A convenient preparation of N ε-methyl-l-lysine derivatives and its application to the synthesis of histone tail peptides

A convenient route is established for the preparation of N ^α-Fmoc-N ^ε-(Boc, methyl)-l-lysine and N ^α-Fmoc-N ^ε-dimethyl-l-lysine as building blocks to be used for the synthesis of methylated peptides. This methodology is based on the use of malonate derivatives and dibromobutane to produce key intermediates, l-2-amino-6-bromohexanoic acid derivatives, which could be modified to the required group at the ε-position. Fmoc-protection is accessible, so these compounds can be used in solution as well as in solid-phase peptide synthesis. Also the peptides containing these methylated lysines have been proved to resist the action of trypsin and lysyl endopeptidase. Thus, this new method could be considered as an improvement of the synthesis of N ^ε-methyl-l-lysine derivatives.     

Design and synthesis of mono and bicyclic tetrapeptides thioester as potent inhibitor of histone deacetylases

Inhibitors of histone deacetylases (HDACs) are a promising class of anticancer agents that have an effect on gene regulation. The naturally occurring cyclic depsipeptide FK228 containing disulfide and Largazole possessing thioester functionalities act as pro-drugs and share the same HDAC inhibition mechanism in cell. Inspired from these facts, we have reported bicyclic tetrapeptide disulfide HDAC inhibitors resembling FK228 with potent activity and enhanced selectivity. In the present study, we report the design and synthesis of several mono and bicyclic tetrapeptide thioester HDAC inhibitors that share the inhibition mechanism similar to Largazole. Most of the compounds showed HDAC1 and HDAC4 inhibition and p21 promoting activity in nanomolar ranges. Among these the monocyclic peptides 1, 2 and bicyclic peptide, 4 are notable demanding more advanced research to be promising anticancer drug candidates.     

Life history of Juniperus sabina L. adapted to the sand shifting environment in the Mu Us Sandy Land,China: A review

We have reviewed publications on the physiological and ecological features of the growth and regeneration processes of Juniperus sabina L. which grows in semiarid sandy land in the Ordos plateau in northern China where desertification has progressed over time. J. sabina is a key native plant species used for ecological restoration in this region. The life history of J. sabina in this sandy land that has been revealed through this review includes several unique features: (1) both vegetative and seed propagations are observed, but seed propagation is not successful in the location where the mature J. sabina stands. Instead, seed propagation can occur at a different place with different landscapes from the mature stands. (2) Nurse plants play an important role in providing the microclimatic environment necessary for the growth of J. sabina seedlings and young plants. (3) During the horizontal and vertical growth processes of the J. sabina patch, the root system was affected by burial in shifting sand and consequently acquired greater access to the water supply in deeper soil horizons, which could support larger growth. These characteristics suggested that the regeneration by seed propagation and growth strategy of J. sabina in this region was strongly affected by sand movement and the landscape that is generated by sand movement.

     

Effects of livestock grazing intensity on soil arbuscular mycorrhizal fungi and glomalin-related soil protein in a mountain forest steppe and a desert steppe of Mongolia

Arbuscular mycorrhizal fungi (AMF) are important components of the grassland ecosystems in terms of plant phosphorus uptake and accumulation of glomalin-related soil protein (GRSP). Though Mongolian grasslands are seriously degraded by livestock grazing, the effects of grazing on soil AMF and GRSP remain unclear. Here, we examined community composition and diversity of AMF as well as amount of GRSP at three different grazing intensities: lightly grazed (LG), moderately grazed (MG), and heavily grazed (HG) under two different types of grassland, mountain forest steppe at Hustai and desert steppe at Mandalgobi. The diversity and biomass of AMF-host and non-AMF plants strongly affected the overall AMF community composition and its diversity. When we separately analyzed the factors affecting soil AMF diversity at Hustai and Mandalgobi, decrease in the shoot biomass of Poaceae plants at Hustai and decreases in the species number and shoot biomass of AMF-host plants at Mandalgobi were significantly correlated with AMF diversity. GRSP decreased with increasing grazing intensity, which was significantly correlated with soil pH and total root biomass at Hustai. The decrease in plant biomass caused by grazing thus led to GRSP reduction. Our results showed that change in soil AMF community caused by livestock grazing were associated with change in the biomass and diversity of functional vegetation groups such as Poaeceae, AMF-host and non-AMF plants, indicating the importance to focus on such functional vegetation groups to evaluate the effect of grazing on AMF.

     

Kinetic analysis of the entire RNA amplification process by Qbeta replicase

The kinetics of the RNA replication reaction by Qbeta replicase were investigated. Qbeta replicase is an RNA-dependent RNA polymerase responsible for replicating the RNA genome of coliphage Qbeta and plays a key role in the life cycle of the Qbeta phage. Although the RNA replication reaction using this enzyme has long been studied, a kinetic model that can describe the entire RNA amplification process has yet to be determined. In this study, we propose a kinetic model that is able to account for the entire RNA amplification process. The key to our proposed kinetic model is the consideration of nonproductive binding (i.e. binding of an enzyme to the RNA where the enzyme cannot initiate the reaction). By considering nonproductive binding and the notable enzyme inactivation we observed, the previous observations that remained unresolved could also be explained. Moreover, based on the kinetic model and the experimental results, we determined rate and equilibrium constants using template RNAs of various lengths. The proposed model and the obtained constants provide important information both for understanding the basis of Qbeta phage amplification and the applications using Qbeta replicase.     

Evaluating the relationships between the postural adaptation of patients with profound cerebral palsy and the configuration of the Seating Buggy's seating support surface

We are currently investigating the physiological polymorphism of wheelchair users with profound cerebral palsy and the properties of the Seating Buggy (developed by S. Nishimura, 1998) to clarify important and general elements of wheelchairs for widespread use. Cerebral palsy is a diagnostic term used to describe a group of motor syndromes resulting from disorders in early brain development. Recently, it has been shown that the Seating Buggy produces functional head-neck alignments and active control of sitting balance for people with profound cerebral palsy. The Seating Buggy is a wheelchair for the profoundly disabled and features a wide adjustment range from heights of 120 cm to 175 cm. Its seating support surface is comprised of a sling-seat. To examine the relationships between the postural adaptation of patients with profound cerebral palsy and the configuration of the Seating Buggy's seating, we assessed the postural alignment of the Seating Buggy's user and then measured the configuration of its resulting seating support surface with a three dimensional scanning system. Twenty-one subjects were used for the purposes of this investigation in their everyday environment. Postural adaptation and wheelchair fitting in the Seating Buggy were assessed from the viewpoint of the Active Balanced Seating by a seating expert. The subjects fell into two categories, as follows: 11 for appropriate or nearly appropriate fitting, and 10 for ill-fitting. The depth of thoracic support and the forward distance of lumbar support for those who claimed that it was ill-fitting were significantly reduced compared with that of those who claimed that the Seating Buggy offered an appropriate or nearly appropriate fitting. It was suggested that the properly adjusted depth of thoracic support and distance of the lumbar support were related to the resulting satisfactory head-neck alignment and sitting balance of the patients with profound cerebral palsy.     

Development of a reporter peptide that catalytically produces a fluorescent signal through α-complementation

In α-complementation, inactive N-terminal (α-domain) and C-terminal (ω-domain) fragments of β-galactosidase associate to reconstitute the active protein. To date, the effect of α-domain size on α-complementation activity has not been systematically investigated. In this study, we compared the complementation activities of α-domains of various sizes using an in vitro system. We found that the complementation activities are similar for α-domains comprising between 45 and 229 N-terminal residues but are significantly decreased for those containing less than 37 residues. However, these smaller α-domains (15 and 25 residues) exhibited sufficient α-complementation activity for application as reporters.     

A New Therapeutic Assessment Score for Advanced Hepatocellular Carcinoma Patients Receiving Hepatic Arterial Infusion Chemotherapy

MethodsWe retrospectively analyzed 90 advanced HCC patients with elevated baseline alpha-fetoprotein (AFP) and/or des-gamma-carboxy prothrombin (DCP) levels and analyzed various parameters for their possible use as predictors of response and survival. AFP and DCP responses were assessed after half a course of HAIC (2 weeks); a positive-response was defined as a reduction of ≥ 20% from baseline.ResultsMultivariate analysis identified DCP response (odds ratio 16.03, p < 0.001) as an independent predictor of treatment response. In multivariate analysis, Child-Pugh class A (hazard ratio [HR] 1.99, p = 0.018), AFP response (HR 2.17, p = 0.007), and DCP response (HR 1.90, p = 0.030) were independent prognostic predictors. We developed an Assessment for Continuous Treatment with HAIC (ACTH) score, including the above 3 factors, which ranged from 0 to 3. Patients stratified into two groups according to this score showed significantly different prognoses (≤1 vs. ≥2 points: median survival time, 15.1 vs. 8.7 months; p = 0.003).ConclusionsThe ACTH score may be useful in the therapeutic assessment of HCC patients receiving HAIC.     

Phosphorylation of CHO1 by Lats1/2 regulates the centrosomal activation of LIMK1 during cytokinesis

Large tumor suppressor 1 and 2 (Lats1/2) regulate centrosomal integrity, chromosome segregation and cytokinesis. As components of the centralspindlin complex, the kinesin-like protein CHO1 and its splicing variant MKLP1 colocalize with chromosome passenger proteins and GTPases and regulate the formation of the contractile ring and cytokinesis; however, the regulatory mechanisms of CHO1/MKLP1 remain elusive. Here, we show that Lats1/2 phosphorylate Ser716 in the F-actin-interacting region of CHO1, which is absent in MKLP1. Phosphorylated CHO1 localized to the centrosomes and midbody, and the actin polymerization factor LIM-kinase 1 (LIMK1) was identified as its binding partner. Overexpression of constitutively phosphorylated and non-phosphorylated CHO1 altered the mitotic localization and activation of LIMK1 at the centrosomes in HeLa cells, leading to the inhibition of cytokinesis through excessive phosphorylation of Cofilin and mislocalization of Ect2. These results suggest that Lats1/2 stringently control cytokinesis by regulating CHO1 phosphorylation and the mitotic activation of LIMK1 on centrosomes.     

Rapid Exchange of Bound ADP on the Staphylococcus aureus Replication Initiation Protein DnaA

In Escherichia coli, regulatory inactivation of the replication initiator DnaA occurs after initiation as a result of hydrolysis of bound ATP to ADP, but it has been unknown how DnaA is controlled to coordinate cell growth and chromosomal replication in Gram-positive bacteria such as Staphylococcus aureus. This study examined the roles of ATP binding and its hydrolysis in the regulation of the S. aureus DnaA activity. In vitro, S. aureus DnaA melted S. aureus oriC in the presence of ATP but not ADP by a mechanism independent of ATP hydrolysis. Unlike E. coli DnaA, binding of ADP to S. aureus DnaA was unstable. As a result, at physiological concentrations of ATP, ADP bound to S. aureus DnaA was rapidly exchanged for ATP, thereby regenerating the ability of DnaA to form the open complex in vitro. Therefore, we examined whether formation of ADP-DnaA participates in suppression of replication initiation in vivo. Induction of the R318H mutant of the AAA+ sensor 2 protein, which has decreased intrinsic ATPase activity, caused over-initiation of chromosome replication in S. aureus, suggesting that formation of ADP-DnaA suppresses the initiation step in S. aureus. Together with the biochemical features of S. aureus DnaA, the weak ability to convert ATP-DnaA into ADP-DnaA and the instability of ADP-DnaA, these results suggest that there may be unidentified system(s) for reducing the cellular ratio of ATP-DnaA to ADP-DnaA in S. aureus and thereby delaying the re-initiation of DNA replication.