experimental conditions (thyroidectomy, continuous thyroxine treatment, regeneration after partial pancreatectomy, thyroidectomy with partial pancreatectomy, partial pancreatectomy, partial pancreatectomy with continuous thyroxine treatment). Five types of B-cells have been distinguished. It has been supposed that "light" B-cell 1 is related to the stage of secretory granule extrusion, "light" B-cell 2 reflects the extrusion of secretory material and the early stages of secretory granule synthesis; "dark" B-cell 1 is involved in the intensive synthesis, formation and extrusion of secretory material, and "dark" B-cell 2 in the intensive secretory granule synthesis, formation and storage.