Targeting Protein for Xenopus Kinesin-like Protein 2 (TPX2) Regulates γ-Histone 2AX (γ-H2AX) Levels upon Ionizing Radiation

The microtubule-associated protein targeting protein for Xenopus kinesin-like protein 2 (TPX2) plays a key role in spindle assembly and is required for mitosis in human cells. In interphase, TPX2 is actively imported into the nucleus to prevent its premature activity in microtubule organization. To date, no function has been assigned to nuclear TPX2. We now report that TPX2 plays a role in the cellular response to DNA double strand breaks induced by ionizing radiation. Loss of TPX2 leads to inordinately strong and transient accumulation of ionizing radiation-dependent Ser-139-phosphorylated Histone 2AX (γ-H2AX) at G₀ and G₁ phases of the cell cycle. This is accompanied by the formation of increased numbers of high intensity γ-H2AX ionizing radiation-induced foci. Conversely, cells overexpressing TPX2 have reduced levels of γ-H2AX after ionizing radiation. Consistent with a role for TPX2 in the DNA damage response, we found that the protein accumulates at DNA double strand breaks and associates with the mediator of DNA damage checkpoint 1 (MDC1) and the ataxia telangiectasia mutated (ATM) kinase, both key regulators of γ-H2AX amplification. Pharmacologic inhibition or depletion of ATM or MDC1, but not of DNA-dependent protein kinase (DNA-PK), antagonizes the γ-H2AX phenotype caused by TPX2 depletion. Importantly, the regulation of γ-H2AX signals by TPX2 is not associated with apoptosis or the mitotic functions of TPX2. In sum, our study identifies a novel and the first nuclear function for TPX2 in the cellular responses to DNA damage.     

Residual force enhancement during voluntary contractions of knee extensors and flexors at short and long muscle lengths

Residual force enhancement (RFE) is a term describing the observation that muscle tension during a contraction that includes a stretch and hold remains above that during an isometric contraction at the hold length. RFE has been observed during in vitro and in vivo experiments, but results involving voluntary contractions are mixed, particularly with respect to large muscles. The purpose of this study was to determine if RFE can be observed in large muscles such as knee extensors and flexors at joint configurations corresponding to the ascending and descending limbs of the muscle force-length curve. Two groups of twenty participants (ten males and ten females per group) performed maximum voluntary contractions on a Biodex machine in purely isometric conditions and in isometric conditions immediately following eccentric stretch. Knee extension trials were performed at 40° (short muscles) and 100° (long muscles) flexion from full extension (0°), and knee flexion trials were performed at 70° (short muscles) and 10° (long muscles) flexion. Stretch-isometric trials terminated at these angles following 30° of eccentric motion at 30°/s. Statistically-significant RFE was observed for both tasks at long-muscle joint configurations, but was not observed for either task at short-muscle joint configurations. Passive torque enhancement was also observed following muscle relaxation at long-muscle joint configurations for both tasks, but for only knee flexion at short-muscle joint configurations. These results reinforce for voluntary contractions of large muscles the RFE behavior observed in smaller muscles, and provide further evidence that RFE occurs primarily on the descending limb of the muscle force-length curve.     

Tactile feedback plays a critical role in maximum finger force production

This study investigates the role of cutaneous feedback on maximum voluntary force (MVF), finger force deficit (FD) and finger independence (FI). FD was calculated as the difference between the sum of maximal individual finger forces during single-finger pressing tasks and the maximal force produced by those fingers during an all-finger pressing task. FI was calculated as the average non-task finger forces normalized by the task-finger forces and subtracted from 100 percent. Twenty young healthy right-handed males participated in the study. Cutaneous feedback was removed by administering ring block digital anesthesia on the 2nd, 3rd, 4th and 5th digits of the right hands. Subjects were asked to press force sensors with maximal effort using individual digits as well as all four digits together, with and without cutaneous feedback. Results from the study showed a 25% decrease in MVF for the individual fingers as well as all the four fingers pressing together after the removal of cutaneous feedback. Additionally, more than 100% increase in FD after the removal of cutaneous feedback was observed in the middle and ring fingers. No changes in FI values were observed between the two conditions. Results of this study suggest that the central nervous system utilizes cutaneous feedback and the feedback mechanism plays a critical role in maximal voluntary force production by the hand digits.     

Draft Genome Sequence of Klebsiella pneumoniae subsp. pneumoniae DSM 30104T

Klebsiella pneumoniae is a Gram-negative, rod-shaped, nonmotile, and opportunistic pathogenic species with clinical importance. It is a part of natural flora of humans and animals. Here we report the draft genome sequence of the type strain of Klebsiella pneumoniae subsp. pneumoniae (DSM 30104^T) to provide taxonomic and functional insights into the species.     

Label-free detection of kanamycin based on the aptamer-functionalized conducting polymer/gold nanocomposite

Highly sensitive label-free detection of kanamycin is achieved with an aptamer sensor based on a conducting polymer/gold self-assembled nanocomposite. The sensor probe is fabricated by covalently immobilizing an in vitro selected DNA aptamer for kanamycin onto gold nanoparticle (AuNP)-comprised conducting polymer, poly-[2, 5-di-(2-thienyl)-1H-pyrrole-1-(p-benzoic acid)] (poly-DPB). The self-assembling of DPB on AuNP is investigated by TEM and UV-vis spectroscopy and the modification of the aptamer sensor is characterized using XPS and electrochemical impedance spectroscopy. The probe is applied to detect kanamycin by using voltammetric techniques. The sensor shows a pair of redox peaks around 0.26/ 0.08 V (vs. Ag/AgCl) for kanamycin captured by the aptamer-immobilized probe. The parameters that can affect the response, such as aptamer concentration, incubation time, temperature, and pH are optimized. The calibration plot shows a linear range from 0.05 μM to 9.0 μM kanamycin with a detection limit of 9.4±0.4 nM. The proposed aptamer sensor is examined with a real sample.     

A novel method of transgene delivery into triticale plants using the Agrobacterium transferred DNA-derived nano-complex

Genetic transformation of monocotyledonous plants still presents a challenge for plant biologists and biotechnologists because monocots are difficult to transform with Agrobacterium tumefaciens, whereas other transgenesis methods, such as gold particle-mediated transformation, result in poor transgene expression because of integration of truncated DNA molecules. We developed a method of transgene delivery into monocots. This method relies on the use of an in vitro-prepared nano-complex consisting of transferred DNA, virulence protein D2, and recombination protein A delivered to triticale microspores with the help of a Tat2 cell-penetrating peptide. We showed that this approach allowed for single transgene copy integration events and prevented degradation of delivered DNA, thus leading to the integration of intact copies of the transgene into the genome of triticale plants. This resulted in transgene expression in all transgenic plants regenerated from microspores transfected with the full transferred DNA/protein complex. This approach can easily substitute the bombardment technique currently used for monocots and will be highly valuable for plant biology and biotechnology.     

Divergence of flowering genes in soybean

Soybean genome sequences were blasted with Arabidopsis thaliana regulatory genes involved in photoperiod-dependent flowering. This approach enabled the identification of 118 genes involved in the flowering pathway. Two genome sequences of cultivated (Williams 82) and wild (IT182932) soybeans were employed to survey functional DNA variations in the flowering-related homologs. Forty genes exhibiting nonsynonymous substitutions between G. max and G. soja were catalogued. In addition, 22 genes were found to co-localize with QTLs for six traits including flowering time, first flower, pod maturity, beginning of pod, reproductive period, and seed filling period. Among the genes overlapping the QTL regions, two LHY/CCA1 genes, GI and SFR6 contained amino acid changes. The recently duplicated sequence regions of the soybean genome were used as additional criteria for the speculation of the putative function of the homologs. Two duplicated regions showed redundancy of both flowering-related genes and QTLs. ID 12398025, which contains the homeologous regions between chr 7 and chr 16, was redundant for the LHY/CCA1 and SPA1 homologs and the QTLs. Retaining of the CRY1 gene and the pod maturity QTLs were observed in the duplicated region of ID 23546507 on chr 4 and chr 6. Functional DNA variation of the LHY/CCA1 gene (Glyma07g05410) was present in a counterpart of the duplicated region on chr 7, while the gene (Glyma16g01980) present in the other portion of the duplicated region on chr 16 did not show a functional sequence change. The gene list catalogued in this study provides primary insight for understanding the regulation of flowering time and maturity in soybean.     

TPA-induced cell transformation provokes a complex formation between Pin1 and 90?kDa ribosomal protein S6 kinase 2

The antioxidant properties of organoselenium compounds have been extensively investigated because oxidative stress is a hallmark of a variety of chronic human diseases. Here, we reported the influence of substituent groups in the antioxidant activity of β-selenoamines. We have investigated whether they exhibited glutathione peroxidase-like (GPx-like) activity and whether they could be substrate of thioredoxin reductase (TrxR). In the DPPH assay, the β-selenium amines did not exhibit antioxidant activity. However, the β-selenium amines with p-methoxy and tosyl groups prevented the lipid peroxidation. The β-selenium amine compound with p-methoxy substituent group exhibited thiol-peroxidase-like activity (GPx-like activity) and was reduced by the hepatic TrxR. These results contribute to understand the influence of structural alteration of non-conventional selenium compounds as synthetic mimetic of antioxidant enzymes of mammalian organisms.     

Host–parasite interactions and host species susceptibility of the marine oomycete parasite, Olpidiopsis sp., from Korea that infects red algae

Porphyra farms in Korea occasionally suffer from Olpidiopsis infection. As Porphyra farming proceeds from October to March, this obligatory biotrophic parasite may need an alternative host to survive during other months of the year. To find a possible alternative summer host, we collected algae from Wando, Korea, where extensive Porphyra plantations are located, and discovered an oomycete assignable to the genus Olpidiopsis from Heterosiphonia pulchra. Host susceptibility tests showed that this oomycete could also infect Heterosiphonia japonica, Dasya sp., Dasysiphonia chejuensis, and also blades of Porphyra tenera. The minimum incubation time for this Olpidiopsis sp. to infect its hosts was approximately 4 h. Zoosporangia matured in 2 days and biflagellate zoospores were released. Free zoospores remained infective in seawater for up to 7 days. The infection of Olpidiopsis sp. to H. japonica was cell-type specific and extended rhizoid-like apical cells of determinate branches were preferentially infected. FITC-conjugated lectin staining showed specific binding of concanavalin A (ConA) to extended rhizoid-like apical cells. Attachment of Olpidiopsis sp. zoospores to the host cells was inhibited by α-mannosidase. Monosaccharide inhibition experiments showed that d(+)-mannose, complementary to the lectin ConA, could also block the infection, suggesting a lectin–carbohydrate interaction during host–parasite recognition.     

Cardioprotective effects of 3-phosphoinositide-dependent protein kinase-1 on hypoxic injury in cultured neonatal rat cardiomyocytes and myocardium in a rat myocardial infarct model

3-Phosphoinositide-dependent protein kinase-1 (PDK1) is involved in numerous cellular responses. In this study, we investigated the protective effects of PDK1 gene expression against hypoxic conditions in cultured rat CMCs (rCMCs) and in a rat myocardial infarction (MI) model using the lentiviral vector (LeV) system. LeV-PDK1 transfer effectively reduced the apoptotic cell death caused by hypoxic injury as compared to LeV-GFP transfer in rCMCs the expression of survival proteins increased in the LeV-PDK1 group, whereas apoptosis signaling decreased in the rCMCs and in infarcted hearts treated with LeV-PDK1. LeV-PDK1 transfer also reduced apoptosis and infarct size and attenuated myocardial wall thinning and ventricular remodeling in a rat MI model. These findings suggest that PDK1 has a protective role in the injured ischemic myocardium via overexpression of the cell survival pathway in CMCs. Hence PDK1 can be used as a treatment strategy for myocardial salvage inin hypoxic injury.