A Saturation-Mutagenesis Analysis of the Interplay Between Stability and Activation in Ras

Cancer mutations in Ras occur predominantly at three hotspots: Gly 12, Gly 13, and Gln 61. Previously, we reported that deep mutagenesis of H?Ras using a bacterial assay identified many other activating mutations (Bandaru et al., 2017). We now show that the results of saturation mutagenesis of H?Ras in mammalian Ba/F3 cells correlate well with results of bacterial experiments in which H-Ras or K-Ras are co-expressed with a GTPase?activating protein (GAP). The prominent cancer hotspots are not dominant in the Ba/F3 data. We used the bacterial system to mutagenize Ras constructs of different stabilities and discovered a feature that distinguishes the cancer hotspots. While mutations at the cancer hotspots activate Ras regardless of construct stability, mutations at lower-frequency sites (e.g., at Val 14 or Asp 119) can be activating or deleterious, depending on the stability of the Ras construct. We characterized the dynamics of three non-hotspot activating Ras mutants by using NMR to monitor hydrogen?deuterium exchange (HDX). These mutations result in global increases in HDX rates, consistent with the destabilization of Ras. An explanation for these observations is that mutations that destabilize Ras increase nucleotide dissociation rates, enabling activation by spontaneous nucleotide exchange. A further stability decrease can lead to insufficient levels of folded Ras – and subsequent loss of function. In contrast, the cancer hotspot mutations are mechanism-based activators of Ras that interfere directly with the action of GAPs. Our results demonstrate the importance of GAP surveillance and protein stability in determining the sensitivity of Ras to mutational activation.

Characteristics of Older Adults Accessing Medical Assistance in Dying (MAiD): a Descriptive Study

Background Medical Assistance in Dying (MAiD) is an end-of-life option for Canadians accounting for 2% of all deaths in Canada in 2019. Adults over 80 years old represent a significant proportion of these deaths, yet little is known about how they compare with their younger counterparts. Methods This study retrospectively reviewed our tertiary care institution’s MAiD database to compare MAiD recipients <65, 65–80, and >80 years of age. Extracted data included basic demographics, illness characteristics, functional status, social living arrangements/contacts, and outcomes of MAiD assessments. Results Of 267 patients assessed for MAiD, 38.2% were over 80. Compared to the younger groups, those over 80 were more likely to be female, to live alone, and to be widowed; however, they did not self-identify as ‘socially isolated’. The majority fit into the illness categories of malignancy, cardiopulmonary or neurologic diseases, but those over 80 were more likely to have other more chronic/subacute conditions leading to the MAiD request. Conclusions Older adults accessing MAiD are distinct in that they tend to be increasingly frail and without a predominant underlying diagnosis as compared with younger adults, but rather have an accumulation of losses resulting in global functional decline and subsequent loss of autonomy and independence.

Dress and Dress Fasteners from the Pagan Norse Graves of Scotland: Origins and Identity

The pagan Norse graves of Scotland provide a tantalising glimpse of what the interred were wearing at the time of burial. However, the survival of actual clothing, frequently in a mineralised state, is rare. Yet dress accessories—typically brooches, cloak pins and belt fittings—help to create a more comprehensive picture of the dress worn, and of contemporary fashion. The origins of these fittings can be identified by their form, ornament and metallic composition. Some, such as the paired oval brooches found in female burials, are typically Scandinavian and indicative of the wearing of a pinafore-like dress, corroborated on occasion by surviving textile loops mineralised within their shells. However, others, including penannular brooches, ringed pins and bossed belt fittings, are of Insular origin and illustrate an emerging Hiberno-Scandinavian identity. Such evidence for cultural interaction through dress provides a fascinating insight into contemporary perceptions of identity, just prior to the abandonment of the pagan ritual with the subsequent loss of this rich source of evidence.

Sustained Post-Developmental T-Bet Expression Is Critical for the Maintenance of Type One Innate Lymphoid Cells In Vivo

Innate lymphoid cells (ILC) play a significant role in the intestinal immune response and T-bet+ CD127+ group 1 cells (ILC1) have been linked to the pathogenesis of human inflammatory bowel disease (IBD). However, the functional importance of ILC1 in the context of an intact adaptive immune response has been controversial. In this report we demonstrate that induced depletion of T-bet using a Rosa26-Cre-ERT2 model resulted in the loss of intestinal ILC1, pointing to a post-developmental requirement of T-bet expression for these cells. In contrast, neither colonic lamina propria (cLP) ILC2 nor cLP ILC3 abundance were altered upon induced deletion of T-bet. Mechanistically, we report that STAT1 or STAT4 are not required for intestinal ILC1 development and maintenance. Mice with induced deletion of T-bet and subsequent loss of ILC1 were protected from the induction of severe colitis in vivo. Hence, this study provides support for the clinical development of an IBD treatment based on ILC1 depletion via targeting T-bet or its downstream transcriptional targets.

Global shifts in species richness have shaped carpet shark evolution

Background The evolutionary processes that shape patterns of species richness in marine ecosystems are complex and may differ between organismal groups. There has been considerable interest in understanding the evolutionary processes that led to marine species richness being concentrated in specific geographical locations. In this study we focus on the evolutionary history of a group of small-to-medium sized sharks known as carpet sharks. While a few carpet shark species are widespread, the majority of carpet shark species richness is contained within a biodiversity hotspot at the boundary of the Indian and Pacific oceans. We address the significance of this biodiversity hotspot in carpet shark evolution and speciation by leveraging a rich fossil record and molecular phylogenetics to examine the prehistoric distribution of carpet sharks. Results We find that carpet sharks species richness was greatest in shallow seas connected to the Atlantic Ocean during the Late Cretaceous, but that there was a subsequent loss of biodiversity in Atlantic waters. Fossil evidence from sites in close geographic proximity to the current center of carpet shark diversity are generally restricted to younger geologic strata. Conclusions From this data we conclude that (1) center of carpet shark biodiversity has shifted during the last 100 million years, (2) carpet sharks have repeatedly dispersed to nascent habitat (including to their current center of diversity), and (3) the current center of carpet shark biodiversity conserves lineages that have been extirpated from this prehistoric range and is a source of new carpet shark species. Our findings provide insights into the roles of marine biodiversity hotspots for higher-tropic level predators and the methods applied here can be used for additional studies of shark evolution.

Mouse model and human patient data suggest critical roles for Pten and p53 in suppressing POLE mutant tumor development

Mutations in the exonuclease domain of POLE are associated with tumors harboring very high mutation burdens. The mechanisms linking this significant mutation accumulation and tumor development remain poorly understood. Pole+/P286R;Trp53+/- mice showed accelerated cancer mortality compared to Pole+/P286R;Trp53+/+ mice. Cells from Pole+/P286R mice showed increased p53 activation, and subsequent loss of p53 permitted rapid growth, implicating canonical p53 loss of heterozygosity in POLE mutant tumor growth. Somewhat surprisingly, however, p53 status had no effect on tumor mutation burden or single base substitution signatures in POLE mutant tumors from mice or humans. Pten has important roles in maintaining genome stability. We find that PTEN mutations are highly enriched in human POLE mutant tumors, including many in POLE signature contexts. One such signature mutation, PTEN-F341V, was previously shown in a mouse model to specifically decrease nuclear Pten and lead to increased DNA damage. We found tumors in Pole+/P286R mice that spontaneously acquired PtenF341V mutations and were associated with significantly reduced nuclear Pten and elevated DNA damage. Taken together with recent published work, our results support the idea that POLE-mediated hypermutagenesis is necessary, but not entirely sufficient, for tumorigenesis. Disabling surveillance of nuclear DNA damage is a likely sufficient factor.

The Global Fund: why anti-corruption, transparency and accountability matter

Background The creation of the Global Fund to Fight AIDS, Tuberculosis and Malaria, also known as the Global Fund, was prompted by the lack of a timely and effective global response, and the need for financing to fight against three devastating diseases: HIV/AIDS, tuberculosis, and malaria. During the formation of the Global Fund, necessary anti-corruption, transparency, and accountability (ACTA) structures were not put in place to prevent fraud and corruption in its grants, which resulted in the misuse of funds by grant recipients and an eventual loss of donor confidence in 2011. The Global Fund has instituted various ACTA mechanisms to address this misuse of funding and the subsequent loss of donor confidence, and this paper seeks to understand these implementations and their impacts over the past decade, in an effort to probe ACTA more deeply. Results By restructuring the governing committees in 2011, and the Audit and Finance; Ethics and Governance; and Strategy Committees in 2016, the Global Fund has delineated committee mandates and strengthened the Board’s oversight of operations. Additionally, the Global Fund has adopted a rigorous risk management framework which it has worked into all aspects of its functioning. An Ethics and Integrity Framework was adopted in 2014 and an Ethics Office was established in 2016, resulting in increased conflict of interest disclosures and greater considerations of ethics within the organization. The Global Fund’s Office of the Inspector General (OIG) has effectively performed internal and external audits and investigations on fraud and corruption, highlighted potential risks for mitigation, and has implemented ACTA initiatives, such as the I Speak Out Now! campaign to encourage whistleblowing and educate on fraud and corruption. Conclusions From 2011 onwards, the Global Fund has developed a number of ACTA mechanisms which, in particular, resulted in reduced grant-related risks and procurement fraud as demonstrated by the decreased classification from high to moderate in 2017, and the reduction of investigations in 2018 respectively. However, it is crucial that the Global Fund continues to evaluate the effectiveness of these mechanisms; monitor for potential perverse impacts; and make necessary changes, when and where they are needed.

Arthroscopic Repair of Batter’s Shoulder

Background: Batter’s shoulder is a condition in which the posterior labrum is typically torn during the baseball swinging motion, producing a traumatic tear and posterior instability. The injury commonly occurs in the batter’s lead shoulder due to repetitive microtrauma, raising concern for switch-hitters due to the cumulative stress of throwing and swinging on the lead shoulder. Instability is commonly caused by a posterior humeral force and relative shoulder adduction, which is most prevalent during a swing attempt at a low and outside pitch. Indications: Damage to the labrum during the acute traumatic event can cause residual pain and recurrent instability of the shoulder. Indications include failed conservative management. The patient demonstrated a full-thickness longitudinal tear that was grossly unstable with gentle probing. Technique Description: After establishing presence of an unstable posterior labral tear during diagnostic arthroscopy, a 7-o’clock portal is established for the labral repair. A knotless suture anchor construct was utilized for its low-profile features. Care is taken to avoid both tangling of sutures and overtensioning of the repair. Results: Patients return to live batting practice at 6 months postoperatively and most patients return to the same level of play following surgical management. Discussion/Conclusion: Avoid overtightening of labral repair and subsequent loss of range of motion. Use of a low-profile knotless suture anchor is the senior author’s preferred method of surgical management. A majority of patients surgically managed for unstable posterior labral injuries return to the same level of play.

INJECTION SITE LIPODYSTROPHY WITH FAT NECROSIS: A RARE CASE REPORT

Lipodystrophies are dermatological complications due to repetitive subcutaneous injections leading to subsequent loss of adipose tissue at the site of injection. Fat necrosis occursdue to concentrated pressure on microlobules of adipocytes causing disruption of septa between these microlobules, leading to permanent changes in the injured site. In this report we present a case of 63 year old female who developed lipodystrophy following subcutaneous injection in left lower abdomen.

Primaries will launch tense Argentine electoral cycle

Significance The primaries, pushed back from this month to September 12, will generate few surprises but offer an opportunity to gauge the relative support of each coalition ahead of the midterm elections in which half of the 257-seat Lower House and 24 of 54 Senate seats will be contested. Impacts The next presidential contest may well be between two candidates whose rejection rates far outweigh their approval. The incumbent is unlikely to be the government’s presidential candidate in 2023. Government moderates, including the president, will be blamed for a poor election result and for any subsequent loss of support.