Hydroxylation of recombinant human collagen type I alpha 1 in transgenic maize co-expressed with a recombinant human prolyl 4-hydroxylase

<p>Abstract</p> <p>Background</p> <p>Collagens require the hydroxylation of proline (Pro) residues in their triple-helical domain repeating sequence Xaa-Pro-Gly to function properly as a main structural component of the extracellular matrix in animals at physiologically relevant conditions. The regioselective proline hydroxylation is catalyzed by a specific prolyl 4-hydroxylase (P4H) as a posttranslational processing step.</p> <p>Results</p> <p>A recombinant human collagen type I α-1 (rCIα1) with high percentage of hydroxylated prolines (Hyp) was produced in transgenic maize seeds when co-expressed with both the α- and β- subunits of a recombinant human P4H (rP4H). Germ-specific expression of rCIα1 using maize globulin-1 gene promoter resulted in an average yield of 12 mg/kg seed for the full-length rCIα1 in seeds without co-expression of rP4H and 4 mg/kg seed for the rCIα1 (rCIα1-OH) in seeds with co-expression of rP4H. High-resolution mass spectrometry (HRMS) analysis revealed that nearly half of the collagenous repeating triplets in rCIα1 isolated from rP4H co-expressing maize line had the Pro residues changed to Hyp residues. The HRMS analysis determined the Hyp content of maize-derived rCIα1-OH as 18.11%, which is comparable to the Hyp level of yeast-derived rCIα1-OH (17.47%) and the native human CIa1 (14.59%), respectively. The increased Hyp percentage was correlated with a markedly enhanced thermal stability of maize-derived rCIα1-OH when compared to the non-hydroxylated rCIα1.</p> <p>Conclusions</p> <p>This work shows that maize has potential to produce adequately modified exogenous proteins with mammalian-like post-translational modifications that may be require for their use as pharmaceutical and industrial products.</p

Mental Toughness in South African Youth: Relationships With Forgivingness and Attitudes Towards Risk

Young people are particularly vulnerable to health risk behaviors and interpersonal violence, stimulating scholars’ attention towards identifying factors that may reduce the likelihood that these actions will occur. Associated with positive outcomes in a variety of domains, mental toughness in young people might protect them from engaging in potentially deleterious interpersonal or health-risk behaviors, while potentially promoting positive psychological behaviors. Within this framework, the present study investigated the relationships between mental toughness, attitudes towards physical and psychological risk-taking, and trait forgiveness in a sample of 123 (males = 54, females = 69) South African youth (M age = 23.97 years, SD = 4.46). Univariate and multivariate analyses indicated higher levels of mental toughness were associated with being more forgiving, (η2pηp2 = .036), perceiving physical risk-taking more positively (η2pηp2 = .062), but having more negative attitudes towards psychological risk-taking (η2pηp2 = .036). These findings give credence to mental toughness as a psychological characteristic involved in youth risk-taking perceptions and interpersonal functioning. Future research might explore the integration of mental toughness into the development of future youth risk behavior interventions

Increased insolation threshold for runaway greenhouse processes on Earth like planets

Because the solar luminosity increases over geological timescales, Earth climate is expected to warm, increasing water evaporation which, in turn, enhances the atmospheric greenhouse effect. Above a certain critical insolation, this destabilizing greenhouse feedback can "runaway" until all the oceans are evaporated. Through increases in stratospheric humidity, warming may also cause oceans to escape to space before the runaway greenhouse occurs. The critical insolation thresholds for these processes, however, remain uncertain because they have so far been evaluated with unidimensional models that cannot account for the dynamical and cloud feedback effects that are key stabilizing features of Earth's climate. Here we use a 3D global climate model to show that the threshold for the runaway greenhouse is about 375 W/m$^2$, significantly higher than previously thought. Our model is specifically developed to quantify the climate response of Earth-like planets to increased insolation in hot and extremely moist atmospheres. In contrast with previous studies, we find that clouds have a destabilizing feedback on the long term warming. However, subsident, unsaturated regions created by the Hadley circulation have a stabilizing effect that is strong enough to defer the runaway greenhouse limit to higher insolation than inferred from 1D models. Furthermore, because of wavelength-dependent radiative effects, the stratosphere remains cold and dry enough to hamper atmospheric water escape, even at large fluxes. This has strong implications for Venus early water history and extends the size of the habitable zone around other stars.Comment: Published in Nature. Online publication date: December 12, 2013. Accepted version before journal editing and with Supplementary Informatio

Research workshop to research work: initial steps in establishing health research systems on Malaita, Solomon Islands

<p>Abstract</p> <p>Introduction</p> <p>Atoifi Adventist Hospital is a 90 bed general hospital in East Kwaio, Malaita, Solomon Islands providing services to the population of subsistence villagers of the region. Health professionals at the hospital and attached College of Nursing have considerable human capacity and willingness to undertake health research. However they are constrained by limited research experience, training opportunities, research systems, physical infrastructure and access to resources. This brief commentary describes an 'Introduction to Health Research' workshop delivered at Atoifi Adventist Hospital in September 2009 and efforts to move from 'research workshop' to 'research work'.</p> <p>The Approach</p> <p>Using a participatory-action research approach underpinned by decolonising methodologies, staff from Atoifi Adventist Hospital and James Cook University (Queensland, Australia) collaboratively designed, implemented and evaluated a health research workshop. Basic health research principles and methods were presented using active learning methodologies. Following the workshop, Atoifi Adventist Hospital and Atoifi College of Nursing staff, other professionals and community members reported an increased awareness and understanding of health research. The formation of a local Research Committee, improved ethics review procedures and the identification of local research mentors followed the week long workshop. The workshop has acted as a catalyst for research activity, increasing structural and human resource capacity for local health professionals and community leaders to engage in research.</p> <p>Discussion and Conclusions</p> <p>Participants from a variety of educational backgrounds participated in, and received benefit from, a responsive, culturally and linguistically accessible health research workshop. Improving health research systems at a remote hospital and aligning these with local and national research agendas is establishing a base to strengthen public health research and practice on Malaita, Solomon Islands.</p

Transgene-Induced Gene Silencing Is Not Affected by a Change in Ploidy Level

BACKGROUND: Whole genome duplication, which results in polyploidy, is a common feature of plant populations and a recurring event in the evolution of flowering plants. Polyploidy can result in changes to gene expression and epigenetic instability. Several epigenetic phenomena, occurring at the transcriptional or post-transcriptional level, have been documented in allopolyploids (polyploids derived from species hybrids) of Arabidopsis thaliana, yet findings in autopolyploids (polyploids derived from the duplication of the genome of a single species) are limited. Here, we tested the hypothesis that an increase in ploidy enhances transgene-induced post-transcriptional gene silencing using autopolyploids of A. thaliana. METHODOLOGY/PRINCIPAL FINDINGS: Diploid and tetraploid individuals of four independent homozygous transgenic lines of A. thaliana transformed with chalcone synthase (CHS) inverted repeat (hairpin) constructs were generated. For each line diploids and tetraploids were compared for efficiency in post-transcriptional silencing of the endogenous CHS gene. The four lines differed substantially in their silencing efficiency. Yet, diploid and tetraploid plants derived from these plants and containing therefore identical transgene insertions showed no difference in the efficiency silencing CHS as assayed by visual scoring, anthocyanin assays and quantification of CHS mRNA. CONCLUSIONS/SIGNIFICANCE: Our results in A. thaliana indicated that there is no effect of ploidy level on transgene-induced post-transcriptional gene silencing. Our findings that post-transcriptional mechanisms were equally effective in diploids and tetraploids supports the use of transgene-driven post-transcriptional gene silencing as a useful mechanism to modify gene expression in polyploid species

AXR1-ECR1 and AXL1-ECR1 heterodimeric RUB-activating enzymes diverge in function in Arabidopsis thaliana

RELATED TO UBIQUITIN (RUB) modification of CULLIN (CUL) subunits of the CUL-RING ubiquitin E3 ligase (CRL) superfamily regulates CRL ubiquitylation activity. RUB modification requires E1 and E2 enzymes that are analogous to, but distinct from, those activities required for UBIQUITIN (UBQ) attachment. Gene duplications are widespread in angiosperms, and in line with this observation, components of the RUB conjugation pathway are found in multiples in Arabidopsis. To further examine the extent of redundancy within the RUB pathway, we undertook biochemical and genetic characterizations of one such duplication event- the duplication of the genes encoding a subunit of the RUB E1 into AUXIN RESISTANT1 (AXR1) and AXR1-LIKE1 (AXL1). In vitro, the two proteins have similar abilities to function with E1 C-TERMINAL-RELATED1 (ECR1) in catalyzing RUB1 activation and RUB1-ECR1 thioester formation. Using mass spectrometry, endogenous AXR1 and AXL1 proteins were found in complex with 3HA-RUB1, suggesting that AXR1 and AXL1 exist in parallel RUB E1 complexes in Arabidopsis. In contrast, AXR1 and AXL1 differ in ability to correct phenotypic defects in axr1-30, a severe loss-of-function AXR1 mutant, when the respective coding sequences are expressed from the same promoter, suggesting differential in vivo functions. These results suggest that while both proteins function in the RUB pathway and are biochemically similar in RUB-ECR1 thioester formation, they are not functionally equivalent

Giant rafted pumice blocks from the most recent eruption of Taupo volcano, New Zealand: Insights from palaeomagnetic and textural data

Giant blocks of pumice lie strewn along a former shoreline of intracaldera Lake Taupo, New Zealand, and are the sole subaerial evidence of the most recent volcanism at the Taupo supervolcano. Geochemically they are identical to material erupted during the complex and multiphase 1.8 ka Taupo eruption, which they post-date by one to two decades. The blocks, some of which are >10 m long, show complex jointing patterns indicative of both surface chilling and continued interior expansion, as well as heterogeneous vesicularity, with dense rims (mean density 917 kg/m3) grading via an intervening transition zone (mean density 844 kg/m3) into a more highly vesicular interior (mean density 815 kg/m3). Analysis of thermal demagnetisation data indicates significant reorientation of the blocks as they cooled through a series of blocking temperatures. Some parts of block rims cooled to below 580 °C well before emplacement on the shore, whereas other parts in the interior and transition zones, which cooled more slowly, acquired different orientations before stranding. Some block interiors cooled after blocks were finally deposited, and record the direction of the 1.8 ka field. The blocks are believed to be derived from one or both of a pair of rhyolitic lava domes that developed on the bed of Lake Taupo several decades after the climactic Taupo eruption over the inferred vent area.These, and similar giant rafted pumice blocks in other marine and lacustrine settings raise a number of questions about how volatile-rich felsic magma can be erupted underwater with only limited thermal fragmentation. Furthermore, the prolonged flotation of out-sized fragments of vesiculated magma formed during subaqueous dome-growth contrasts with the rapid sinking of smaller pieces of hot plinian pumice under laboratory conditions. The genesis of pumice forming the blocks is not entirely clear. Most simply the blocks may represent part of a vesiculated carapace of a growing lava dome, broken loose as the dome grew and deformed then rising buoyantly to the surface. Parts of the carapace could also be released by local magma-water explosions. Some textures of the pumice, however, suggest fresher magma released from beneath the carapace. This may suggest that silicic dikes and pillows/pods intruded into a growing mound of silicic hyaloclastite, itself formed by quench fragmentation and thermal granulation of the dike margins. This fragmental cover would have inhibited cooling of a still-hot and actively vesiculating interior, which was then released to float to the surface by gravitational destabilisation and collapse of the growing pile. Following their formation, the large fragments of pumice floated to the lake's surface, where they were blown ashore to become embedded in accumulating transgressive shoreface sediments and continue cooling

Loss of susceptibility as a novel breeding strategy for durable and broad-spectrum resistance

Recent studies on plant immunity have suggested that a pathogen should suppress induced plant defense in order to infect a plant species, which otherwise would have been a nonhost to the pathogen. For this purpose, pathogens exploit effector molecules to interfere with different layers of plant defense responses. In this review, we summarize the latest findings on plant factors that are activated by pathogen effectors to suppress plant immunity. By looking from a different point of view into host and nonhost resistance, we propose a novel breeding strategy: disabling plant disease susceptibility genes (S-genes) to achieve durable and broad-spectrum resistance

Insulated molecular wires: inhibiting orthogonal contacts in metal complex based molecular junctions

Metal complexes are receiving increased attention as molecular wires in fundamental studies of the transport properties of metal|molecule|metal junctions. In this context we report the single-molecule conductance of a systematic series of d8 square-planar platinum(II) trans-bis(alkynyl) complexes with terminal trimethylsilylethynyl (C[triple bond, length as m-dash]CSiMe3) contacting groups, e.g. trans-Pt{C[triple bond, length as m-dash]CC6H4C[triple bond, length as m-dash]CSiMe3}2(PR3)2 (R = Ph or Et), using a combination of scanning tunneling microscopy (STM) experiments in solution and theoretical calculations using density functional theory and non-equilibrium Green's function formalism. The measured conductance values of the complexes (ca. 3–5 × 10−5G0) are commensurate with similarly structured all-organic oligo(phenylene ethynylene) and oligo(yne) compounds. Based on conductance and break-off distance data, we demonstrate that a PPh3 supporting ligand in the platinum complexes can provide an alternative contact point for the STM tip in the molecular junctions, orthogonal to the terminal C[triple bond, length as m-dash]CSiMe3 group. The attachment of hexyloxy side chains to the diethynylbenzene ligands, e.g. trans-Pt{C[triple bond, length as m-dash]CC6H2(Ohex)2C[triple bond, length as m-dash]CSiMe3}2(PPh3)2 (Ohex = OC6H13), hinders contact of the STM tip to the PPh3 groups and effectively insulates the molecule, allowing the conductance through the full length of the backbone to be reliably measured. The use of trialkylphosphine (PEt3), rather than triarylphosphine (PPh3), ancillary ligands at platinum also eliminates these orthogonal contacts. These results have significant implications for the future design of organometallic complexes for studies in molecular junctions