A mutational hot spot in keratin 10 (KRT 10) in patients with epidermolytic hyperkeratosis

Epidermolytic hyperkeratosis (EHK), (bullous congenital ichthyosiform erythroderma), is an autosomal dominant human skin disorder. Recently, we and others have described mutations in keratins 1 and 10 (K1 and K10) in patients with this disease. Structure-function models predict that these mutations would impair normal filament assembly and function. We have extended our earlier studies to include 8 more incidences of EHK. In half of these families, we were unable to locate a mutation within the rod domains of either K1 or K10. However, polymorphic restriction site and sequence analysis of the other families revealed a mutational hot spot within the 1A alpha-helical segment of K10. These involve Arginine to Histidine, Arginine to Cysteine and Arginine to Leucine substitutions at residue 10 of the rod domain. Interestingly, mutations in the corresponding Arginine residue in keratin K14 have been identified in patients with epidermolysis bullosa simplex. The large number of mutations found at this position in both keratins K10 and K14 suggests that other epithelia cell disorders will be discovered that are caused by the corresponding mutation in related type I keratin gene

Reduced P53 levels ameliorate neuromuscular junction loss without affecting motor neuron pathology in a mouse model of spinal muscular atrophy

Spinal Muscular Atrophy (SMA) is a childhood motor neuron disease caused by mutations or deletions within the SMN1 gene. At endstages of disease there is profound loss of motor neurons, loss of axons within ventral roots and defects at the neuromuscular junctions (NMJ), as evidenced by pathological features such as pre-synaptic loss and swelling and post-synaptic shrinkage. Although these motor unit defects have been widely described, the time course and interdependancy of these aspects of motor unit degeneration are unclear. Recent reports have also revealed an early upregulation of transcripts associated with the P53 signalling pathway. The relationship between the upregulation of these transcripts and pathology within the motor unit is also unclear. In this study, we exploit the prolonged disease timecourse and defined pre-symptomatic period in the Smn mouse model to perform a temporal analysis of the different elements of motor unit pathology. We demonstrate that NMJ loss occurs prior to cell body loss, and coincides with the onset of symptoms. The onset of NMJ pathology also coincides with an increase in P53-related transcripts at the cell body. Finally, using a tamoxifen inducible P53 knockout, we demonstrate that post-natal reduction in P53 levels can reduce NMJ loss, but does not affect other aspects of NMJ pathology, motor neuron loss or the phenotype of the Smn mouse model. Together this work provides a detailed temporal description of pathology within motor units of an SMA mouse model, and demonstrates that NMJ loss is a P53-dependant process. This work supports the role for P53 as an effector of synaptic and axonal degeneration in a die-back neuropathy

Abstracts from the 8th International Conference on cGMP Generators, Effectors and Therapeutic Implications

This work was supported by a restricted research grant of Bayer AG

The evolving SARS-CoV-2 epidemic in Africa: Insights from rapidly expanding genomic surveillance

INTRODUCTION Investment in Africa over the past year with regard to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) sequencing has led to a massive increase in the number of sequences, which, to date, exceeds 100,000 sequences generated to track the pandemic on the continent. These sequences have profoundly affected how public health officials in Africa have navigated the COVID-19 pandemic. RATIONALE We demonstrate how the first 100,000 SARS-CoV-2 sequences from Africa have helped monitor the epidemic on the continent, how genomic surveillance expanded over the course of the pandemic, and how we adapted our sequencing methods to deal with an evolving virus. Finally, we also examine how viral lineages have spread across the continent in a phylogeographic framework to gain insights into the underlying temporal and spatial transmission dynamics for several variants of concern (VOCs). RESULTS Our results indicate that the number of countries in Africa that can sequence the virus within their own borders is growing and that this is coupled with a shorter turnaround time from the time of sampling to sequence submission. Ongoing evolution necessitated the continual updating of primer sets, and, as a result, eight primer sets were designed in tandem with viral evolution and used to ensure effective sequencing of the virus. The pandemic unfolded through multiple waves of infection that were each driven by distinct genetic lineages, with B.1-like ancestral strains associated with the first pandemic wave of infections in 2020. Successive waves on the continent were fueled by different VOCs, with Alpha and Beta cocirculating in distinct spatial patterns during the second wave and Delta and Omicron affecting the whole continent during the third and fourth waves, respectively. Phylogeographic reconstruction points toward distinct differences in viral importation and exportation patterns associated with the Alpha, Beta, Delta, and Omicron variants and subvariants, when considering both Africa versus the rest of the world and viral dissemination within the continent. Our epidemiological and phylogenetic inferences therefore underscore the heterogeneous nature of the pandemic on the continent and highlight key insights and challenges, for instance, recognizing the limitations of low testing proportions. We also highlight the early warning capacity that genomic surveillance in Africa has had for the rest of the world with the detection of new lineages and variants, the most recent being the characterization of various Omicron subvariants. CONCLUSION Sustained investment for diagnostics and genomic surveillance in Africa is needed as the virus continues to evolve. This is important not only to help combat SARS-CoV-2 on the continent but also because it can be used as a platform to help address the many emerging and reemerging infectious disease threats in Africa. In particular, capacity building for local sequencing within countries or within the continent should be prioritized because this is generally associated with shorter turnaround times, providing the most benefit to local public health authorities tasked with pandemic response and mitigation and allowing for the fastest reaction to localized outbreaks. These investments are crucial for pandemic preparedness and response and will serve the health of the continent well into the 21st century

Tissue‐ and diet‐dependent stable carbon and nitrogen isotope discrimination: a calibration study in a captive shorebird species

In ecology, stable‐isotope ratios are widely used to determine diets of organisms and reconstruct food webs. This is usually done by analyzing the stable‐isotope ratios of nitrogen (δ15N), which increase with increasing trophic level, and those of carbon (δ13C), which correlate with the δ13C value of food source(s) and generally differ between terrestrial and marine food sources. Assimilation of food changes stable‐isotope ratios, resulting in different values between the food source and its consumer. These differences are known as isotope trophic discrimination factors and, if known, can be used to determine from the stable‐isotope ratios in the consumer's tissue what the consumer has been eating. What is often ignored is that discrimination factors can differ between consumer's food sources and also between tissue types. Therefore, we performed a controlled feeding study in red knots Calidris canutus to determine discrimination factors between different food sources and red knot tissues. We kept two groups of red knots in captivity on a stable diet, one group feeding on mudsnails and the other on Trouvit pellets, for several months, during which the birds molted their feathers. We analyzed δ13C and δ15N in both food sources and in five red knot tissues (blood cells, blood plasma and three feather types) and subsequently calculated the isotope discrimination factors. We confirmed that the discrimination factors differed between tissues, and also between diets. Our values deviated from general averages reported in reviews on a wide range of animals/birds, but were very similar to values from previous red knot and dunlin studies. We therefore think that our discrimination factors can be used in future stable isotope studies, not only on red knots, but also on other marine shorebird species and plea for careful consideration of using the right discrimination factors. Keywords: δ13C, δ15N, discrimination factor, red knot, shorebird, stable isotop

Tissue- and diet-dependent stable carbon and nitrogen isotope discrimination: a calibration study in a captive shorebird species

In ecology, stable-isotope ratios are widely used to determine diets of organisms and reconstruct food webs. This is usually done by analyzing the stable-isotope ratios of nitrogen (?15N), which increase with increasing trophic level, and those of carbon (?13C), which correlate with the ?13C value of food source(s) and generally differ between terrestrial and marine food sources. Assimilation of food changes stable-isotope ratios, resulting in different values between the food source and its consumer. These differences are known as isotope trophic discrimination factors and, if known, can be used to determine from the stable-isotope ratios in the consumer?s tissue what the consumer has been eating. What is often ignored is that discrimination factors can differ between consumer?s food sources and also between tissue types. Therefore, we performed a controlled feeding study in red knots Calidris canutus to determine discrimination factors between different food sources and red knot tissues. We kept two groups of red knots in captivity on a stable diet, one group feeding on mudsnails and the other on Trouvit pellets, for several months, during which the birds molted their feathers. We analyzed ?13C and ?15N in both food sources and in five red knot tissues (blood cells, blood plasma and three feather types) and subsequently calculated the isotope discrimination factors. We confirmed that the discrimination factors differed between tissues, and also between diets. Our values deviated from general averages reported in reviews on a wide range of animals/birds, but were very similar to values from previous red knot and dunlin studies. We therefore think that our discrimination factors can be used in future stable isotope studies, not only on red knots, but also on other marine shorebird species and plea for careful consideration of using the right discrimination factors

Detection of codon 12 mutation in the k-ras oncogene in pancreatic tumors Detecção de mutação no códon 12 do oncogene K-ras em tumores pancreáticos

Mutations at codons 12, 13, or 61 of the H-ras, K-ras, and N-ras have been detected in human neoplasias by a variety of techniques. Some of these techniques are very sensitive and can detect K-ras mutation in 90% of the cases of pancreatic adenocarcinomas. We analyzed 11 samples of pancreatic adenocarcinoma, three samples of pancreatic mucinous cystadenoma, and two samples without tumors in formalin-fixed paraffin embedded tissue sections. K-ras mutations at codon 12 were detected by a two-step PCR-enriched technique in all the samples of pancreatic adenocarcinoma, but not in cystadenoma or control samples. This technique may be useful for early detection of pancreatic cancer.<br>Muitos dos oncogenes detectados em neoplasias malignas humanas pertencem à família do gene ras. Mutações nos códons 12, 13 ou 61 em um dos tres genes ras; H-ras, K-ras e N-ras, convertem esses genes em oncogenes ativos. Ensaios rápidos para detecção dessas mutações pontuais, tais como a reação em cadeia de polimertização têm sido desenvolvidos nas últimas décadas e usados para investigar o papel dos genes ras mutados na patogênese de tumores humanos. As mutações no gene ras podem ser encontradas numa variedade de tipos de tumores. Incidências mais altas aparecem em adenocarcinomas do pâncreas (90%) e cólon (50%). Analisamos 11 amostras de tumores primários de pâncreas com diferentes metástases, três amostras de cistadenoma mucinoso e dois casos de ausência de tumor de material incluído em parafina, de onde extraímos o DNA para realização das amplificações. Os resultados mostraram que todos os casos de tumores apresentaram a banda de 135 pares de bases correspondente ao gene mutado e para os normais, a banda característica de 106 pares de bases. Nos três casos de cistadenoma mucinosos, não detectamos a banda de 135 pares de bases , apenas a banda de 106 pares de bases