Necrosis: a specific form of programmed cell death?

For a long time necrosis was considered as an alternative to programmed cell death, apoptosis. Indeed, necrosis has distinct morphological features and it is accompanied by rapid permeabilization of plasma membrane. However, recent data indicate that, in contrast to necrosis caused by very extreme conditions, there are many examples when this form of cell death may be a normal physiological and regulated (programmed) event. Various stimuli (e.g., cytokines, ischemia, heat, irradiation, pathogens) can cause both apoptosis and necrosis in the same cell population. Furthermore, signaling pathways, such as death receptors, kinase cascades, and mitochondria, participate in both processes, and by modulating these pathways, it is possible to switch between apoptosis and necrosis. Moreover, antiapoptotic mechanisms (e.g., Bcl-2/Bcl-x proteins, heat shock proteins) are equally effective in protection against apoptosis and necrosis. Therefore, necrosis, along with apoptosis, appears to be a specific form of execution phase of programmed cell death, and there are several examples of necrosis during embryogenesis, a normal tissue renewal, and immune response. However, the consequences of necrotic and apoptotic cell death for a whole organism are quite different. In the case of necrosis, cytosolic constituents that spill into extracellular space through damaged plasma membrane may provoke inflammatory response; during apoptosis these products are safely isolated by membranes and then are consumed by macrophages. The inflammatory response caused by necrosis, however, may have obvious adaptive significance (i.e., emergence of a strong immune response) under some pathological conditions (such as cancer and infection). On the other hand, disturbance of a fine balance between necrosis and apoptosis may be a key element in development of some diseases.     

Salinity Adaptations and Bathymetric Distribution of Bivalve Mollusks Portlandia arcticaand Nuculana pernula in the White Sea

We examined changes in the oxygen consumption rate, the number of active animals, rate of losses of salts, and the level of cell hydration under conditions of decreased salinity in the mollusks Portlandia arctica(Gray) and Nuculana pernula(O.F. Müller), inhabiting the White Sea in the deepest depths under high salinity (about 30) and negative temperatures of –1.3 to –1.4°C throughout the year. It has been shown that the mollusks demonstrate a relatively great tolerance of desalination at the organism level and a capability to regulate cell volume under decreased salinity of the environment. We concluded that the depletion of fauna of the White Sea in the deepest depths is not connected with the tolerance level of deep-sea animals to salinity fluctuations, but rather is due to environmental conditions (liquid silts and negative temperatures).     

Synthesis of N-Acetylglucosamine Aryl β-Glycosides Catalyzed by Crown Compounds

Glycosylation of various phenols with -D-glucosaminyl chloride peracetate in a solid phase–liquid system catalyzed by crown compounds was studied. The highest yields of aryl -glycosides were observed at room temperature in acetonitrile using anhydrous potassium carbonate as a base. The optimum phenol–glycosyl donor–base–crown ether ratio was 1 : 1 : 1 : 0.2.     

Expression of a novel HsMCAK mRNA splice variant,tsMCAK gene,in human testis

Identification of specifically expressed genes in the adult or fetal testis is very important for the study of genes related to the development and function of the testis. In this study, a human adult testis cDNA microarray was constructed and hybridized with 33P-labeled human adult and embryo testis cDNA probes, respectively. After differential display analyzing, a number of new genes related to the development of testis and spermatogenesis had been identified. One of these new genes is tsMCAK. tsMCAK was expressed 2.62 folds more in human adult testis than fetal testis. The full length of tsMCAK is 2401 bp and contains a 2013 bp open reading frame, encoding a 671-amino-acid protein. Sequence analysis showed that it has a central kinesin motor domain and is homologous to HsMCAK gene of the somatic cells. Blasting human genome database localized tsMCAK to human chromosome 1P34 and further investigation showed that it is a splice variant of HsMCAK. The tissue distribution of tsMCAK was determined by RT-PCR and it is expressed highly and specifically in the testis. Southern blot studies of its expression in patients with infertility indicated its specific expression in spermatogenic cells and its correlation with male infertility. The above results suggested that tsMCAK is a candidate gene for the testis-specific KRPs and its specific expression in the testis was correlated with spermatogenesis and may be correlated with male infertility.     

Synthesis of Oligonucleotide Derivatives with Aryl(trifluoromethyl)diazirine Moiety for the Photoaffinity Modification of Proteins and Nucleic Acids

1-(4-(3-(Trifluoromethyl)-3H-diazirin-3-yl)benzamido)-3-O-(4,4"-dimethoxytrityl)-2,3-propanediol phosphoramidite was synthesized and used as a modified unit in the automatic synthesis of oligodeoxyribonucleotides. Pentadecathymidylates with various numbers of 2,3-propanediol moieties substituted with aryl(trifluoromethyl)diazirinyl (ATFMD) were obtained, and the thermal stability of their duplexes with (dA)₁₅ were studied. One ATFMD-propanediol residue was shown to reduce the thermal stability of the duplex by 8–9°C. The irradiation of the ATFMD-containing duplexes by UV light with the wavelength of 350 nm was found to cause the cross-linking reaction of the ATFMD-containing strand with the complementary strand and the formation of the cross-linked duplexes. The photomodification efficiency was independent of the oligonucleotide sequence, with each ATFMD group providing for 5% cross-linking. The irradiation of an ATFMD-containing duplex, a substrate of the HIV-1 integrase, in the presence of this enzyme resulted in the covalent DNA–protein complex. The oligonucleotides with the 1-(4-(3-(trifluoromethyl)-3H-diazirin-3-yl)benzamido)-2,3-propanediol moiety in their chains can be used for the photoaffinity modification of both nucleic acids and proteins that recognize them.     

Nuclear wave packet motion between P* and P(+)B(A)(-) potential surfaces with a subsequent electron transfer to H(A) in bacterial reaction centers at 90 K. Electron transfer pathway

In Rhodobacter sphaeroides R-26 reaction centers (RCs) the nuclear wave packet induced by 25 fs excitation at 90 K moves on the primary electron donor P* potential energy hypersurface with initial frequency at approximately 130 cm(-1) (monitored by stimulated emission measurement). At the long-wavelength side of P* stimulated emission at 935 nm the wave packet is transferred to the surface with P(+)B(A)(-) character at 120, 380, 1.2 fs, etc. delays (monitored by measurement of the primary electron acceptor B(A)(-) band at 1020 nm). However, only beginning from 380 fs delay and later the relative stabilization of the state P(+)B(A)(-) is observed. This is accompanied by the electron transfer to bacteriopheophytin H(A) (monitored by H(A) band measurement at 760 nm). The most active mode of 32 cm(-1) in the electron transfer and its overtones up to the seventh were found in the Fourier transform spectrum of the oscillatory part of the kinetics of the P* stimulated emission and of the P(+)B(A)(-) and P(+)H(A)(-) formation. This mode and its overtones are apparently populated via the 130 cm(-1) vibrational mode. The deuteration of the sample shifts the fundamental frequency (32 cm(-1)) and all overtones by the same factor of approximately 1.3. This mode and its overtones are suppressed by a factor of approximately 4.7 in the dry film of RCs. The results obtained indicate that the 32 cm(-1) mode might be related to a rotation of hydrogen-containing groups (possibly the water molecule) participating in the modulation of the primary electron transfer from P* to B(A)(-) in at least 35% of RCs. The Brookhaven Protein Data Bank (1PRC) displays the water molecule located at the position HOH302 between His M200 (axial ligand for P(B)) and the oxygen of ring V of B(A) which might be a part (approximately 35%) of the molecular pathway for electron transfer from P* to B(A).     

Involvement of chloroplasts in the programmed death of plant cells

The effect of cyanide, an apoptosis inducer, on pea leaf epidermal peels was investigated. Illumination stimulated the CN^–-induced destruction of guard cells (containing chloroplasts and mitochondria) but not of epidermal cells (containing mitochondria only). The process was prevented by antioxidants (-tocopherol, 2,5-di-tret-butyl-4-hydroxytoluene, and mannitol), by anaerobiosis, by the protein kinase C inhibitor staurosporine, and by cysteine and serine protease inhibitors. Electron acceptors (menadione, p-benzoquinone, diaminodurene, TMPD, DCPIP, and methyl viologen) suppressed CN^–-induced apoptosis of guard cells, but not epidermal cells. Methyl viologen had no influence on the removal of CN^–-induced nucleus destruction in guard cells under anaerobic conditions. The light activation of CN^–-induced apoptosis of guard cells was suppressed by DCMU (an inhibitor of the electron transfer in Photosystem II) and by DNP-INT (an antagonist of plastoquinol at the Q_o site of the chloroplast cytochrome b ₆ f complex). It is concluded that apoptosis initiation in guard cells depends on the simultaneous availability of two factors, ROS and reduced quinones of the electron transfer chain. The conditions for manifestation of programmed cell death in guard and epidermal cells of the pea leaf were significantly different.     

Endogenous DNases as a Tool for Isolation of Nuclear Matrix: Critical Parameters of Nucleolysis

The degree of nucleolysis is of critical significance for isolation of nuclear matrix (NM) specifically enriched in transcribed DNA sequences, as demonstrated using an example of inactive (c-fos, c-myc, andC ) and active (p53, albumin, and28S rRNA) genes in resting hepatocytes. The optimal degree of nucleolysis is characterized by degradation of loop domains of chromatin, with the relatively uniform molecular weight distribution of DNA being preserved. Deviation from these parameters leads to nonspecific fragmentation of chromatin in various gene loci and isolation of NM samples nonspecifically enriched with or depleted of transcribed DNA sequences. Under optimal hydrolytic conditions, the transcribed chromatin is more resistant to endogenous DNase attack, which allows selective conservation of its association with the nuclear matrix