Benchmarking trial between France and Australia comparing management of primary rectal cancer beyond TME and locally recurrent rectal cancer (PelviCare Trial): rationale and design.

BACKGROUND: Among patients with rectal cancer, 5-10% have a primary rectal cancer beyond the total mesorectal excision plane (PRC-bTME) and 10% recur locally following primary surgery (LRRC). In both cases, patients 'care remains challenging with a significant worldwide variation in practice regarding overall management and criteria for operative intervention. These variations in practice can be explained by structural and organizational differences, as well as cultural dissimilarities. However, surgical resection of PRC-bTME and LRRC provides the best chance of long-term survival after complete resection (R0). With regards to the organization of the healthcare system and the operative criteria for these patients, France and Australia seem to be highly different. A benchmarking-type analysis between French and Australian clinical practice, with regards to the care and management of PRC-bTME and LRRC, would allow understanding of patients' care and management structures as well as individual and collective mechanisms of operative decision-making in order to ensure equitable practice and improve survival for these patients. METHODS/DESIGN: The current study is an international Benchmarking trial comparing two cohorts of 120 consecutive patients with non-metastatic PRC-bTME and LRRC. Patients with curative and palliative treatment intent are included. The study design has three main parts: (1) French and Australian cohorts including clinical, radiological and surgical data, quality of life (MOS SF36, FACT-C) and distress level (Distress thermometer) at the inclusion, 6 and 12 months; (2) experimental analyses consisting of a blinded inter-country reading of pelvic MRI to assess operatory decisions; (3) qualitative analyses based on MDT meeting observation, semi-structured interviews and focus groups of health professional attendees and conducted by a research psychologist in both countries using the same guides. The primary endpoint will be the clinical resection rate. Secondary end points will be concordance rate between French and Australian operative decisions based on the inter-country reading MRI, post-operative mortality and morbidity rates, oncological outcomes based on resection status and one-year overall and disease-free survival, patients' quality of life and distress level. Qualitative analysis will compare obstacles and facilitators of operative decision-making between both countries. DISCUSSION: Benchmarking can be defined as a comparison and learning process which will allow, in the context of PRC-bTME and LRRC, to understand and to share the whole process management of these patientsbetween Farnce and Australia. TRIAL REGISTRATION: NCT02551471 . (date of registration: 09/14/2015).

A TSPO-related protein localizes to the early secretory pathway in Arabidopsis, but is targeted to mitochondria when expressed in yeast.

AtTSPO is a TspO/MBR domain-protein potentially involved in multiple stress regulation in Arabidopsis. As in most angiosperms, AtTSPO is encoded by a single, intronless gene. Expression of AtTSPO is tightly regulated both at the transcriptional and post-translational levels. It has been shown previously that overexpression of AtTSPO in plant cell can be detrimental, and the protein was detected in the endoplasmic reticulum (ER) and Golgi stacks, contrasting with previous findings and suggesting a mitochondrial subcellular localization for this protein. To ascertain these findings, immunocytochemistry and ABA induction were used to demonstrate that, in plant cells, physiological levels of AtTSPO colocalized with AtArf1, a mainly Golgi-localized protein in plant cells. In addition, fluorescent protein-tagged AtTSPO was targeted to the secretory pathway and did not colocalize with MitoTracker-labelled mitochondria. These results suggest that the polytopic membrane protein AtTSPO is cotranslationally targeted to the ER in plant cells and accumulates in the Trans-Golgi Network. Heterologous expression of AtTSPO in Saccharomyces cerevisiae, yeast devoid of TSPO-related protein, resulted in growth defects. However, subcellular fractionation and immunoprecipitation experiments showed that AtTSPO was targeted to mitochondria where it colocalized and interacted with the outer mitochondrial membrane porin VDAC1p, reminiscent of the subcellular localization and activity of mammalian translocator protein 18 kDa TSPO. The evolutionarily divergent AtTSPO appears therefore to be switching its sorting mode in a species-dependent manner, an uncommon peculiarity for a polytopic membrane protein in eukaryotic cells. These results are discussed in relation to the recognition and organelle targeting mechanisms of polytopic membrane proteins in eukaryotic cells.

ABA, porphyrins and plant TSPO-related protein.

We have shown that, unexpectedly, AtTSPO (Arabidopsis thaliana TSPO-related protein) is an endoplasmic reticulum and Golgi-localized membrane protein in plant cells.(1) This localization contrasts with that of mammalian 18-kDa translocator protein (at least for the mostly studied isoform, 18-kDa TSPO), a mitochondrial outer membrane protein (reviewed in ref. 2). Whereas the potential functions of 18-kDa TSPO are well documented, involved mainly in mitochondrial physiology,(2) and its interest as drugs target is been explored,(3) the roles of TSPO-related proteins in plant growth and development are yet to be specified. AtTSPO is expressed in dry seeds and can be induced in vegetative tissues by osmotic and salt stress or abscisic acid (ABA) treatment. Moreover, it was shown that the ABA-dependent induction is transient, and that boosting tetrapyrroles biosynthesis through 5-aminolevulinic acid (ALA) feeding enhanced downregulation of AtTSPO, suggesting an inherent post-translational regulation mechanism also involving ABA and likely porphyrins. We present additional evidence that ABA can help stabilize constitutively expressed AtTSPO and that ALA feeding to knockout mutant seeds, induces substantial germination delay. Here we discuss the possible link between ABA and tetrapyrroles in AtTSPO expression and post-translational regulation.

The Arabidopsis TSPO-related protein is a stress and abscisic acid-regulated, endoplasmic reticulum-Golgi-localized membrane protein.

The Arabidopsis gene At2g47770 encodes a membrane-bound protein designated AtTSPO (Arabidopsis thaliana TSPO-related). AtTSPO is related to the bacterial outer membrane tryptophan-rich sensory protein (TspO) and the mammalian mitochondrial 18-kDa translocator protein (18 kDa TSPO), members of the group of TspO/MBR domain-containing membrane proteins. In this study we show that AtTSPO is mainly detected in dry seeds, but can be induced in vegetative tissues by osmotic or salt stress or abscisic acid (ABA) treatment, corroborating available transcriptome data. Using subcellular fractionation, immunocytochemistry and fluorescent protein tagging approaches we present evidence that AtTSPO is targeted to the secretory pathway in plants. Induced or constitutively expressed AtTSPO can be detected in the endoplasmic reticulum and the Golgi stacks of plant cells. AtTSPO tagged with fluorescent protein in transgenic plants (Arabidopsis and tobacco) was mainly detected in the Golgi stacks of leaf epidermal cells. Constitutive expression of AtTSPO resulted in increased sensitivity to NaCl, but not to osmotic stress, and in reduced greening of cultured Arabidopsis cells under light growing conditions. Transgenic Arabidopsis plants overexpressing AtTSPO were more sensitive to ABA-induced growth inhibition, indicating that constitutive expression of AtTSPO may enhance ABA sensitivity. AtTSPO is rapidly downregulated during seed imbibition, and the ABA-dependent induction in plant is transient. Downregulation of AtTSPO seems to be boosted by treatment with aminolevulinic acid. Taken together, these results suggest that AtTSPO is a highly regulated protein, induced by abiotic stress to modulate, at least in part, transient intracellular ABA-dependent stress perception and/or signalling.

Harnessing the potential of hairy roots: dawn of a new era.

In the past two decades, hairy root research for the production of important secondary metabolites has received a lot of attention. The addition of knowledge to overcome the limiting culture parameters of the regulation of the metabolic pathway by specific molecules and the development of novel tools for metabolic engineering now offer new possibilities to improve the hairy root technique for the production of metabolites. Furthermore, engineering hairy roots for the production of animal proteins of therapeutic interest in confined and controlled in vitro conditions is seen as one of the exciting spin-offs of the technology. Recent progress made in the scale-up of the hairy root cultures has paved the way for industrial exploitation of this system. This review highlights some of the significant progress made in the past three years and discusses the potential implications of that research.

Hairy root research: recent scenario and exciting prospects.

High stability of the production of secondary metabolites is an interesting characteristic of hairy root cultures. For 25 years, hairy roots have been investigated as a biological system for the production of valuable compounds from medicinal plants. A better understanding of the molecular mechanism of hairy root development, which is based on the transfer of Agrobacterium rhizogenes T-DNA into the plant genome, has facilitated its increasing use in metabolic engineering. Hairy roots can also produce recombinant proteins from transgenic roots, and thereby hold immense potential for the pharmaceutical industry. In addition, hairy roots offer promise for phytoremediation because of their abundant neoplastic root proliferation. Recent progress in the scaling-up of hairy root cultures is making this system an attractive tool for industrial processes.