Ryanodine receptor phosphorylation in human heart failure

Heart failure is a complex disorder, characterized by activation of the sympathetic nervous system, leading to dysregulated Ca2+ homeostasis in cardiac myocytes and tissue remodeling. In a variety of diseases, cardiac malfunction is associated with aberrant fluxes of Ca2+ across both the surface membrane and the internal Ca2+ store, the sarcoplasmic reticulum (SR). One prominent hypothesis residues is that in heart failure, the activity of the ryanodine receptor (RyR2) Ca2+ release channel in the SR is increased due to excess phosphorylation and that this contributes to excess SR Ca2+ leak in diastole, reduced SR Ca2+ load and decreased contractility (Huke & Bers, 2008). There is controversy over which serine residues in RyR2 are hyperphosphorylated in animal models of heart failure and whether this is via the CaMKII or the PKA-linked signaling pathway. S2808, S2814 and S2030 in RyR2 have been variously claimed to be hyperphosphorylated. Our aim was to examine the degree of phosphorylation of these residues in RyR2 from failing human hearts. The use of human tissue was approved by the Human Research Ethics Committee, The Prince Charles Hospital, EC28114. Left ventricular tissue samples were obtained from an explanted heart of a patient with endstage heart failure (Emery Dreifuss Muscular Dystrophy with cardiomyopathy) and non-failing tissue was from a patient with cystic fibrosis undergoing heart-lung transplantation with no history of heart disease. SR vesicles were prepared as described by Laver et al. (1995) and examined with SDS-Page and Western Blot. Transferred proteins were probed with antibodies to detect total protein phosphorylation, phosphorylation of RyR2 serine residues S2808, S2814, S2030 and for the key proteins calsequestrin, triadin, junctin and FKBP12.6. To avoid membrane stripping artifact, each membrane was exposed to one phosphorylation-specific antibody and signal densities quantified using Bio-Rad Quantity One software. We found no distinguishable difference between failing and healthy hearts in the protein expression levels of RyR2, triadin, junctin or calsequestrin. We found an expected upregulation of total RyR2 phosphorylation in the failing heart sample, compared to a matched amount of RyR2 (quantified using densiometry) in healthy heart. Probing with antibodies detecting only the phosphorylated form of the specific RyR2 residues showed that the increase in total RyR2 phosphorylation in the failing heart was due to hyperphosphorylation of S2808 and S2814. We found that S2030 phosphorylation levels were unchanged in human heart failure. Interestingly, we found that S2030 has a basal level of phosphorylation in the healthy human heart, different from the absence of basal phosphorylation recently reported in rodent heart (Huke & Bers, 2008). Finally, preliminary results indicate that less FKBP 12.6 is associated with RyR2 in the failing heart, possibly as a consequence of PKA activation. In conclusion, residues S2808 and S2814 are hyperphosphorylated in human heart failure, presumably due to upregulation of the CaMKII and/or PKA signaling pathway as a result of chronic activation of the sympathetic nervous system. Such changes in RyR2 phosphorylation are believed to contribute to the leaky RyR2 phenotype associated with heart failure, which increases the incidence of arrhythmia and contributes to the severely impaired contractile performance of the failing heart. Huke S & Bers DM. (2008). Ryanodine receptor phosphorylation at serine 2030, 2808 and 2814 in rat cardiomyocytes. Biochemical and Biophysical Research Communications 376, 80-85. Laver DR, Roden LD, Ahern GP, Eager KR, Junankar PR & Dulhunty AF. (1995). Cytoplasmic Ca2+ inhibits the ryanodine receptor from cardiac muscle. Journal of Membrane Biology 147, 7-22. Proceedings

Revolutionary self portrait

'Revolutionary Self Portrait' (2009) is a sculptural self-portrait. The work comprises a bust engulfed in hair and a scalloped pedestal stand. Historically, divine energy has frequently been depicted using fluid forms - drapery, clouds and occasionally hair. These forms and associations act as a departure point for this sculpture in which the figure is depicted in a state of inundation by billowing tufts hair. The work was also inspired by the tendency of great 19th century utopian thinkers - for example Marx, Bakunin and Kropotkin - to wear large beards. Within both traditions, the language of heroic subjectivity is amplified by a sculptural extension of the body. In 'Revolutionary Self Portrait' however, this extension threatens to suffocate the subject - a gesture made all the more ironic due to the fact that the artist himself is incapable of growing a beard. The work was selected for the National Artists' Self Portrait Prize, University of Queensland Art Museum, 2009.

Seeing a hard country : Lawrence’s Australian landscape

In 1944 Australian author Eleanor Dark wrote that Australia is a hard country for an outsider to see, citing, in evidence, the writing of the “strange, foreign-looking little man with the beard” -- the self-described by D. H. Lawrence. According to Dark, Lawrence was bewildered by Australia because what his eyes saw was not what they were accustomed to seeing. Kangaroo, she wrote, suggests one long, tormented effort to see. Lawrence appears, for Dark, to be half-blind, struggling, and irritated almost beyond belief with his visit to New South Wales. Eleanor Dark wrote this critique in 1944, long after Lawrence’s 1922 visit, but for her, as for other Australian writers, Kangaroo continued to register as an important book, even if the content rankled. Katharine Susannah Prichard and Christina Stead, both advocates in general of Lawrence, likewise rejected the tenor of Kangaroo, although Lawrence would not have been worried about the response. In 1929 he referred to his irritation with Australia in letters to P.R. “Inky” Stephensen, the Australian nationalist and publisher, and he does not seem to have changed his opinions since writing Kangaroo. Yet the novel continued to be significant for Australian writers, even if as a provocation. My discussion traces the responses of the women authors to Kangaroo, and refers to Lawrence’s letters to Stephensen, as a way of emphasizing this significance, seen especially in relation to ideas about ‘seeing’ and the Australian landscape.

Total Dik - Transmedia Concept development

Total Dik! is a collaborative project between the Queensland University of Technology (QUT) and Queensland Theatre Company (QTC). Total Dik! explores transmedia storytelling in live performance from concept development to delivery and builds on works, By the Way, Meet Vera Stark, (Forrester2012), Hotel Modern’s Kamp (2005) and God’s Beard (2012) that use visual art, puppetry, music and film. The project’s first iteration enabled an interrogation of the integration of media-rich elements with live performers in a theatrical environment. Performative transmedia storytelling draws on the tenets of convergent media theory developed by Jenkins (2007, 2012), Dena (2010) and Philips (2012). This exploratory work, juxtaposing transmedia storytelling techniques with live performance, draws on Samuel Becket’s challenges to theatre orthodoxy, and touches on Brechtian notions of alienation through ‘sleight-of-hand’ or processual unpacking and deconstruction during performance. Total Dik! blends a convergence of technologies, models, green screen capture, and live dimensions of performance in one narrative allowing the work’s creators to test new combinations of transmedia storytelling techniques on a traditional performance platform.

Barbadocladius Cranston & Krosch, a new genus of Orthocladiinae (Diptera : Chironomidae) from South America.

Barbadocladius n. gen. is erected and described in larval, pupal and adult stages for two species: B. andinus sp. nov. and B. limay sp. nov., from Andean streams. The larva is distinctive by virtue of the very large ventromental 'beard' and the anterior parapods with a 'sleeve' of hooklets in addition to apical pectinate claws. The pupa has hooklets on some tergal and sternal intersegmental membranes. The adult, reported only in teneral specimens has hairy eyes, no antennal apical strong seta, no acrostichals, bare and unmarked wings, cylindrical 4th tarsomere subequal in length to the 5th, pulvilli about half the claw length, and hypopygium with anal point, lacking a virga. Molecular phylogenetic analysis eliminates relationships directly to the Eukiefferiella complex (which also have pupal hooklets), or to the Cricotopus group (adults also with hairy eyes), suggesting instead a sister group relationship to a suite of predominantly austral genera of Orthocladiinae.

Green: Exploring the aestheticized use of chroma-key techniques and technologies in two intermedial productions

Peggy Shaw’s RUFF, (USA 2013) and Queensland Theatre Company’s collaboration with Queensland University of Technology, Total Dik!, (Australia 2013) overtly and evocatively draw on an aestheticized use of the cinematic techniques and technologies of Chroma Key to reveal the tensions in their production and add layers to their performances. In doing so they offer invaluable insight where the filmic and theatrical approaches overlap. This paper draws on Eckersall, Grehan and Scheer’s New Media Dramaturgy (2014) to reposition the frame as a contribution to intermedial theatre and performance practices in light of increasing convergence between seemingly disparate discourses. In RUFF, the scenic environment replicates a chroma-key ‘studio’ which facilitates the reconstruction of memory displaced after a stroke. RUFF uses the screen and projections to recall crooners, lounge singers, movie stars, rock and roll bands, and an eclectic line of eccentric family members living inside Shaw. While the show pays tribute to those who have kept her company across decades of theatrical performance, use of non-composited chroma-key technique as a theatrical device and the work’s taciturn revelation of the production process during performance, play a central role in its exploration of the juxtaposition between its reconstructed form and content. In contrast Total Dik! uses real-time green screen compositing during performance as a scenic device. Actors manipulate scale models, refocus cameras and generate scenes within scenes in the construction of the work’s examination of an isolated Dictator. The ‘studio’ is again replicated as a site for (re)construction, only in this case Total Dik! actively seeks to reveal the process of production as the performance plays out. Building on RUFF, and other works such as By the Way, Meet Vera Stark, (2012) and Hotel Modern’s God’s Beard (2012), this work blends a convergence of mobile technologies, models, and green screen capture to explore aspects of transmedia storytelling in a theatrical environment (Jenkins, 2009, 2013). When a green screen is placed on stage, it reads at once as metaphor and challenge to the language of theatre. It becomes, or rather acts, as a ‘sign’ that alludes to the nature of the reconstructed, recomposited, manipulated and controlled. In RUFF and in Total Dik!, it is also a place where as a mode of production and subsequent reveal, it adds weight to performance. These works are informed by Auslander (1999) and Giesenkam (2007) and speak to and echo Lehmann’s Postdramatic Theatre (2006). This paper’s consideration of the integration of studio technique and live performance as a dynamic approach to multi-layered theatrical production develops our understanding of their combinatory use in a live performance environment.

hSSB1 (NABP2/ OBFC2B) is required for the repair of 8-oxo-guanine by the hOGG1-mediated base excision repair pathway

The maintenance of genome stability is essential to prevent loss of genetic information and the development of diseases such as cancer. One of the most common forms of damage to the genetic code is the oxidation of DNA by reactive oxygen species (ROS), of which 8-oxo-7,8-dihydro-guanine (8-oxoG) is the most frequent modification. Previous studies have established that human single-stranded DNA-binding protein 1 (hSSB1) is essential for the repair of double-stranded DNA breaks by the process of homologous recombination. Here we show that hSSB1 is also required following oxidative damage. Cells lacking hSSB1 are sensitive to oxidizing agents, have deficient ATM and p53 activation and cannot effectively repair 8-oxoGs. Furthermore, we demonstrate that hSSB1 forms a complex with the human oxo-guanine glycosylase 1 (hOGG1) and is important for hOGG1 localization to the damaged chromatin. In vitro, hSSB1 binds directly to DNA containing 8-oxoguanines and enhances hOGG1 activity. These results underpin the crucial role hSSB1 plays as a guardian of the genome.