Spermatozoal sensitive biomarkers to defective protaminosis and fragmented DNA

Background

Isg15 covalently modifies murine endometrial proteins in response to early pregnancy. Isg15 can also be severed from targeted proteins by a specific protease called Ubp43 (Usp18). Mice lacking Ubp43 (null) form increased conjugated Isg15 in response to interferon. The Isg15 system has not been examined in chorioallantoic placenta (CP) or mesometrial (MM) components of implantation sites beyond 9.5 days post coitum (dpc). It was hypothesized that deletion of Ubp43 would cause disregulation of Isg15 in implantation sites, and that this would affect pregnancy rates.

Methods

Heterozygous (het) Ubp43 mice were mated and MM and CP implantation sites were collected on 12.5 and 17.5 days post-coitum (dpc).

Results

Free and conjugated Isg15 were greater on 12.5 versus 17.5 dpc in MM. Free and conjugated Isg15 were also present in CP, but did not differ due to genotype on 12.5 dpc. However, null CP had greater free and conjugated Isg15 when compared to het/wt on 17.5 dpc. Null progeny died in utero with fetal genotype ratios (wt:het:null) of 2:5:1 on 12.5 and 2:2:1 on 17.5 dpc. Implantation sites were disrupted within the junctional zone and spongiotrophoblast, contained less vasculature based on lectin B4 staining and contained greater Isg15 mRNA and VEGF protein in Ubp43 null when compared to wt placenta.

Conclusion

It is concluded that Isg15 and its conjugates are present in implantation sites during mid to late gestation and that deletion of Ubp43 causes an increase in free and conjugated Isg15 at the feto-maternal interface. Also, under mixed genetic background, deletion of Ubp43 results in fetal death.     

Yif1B Is Involved in the Anterograde Traffic Pathway and the Golgi Architecture

Yif1B is an intracellular membrane‐bound protein belonging to the Yip family, shown previously to control serotonin 5‐HT_1A receptor targeting to dendrites. Because some Yip proteins are involved in the intracellular traffic between the ER and the Golgi, here we investigated the precise localization of Yif1B in HeLa cells. We found that Yif1B is not resident into the Golgi, but rather belongs to the IC compartment. After analyzing the role of Yif1B in protein transport, we showed that the traffic of the VSVG protein marker was accelerated in Yif1B depleted HeLa cells, as well as in hippocampal neurons from Yif1B KO mice. Conversely, Yif1B depletion in HeLa cells did not change the retrograde traffic of ShTx. Interestingly, in long term depletion of Yif1B as in Yif1B KO mice, we observed a disorganized Golgi architecture in CA1 pyramidal hippocampal neurons, which was confirmed by electron microscopy. However, because short term depletion of Yif1B did not change Golgi structure, it is likely that the implication of Yif1B in anterograde traffic does not rely on its role in structural organization of the Golgi apparatus, but rather on its shuttling between the ER, the IC and the Golgi compartments.      

Physiological responses of two wheat cultivars to soil drought

Young plants of the two wheat cultivars Katya and Prelom, differing in their reaction to drought in the field, were grown in soil in pots, and their water status was assessed as well as the intensity of gas exchange, chlorophyll fluorescence, and accumulation of compatible solutes and hydrogen peroxide after 7 days of dehydration. It was established that cv. Katya displayed markedly better tolerance to soil drying in comparison with cv. Prelom. This was partly due to the more effective control of water balance, activity of the photosynthetic apparatus, and metabolic activity of leaves under stress. Consequently, lower amounts of hydrogen peroxide were accumulated and a lower membrane injury index was determined.     

Lipid raft targeting of hematopoietic protein tyrosine phosphatase by protein kinase C theta-mediated phosphorylation

Protein kinase C theta (PKC theta) is unique among PKC isozymes in its translocation to the center of the immune synapse in T cells and its unique downstream signaling. Here we show that the hematopoietic protein tyrosine phosphatase (HePTP) also accumulates in the immune synapse in a PKC theta-dependent manner upon antigen recognition by T cells and is phosphorylated by PKC theta at Ser-225, which is required for lipid raft translocation. Immune synapse translocation was completely absent in antigen-specific T cells from PKC theta-/- mice. In intact T cells, HePTP-S225A enhanced T-cell receptor (TCR)-induced NFAT/AP-1 transactivation, while the acidic substitution mutant was as efficient as wild-type HePTP. We conclude that HePTP is phosphorylated in the immune synapse by PKC theta and thereby targeted to lipid rafts to temper TCR signaling. This represents a novel mechanism for the active immune synapse recruitment and activation of a phosphatase in TCR signaling.     

Molecular dissection of the prototype foamy virus (PFV) RNA 5′-UTR identifies essential elements of a ribosomal shunt

The prototype foamy virus (PFV) is a nonpathogenic retrovirus that shows promise as a vector for gene transfer. The PFV (pre)genomic RNA starts with a long complex leader that can be folded into an elongated hairpin, suggesting an alternative strategy to cap-dependent linear scanning for translation initiation of the downstream GAG open reading frame (ORF). We found that the PFV leader carries several short ORFs (sORFs), with the three 5′-proximal sORFs located upstream of a structural element. Scanning-inhibitory hairpin insertion analysis suggested a ribosomal shunt mechanism, whereby ribosomes start scanning at the leader 5′-end and initiate at the downstream ORF via bypass of the central leader regions, which are inhibitory for scanning. We show that the efficiency of shunting depends strongly on the stability of the structural element located downstream of either sORFs A/A′ or sORF B, and on the translation event at the corresponding 5′-proximal sORF. The PFV shunting strategy mirrors that of Cauliflower mosaic virus in plants; however, in mammals shunting can operate in the presence of a less stable structural element, although it is greatly improved by increasing the number of base pairings. At least one shunt configuration was found in primate FV (pre)genomic RNAs.     

The effects of a twenty-four-week aquatic training program on muscular strength performance in healthy elderly women

The purpose of the present study was to determine the effectiveness of a 24-week aquatic training (AT) program, which included both aerobic and resistance components, on muscle strength (isometric and dynamic), flexibility, and functional mobility in healthy women over 60 years of age. Twenty-two subjects were assigned randomly to either an AT (n = 12) or a control (C, n = 10) group. Volunteers participated in a supervised shallow-water exercise program for 60 minutes a day, 3 days a week; the exercise program consisted of a 10-minute warm-up and stretching, 25 minutes of endurance-type exercise (dancing) at 80% of heart rate (HR)(max), 20 minutes of upper- and lower-body resistance exercises with specialized water-resistance equipment, and a 5-minute cool down. Maximal isometric torque of knee extensors (KEXT) and knee flexors (KFLEX) were evaluated by a Cybex Norm dynamometer, grip strength (HGR) was evaluated using a Jamar hydraulic dynamometer, and dynamic strength was evaluated via the 3 repetition maximum (3RM) test for chest press, knee extension, lat pull down, and leg press. Jumping performance was evaluated using the squat jump (SJ), functional mobility with the timed up-and-go (TUG) test, and trunk flexion with the sit-and-reach test. Body composition was measured using the bioelectrical impedance method. The AT induced significant improvements in KEXT (10.5%) and KFLEX (13.4%) peak torque, HGR strength (13%), 3RM (25.7-29.4%), SJ (24.6%), sit-and-reach (11.6%), and TUG (19.8%) performance. The AT group demonstrated a significant increase in lean body mass (3.4%). No significant changes in these variables were observed in the C group. The results indicate that AT, with both aerobic and resistance components, is an alternative training method for improving neuromuscular and functional fitness performance in healthy elderly women.     

X-ray crystallography and solution NMR spectroscopy characterization of heptakis(2,3-di-O-acetyl-6-bromo-6-deoxy)cyclomaltoheptaose

Heptakis(2,3-di-O-acetyl-6-bromo-6-deoxy)cyclomaltoheptaose has been characterized in aqueous solution by 1D and 2D NMR spectroscopy and in the solid state by X-ray crystallography. In methanol solution, the acetyl groups were found to interact with both inward and outward-pointing protons. This and the strong deshielding of the bridging carbons, relative to the nonacetylated precursor, indicate macrocyclic flexibility. In the crystalline state the macrocycle exists as a methanol complex. It exhibits elliptical distortion, all glucose residues been tilted with their primary side toward the cavity. The existing strain due to the congestion of 14 acetyl groups at the secondary site is relieved by two glucose rings acquiring the rarely observed skew-boat conformation, (0)S(2), by the increased tilting of two glucose residues, as well as by minor variations of the torsion angles of the acetyl groups. The seven bromine atoms are quite accessible to nucleophiles.     

The size of the synaptic cleft and distinct distributions of filamentous actin,ezrin, CD43, and CD45 at activating and inhibitory human NK cell immune synapses

In this study, we report the organization of cytoskeletal and large transmembrane proteins at the inhibitory and activating NK cell immunological or immune synapse (IS). Filamentous actin accumulates at the activating, but not the inhibitory, NK cell IS. However, surprisingly, ezrin and the associated protein CD43 are excluded from the inhibitory, but not the activating, NK cell IS. This distribution of ezrin and CD43 at the inhibitory NK cell IS is similar to that previously seen at the activating T cell IS. CD45 is also excluded from the inhibitory, but not activating, NK cell IS. In addition, electron microscopy reveals wide and narrow domains across the synaptic cleft. Target cell HLA-C, located by immunogold labeling, clusters where the synaptic cleft spans the size of HLA-C bound to the inhibitory killer Ig-like receptor. These data are consistent with assembly of the NK cell IS involving a combination of cytoskeletal-driven mechanisms and thermodynamics favoring the organization of receptor/ligand pairs according to the size of their extracellular domains.