Vascular Immunotargeting to Endothelial Determinant ICAM-1 Enables Optimal Partnering of Recombinant scFv-Thrombomodulin Fusion with Endogenous Cofactor

The use of targeted therapeutics to replenish pathologically deficient proteins on the luminal endothelial membrane has the potential to revolutionize emergency and cardiovascular medicine. Untargeted recombinant proteins, like activated protein C (APC) and thrombomodulin (TM), have demonstrated beneficial effects in acute vascular disorders, but have failed to have a major impact on clinical care. We recently reported that TM fused with an scFv antibody fragment to platelet endothelial cell adhesion molecule-1 (PECAM-1) exerts therapeutic effects superior to untargeted TM. PECAM-1 is localized to cell-cell junctions, however, whereas the endothelial protein C receptor (EPCR), the key co-factor of TM/APC, is exposed in the apical membrane. Here we tested whether anchoring TM to the intercellular adhesion molecule (ICAM-1) favors scFv/TM collaboration with EPCR. Indeed: i) endothelial targeting scFv/TM to ICAM-1 provides ∼15-fold greater activation of protein C than its PECAM-targeted counterpart; ii) blocking EPCR reduces protein C activation by scFv/TM anchored to endothelial ICAM-1, but not PECAM-1; and iii) anti-ICAM scFv/TM fusion provides more profound anti-inflammatory effects than anti-PECAM scFv/TM in a mouse model of acute lung injury. These findings, obtained using new translational constructs, emphasize the importance of targeting protein therapeutics to the proper surface determinant, in order to optimize their microenvironment and beneficial effects.     

ATMINistrating ATM signaling

The checkpoint kinase ATM (ataxia telangiectasia mutated) transduces genomic stress signals to halt cell cycle progression and promote DNA repair in response to DNA damage. We have recently identified an essential cofactor for ATM, ATMIN (for ATM INteractor). Several observations suggested that ATMIN plays a key role in ATM signalling. ATMIN and ATM protein stability were mutually dependent, which indicated an intimate physical and functional interaction. ATMIN bound ATM using a short carboxy-terminal motif, in a manner analogous to how another ATM cofactor, Nijmegen Breakage Syndrome protein 1 (NBS1), associates with ATM. ATMIN and NBS1 had complementary functions in ATM signalling. ATMIN was required for ATM signalling by chloroquine and hypotonic stress, but not after induction of double-stand breaks by ionizing radiation (IR), whereas NBS1 is required for ATM signalling by IR. This suggested competition of NBS1 and ATMIN for ATM binding in a signal-dependent fashion. Some implications of these findings for the ATM signalling pathway are discussed.     

The Transmembrane Protein of the Human Endogenous Retrovirus - K (HERV-K) Modulates Cytokine Release and Gene Expression

Numerous copies of endogenous retroviruses are present in the genome of mammals including man. Although most of them are defective, some, e.g., the human endogenous retroviruses HERV-K, were found to be expressed under certain physiological conditions. For instance, HERV-K is expressed in germ cell tumours and melanomas as well as in the placenta. Most exogenous retroviruses including the human immunodeficiency virus HIV-1 induce severe immunodeficiencies and there is increasing evidence that the transmembrane envelope (TM) proteins of these retroviruses may be involved. We show here that HERV-K particles released from a human teratocarcinoma cell line, a recombinant TM protein and a peptide corresponding to a highly conserved so-called immunosuppressive domain in the TM protein of HERV-K inhibit the proliferation of human immune cells, induce modulation of the expression of numerous cytokines, and modulate the expression of cellular genes as detected by a microarray analysis. The changes in cytokine release and gene expression induced by the TM protein of HERV-K are similar to those found previously induced by the TM protein of HIV-1. These data suggest that the mechanism of immunosuppression may be similar for different retroviruses and that the expression of the TM protein in tumours and in the placenta may suppress immune responses and thus prevent rejection of the tumour and the embryo.     

New Potential Cell Wall Glucanases of Saccharomyces cerevisiae and Their Involvement in Mating

Biotinylation of intact Saccharomyces cerevisiae cells with a nonpermeant reagent (Sulfo-NHS-LC-Biotin) allowed the identification of seven cell wall proteins that were released from intact cells by dithiothreitol (DTT). By N-terminal sequencing, three of these proteins were identified as the known proteins β-exoglucanase 1 (Exg1p), β-endoglucanase (Bgl2p), and chitinase (Cts1p). One protein was related to the PIR protein family, whereas the remaining three (Scw3p, Scw4p, and Scw10p [for soluble cell wall proteins]) were found to be related to glucanases. Single knockouts of these three potential glucanases did not result in dramatic phenotypes. The double knockout of SCW4 and the homologous gene SCW10 resulted in slower growth, significantly increased release of proteins from intact cells by DTT, and highly decreased mating efficiency when these two genes were disrupted in both mating types. The synergistic behavior of the disruption of SCW4 and SCW10 was partly antagonized by the disruption of BGL2. The data are discussed in terms of a possible counterplay of transglucosidase and glucosidase activities.     

Sex differences in aging,life span and spontaneous tumorigenesis in 129/Sv mice neonatally exposed to metformin

The perinatal (prenatal and early neonatal) period is a critical stage for hypothalamic programming of sexual differentiation as well as for the development of energy and metabolic homeostasis. We hypothesized that neonatal treatment with antidiabetic drug biguanide metformin would positively modify regulation of growth hormone – IGF-1 – insulin signaling pathway slowing down aging and improving cancer preventive patterns in rodents. To test this hypothesis male and female 129/Sv mice were s.c. injected with metformin (100 mg/kg) at the 3rd, 5th and 7th days after birth. Metformin-treated males consumed less food and water and their body weight was decreased as compared with control mice practically over their entire lifespan. There were no significant differences in age-related dynamics of food and water consumption in females and they were heavier than controls. The fraction of mice with regular estrous cycles decreased with age and demonstrated a tendency to decrease in the females neonatally treated with metformin. Neonatal exposure to metformin practically failed to change the extent of hormonal and metabolic parameters in blood serum of male and female mice. In males, neonatal metformin treatment significantly increased the mean life span (+20%, P < 0.05) and slightly increased the maximum life span (+3.5%). In females, the mean life span and median in metformin-treated groups were slightly decreased (−9.1% and −13.8% respectively, P > 0.05) in comparison to controls, whereas mean life span of last 10% survivors and maximum life span were the same as in controls. Almost half (45%) of control male mice and 71.8% male mice neonatally exposed to metformin survived up to 800 d of age, the same age was achieved by 54.3% of mice in control female group and 30% of metformin-treated females (P < 0.03). Thus, neonatal metformin exposure slows down aging and prolongs lifespan in male but not in female mice.     

Morphometric differentiation of Pseudocalanus minutus populations in the Barents Sea

Dvoretsky VG. and Dvoretsky AG. 2011. Morphometric differentiation of Pseudocalanus minutus populations in the Barents Sea. —Acta Zoologica (Stockholm) 00 : 1–12. We investigated spatial variations in the morphometric characteristics (total length of body, lengths of cephalothorax, abdomen and antennules, and their relative proportions) of Pseudocalanus minutus, an abundant copepod species across the Barents Sea in August–September 2007. Females were found to have higher values for the measured parameters than males. The average absolute morphometric characters of both sexes increased from the south to the north. In most cases, parameters were similar in the southern, central, and eastern regions delineated by cluster analyses of oceanographic variables. The morphometric characteristics were strongly correlated with environmental variables in both males and females. Multiple regression analysis showed that temperature together with salinity explained 72–85% (in females) and 38–91% (in males) of the total variations in the log₁₀‐transformed morphometric parameters. According to principal component analysis and discriminant function analysis, two distinct groups could be separated in the Barents Sea. The first group included the copepods from the northern region; the other included populations from the southern, central, and eastern regions. The observed morphological variation can be interpreted as geographical variation connected with hydrological variability.     

Adsorption of nucleosides on mercury surface

Summary All nucleosides currently occurring in nucleic acids are, under the given experimental conditions, surface-active substances. If the concentration is sufficiently high, the most of nucleosides in the electrode surface associates and creates a surface film just as the corresponding bases do. Only with uridine, thymidine, and cytidine, unlike the corresponding bases, no associations in the electrode surface occur.