BBF RFC 37: Fusion protein BioBrick assembly standard with optional linker extension

BBF RFC 37 replaces BBF RFC 36.This RFC 37 describes an extension of the original BioBrick assembly standard (BBF RFC 10) and Freiburg assembly standard (BBF RFC 25). The Fusion Assembly strategy described here is fully compatible with RFC 25 (Freiburg) and RFC 10 standard and supports in-frame fusion and controlled potentially infinite extension of linker between protein fusion domains

Mycoplasma synoviae induces upregulation of apoptotic genes, secretion of nitric oxide and appearance of an apoptotic phenotype in infected chicken chondrocytes

The role of chondrocytes in the development of infectious arthritis is not well understood. Several examples of mycoplasma-induced arthritis in animals indicate that chondrocytes come into direct contact with bacteria. The objective of this study was to analyze the interaction of an arthrogenic Mycoplasma synoviae strain WVU 1853 with chicken chondrocytes. We found that M. synoviae significantly reduces chondrocyte respiration. This was accompanied by alterations in chondrocyte morphology, namely cell shrinkage and cytoplasm condensation, as well as nuclear condensation and formation of plasma membrane invaginations containing nuclear material, which appeared to cleave off the cell surface. In concordance with these apoptosis-like events in chondrocytes, transcription was increased in several pro-apoptotic genes. Twenty-four hours after infection, strong upregulation was assayed in NOS2, Mapk11, CASP8 and Casp3 genes. Twenty-four and 72 h incubation of chondrocytes with M. synoviae induced upregulation of AIFM1, NFκB1, htrA3 and BCL2. Casp3 and NOS2 remained upregulated, but upregulation ceased for Mapk11 and CASP8 genes. Increased production of nitric oxide was also confirmed in cell supernates. The data suggests that chicken chondrocytes infected with M. synoviae die by apoptosis involving production of nitric oxide, caspase 3 activation and mitochondrial inactivation. The results of this study show for the first time that mycoplasmas could cause chondrocyte apoptosis. This could contribute to tissue destruction and influence the development of arthritic conditions. Hence, the study gives new insights into the role of mycoplasma infection on chondrocyte biology and development of infectious arthritis in chickens and potentially in humans

Health and economic burden of skin melanoma in Croatia – cost-of-illness study

Melanoma incidence is increasing, especially in the younger population. The aim of this study was to investigate the cost of this disease in the Croatian population and to identify costs through types of care and types of costs. The secondary goal was to estimate the prevalence of certain types of melanoma (as well as staging distribution) and to connect each stage and its prevalence in Croatia to related costs. A cost-of-illness analysis was performed, mainly including direct costs (monitoring, drugs, primary health care services, hospitalizations, and diagnostics). The calculations were based on data collected from Sestre milosrdnice University Hospital Center in Zagreb and from Cancer Registry Data. The number of patients with melanoma was calculated using the Markov model for melanoma staging and 5-year survival. The estimated total prevalence of melanoma in 2011 in Croatia was 2,180. The total cost of melanoma was estimated to 1,063,488 EUR, with 46% used for hospitalization and chemotherapy, 10% for dermatoscopy, and the remaining 17% being monitoring costs. The average cost per patient was estimated to range between 98 and 4,333 EUR depending on the stage of the disease. The cost of melanoma in the adult population in Croatia in a one-year timeframe accounted for as much as 0.04% of the total Croatian national health care budget for 2011. Study findings indicate the need for a clear strategy to achieve regular screening in order to detect the disease at an early stage.  </p

The Aspergillus giganteus antifungal protein AFPNN5353 activates the cell wall integrity pathway and perturbs calcium homeostasis

Background The antifungal protein AFPNN5353 is a defensin-like protein of Aspergillus giganteus. It belongs to a group of secretory proteins with low molecular mass, cationic character and a high content of cysteine residues. The protein inhibits the germination and growth of filamentous ascomycetes, including important human and plant pathogens and the model organsims Aspergillus nidulans and Aspergillus niger. Results We determined an AFPNN5353 hypersensitive phenotype of non-functional A. nidulans mutants in the protein kinase C (Pkc)/mitogen-activated protein kinase (Mpk) signalling pathway and the induction of the α-glucan synthase A (agsA) promoter in a transgenic A. niger strain which point at the activation of the cell wall integrity pathway (CWIP) and the remodelling of the cell wall in response to AFPNN5353. The activation of the CWIP by AFPNN5353, however, operates independently from RhoA which is the central regulator of CWIP signal transduction in fungi. Furthermore, we provide evidence that calcium (Ca2+) signalling plays an important role in the mechanistic function of this antifungal protein. AFPNN5353 increased about 2-fold the cytosolic free Ca2+ ([Ca2+]c) of a transgenic A. niger strain expressing codon optimized aequorin. Supplementation of the growth medium with CaCl2 counteracted AFPNN5353 toxicity, ameliorated the perturbation of the [Ca2+]c resting level and prevented protein uptake into Aspergillus sp. cells. Conclusions The present study contributes new insights into the molecular mechanisms of action of the A. giganteus antifungal protein AFPNN5353. We identified its antifungal activity, initiated the investigation of pathways that determine protein toxicity, namely the CWIP and the Ca2+ signalling cascade, and studied in detail the cellular uptake mechanism in sensitive target fungi. This knowledge contributes to define new potential targets for the development of novel antifungal strategies to prevent and combat infections of filamentous fungi which have severe negative impact in medicine and agriculture.FWF, P19970-B11, Characterization of the toxicity of PA

pDNA capture using grafted adsorbents

BACKGROUND: ‘Expanded’ composite materials are of interest as an alternative, or as a supplement, to packed-bed chromatography during bioproduct recovery and purification. Functionalized non-woven fabrics and mega-porous bodies are examples of systems that showed promise. However, there is scarce information on their suitability to capture and release plasmid DNA (pDNA), an important type of product employed in gene therapy. RESULTS: Composite adsorbents were prepared using either chemical (CG-DEAE-NW) or gamma-irradiated graft-polymerization (GIR-DEAE-MP), and subsequently modified to have diethylamino ethanol (DEAE) functionality. Capture experiments showed that pDNA can actually reversibly bind to the two mentioned adsorbents, with capacity values of 2.4 and 1.3 mg per mL, respectively. These values are in the range of what can be expected from commercial beaded adsorbents but lower that the values expected from monoliths. CONCLUSIONS: Expanded materials, due to their high voidage, may present limited capacity for pDNA. However, such materials are able to bind proteins and other contaminants from bacterial lysate, opening the way for their utilization in the ‘negative’ mode.Fil: Singh, Naveen Kumar. University of Notre Dame; Estados Unidos. Jacobs University; AlemaniaFil: Dsouza, Roy N.. Jacobs University; AlemaniaFil: Yelemane, Vikas. Jacobs University; AlemaniaFil: Nentwig, Nina. Jacobs University; AlemaniaFil: Grasselli, Mariano. Universidad Nacional de Quilmes; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata; ArgentinaFil: Fernández Lahore, Marcelo. Jacobs University; Alemani

Health and economic burden of skin melanoma in Croatia – cost-of-illness study

Melanoma incidence is increasing, especially in the younger population. The aim of this study was to investigate the cost of this disease in the Croatian population and to identify costs through types of care and types of costs. The secondary goal was to estimate the prevalence of certain types of melanoma (as well as staging distribution) and to connect each stage and its prevalence in Croatia to related costs. A cost-of-illness analysis was performed, mainly including direct costs (monitoring, drugs, primary health care services, hospitalizations, and diagnostics). The calculations were based on data collected from Sestre milosrdnice University Hospital Center in Zagreb and from Cancer Registry Data. The number of patients with melanoma was calculated using the Markov model for melanoma staging and 5-year survival. The estimated total prevalence of melanoma in 2011 in Croatia was 2,180. The total cost of melanoma was estimated to 1,063,488 EUR, with 46% used for hospitalization and chemotherapy, 10% for dermatoscopy, and the remaining 17% being monitoring costs. The average cost per patient was estimated to range between 98 and 4,333 EUR depending on the stage of the disease. The cost of melanoma in the adult population in Croatia in a one-year timeframe accounted for as much as 0.04% of the total Croatian national health care budget for 2011. Study findings indicate the need for a clear strategy to achieve regular screening in order to detect the disease at an early stage.  </p

A Zoomable Mapping of a Musical Parameter Space Using Hilbert Curves

The final publication is available at Computer Music Journal via http://dx.doi.org/10.1162/COMJ_a_0025

HTA and Reimbursement Status of Metastatic Hormone‑Sensitive Prostate Cancer, Non-Metastatic Castration-Resistant Prostate Cancer, and Metastatic Castration-Resistant Prostate Cancer Treatments in Europe: A Patient Access Landscape Review

**Background:** Prostate cancer is the second most common cancer in men, with up to one-third of men being diagnosed in their lifetime. Recently, novel therapies have received regulatory approval with significant improvement in overall survival for metastatic castration-resistant prostate cancer, metastatic hormone-sensitive prostate cancer, and nonmetastatic castration-resistant prostate cancer. To improve decision-making regarding the value of anticancer therapies and support standardized assessment for use by health technology assessment (HTA) agencies, the European Society for Medical Oncology (ESMO) has developed a Magnitude of Clinical Benefit Scale (MCBS). **Objective:** This review aimed to map HTA status, reimbursement restrictions, and patient access for 3 advanced prostate cancer indications across 23 European countries during 2011-2021. **Methods:** HTA, country reimbursement lists, and ESMO-MCBS scorecards were reviewed for evidence and data across 26 European countries. **Results:** The analysis demonstrated that only in Greece, Germany, and Sweden was there full access across all included prostate cancer treatments. Treatments available for metastatic castration-resistant prostate cancer were widely reimbursed, with both abiraterone and enzalutamide accessible in all countries. In 3 countries (Hungary, the Netherlands, and Switzerland), there was a statistically significant difference (_P_<.05) between status of reimbursement and ESMO-MCBS “substantial benefit” (score of 4 or 5) vs “no substantial benefit” (score <4). **Conclusion:** Overall, the impact of the ESMO-MCBS on reimbursement decisions in Europe is unclear, with significant variation across the countries included in this review

The antifungal protein PAF interferes with PKC/MPK and cAMP/PKA signalling of Aspergillus nidulans

The Penicillium chrysogenum antifungal protein PAF inhibits polar growth and induces apoptosis in Aspergillus nidulans. We report here that two signalling cascades are implicated in its antifungal activity. PAF activates the cAMP/protein kinase A (Pka) signalling cascade. A pkaA deletion mutant exhibited reduced sensitivity towards PAF. This was substantiated by the use of pharmacological modulators: PAF aggravated the effect of the activator 8-Br-cAMP and partially relieved the repressive activity of caffeine. Furthermore, the Pkc/mitogen-activated protein kinase (Mpk) signalling cascade mediated basal resistance to PAF, which was independent of the small GTPase RhoA. Non-functional mutations of both genes resulted in hypersensitivity towards PAF. PAF did not increase MpkA phosphorylation or induce enzymes involved in the remodelling of the cell wall, which normally occurs in response to activators of the cell wall integrity pathway. Notably, PAF exposure resulted in actin gene repression and a deregulation of the chitin deposition at hyphal tips of A. nidulans, which offers an explanation for the morphological effects evoked by PAF and which could be attributed to the interconnection of the two signalling pathways. Thus, PAF represents an excellent tool to study signalling pathways in this model organism and to define potential fungal targets to develop new antifungals