Genetic interaction mapping informs integrative structure determination of protein complexes

Determining structures of protein complexes is crucial for understanding cellular functions. Here, we describe an integrative structure determination approach that relies on in vivo measurements of genetic interactions. We construct phenotypic profiles for point mutations crossed against gene deletions or exposed to environmental perturbations, followed by converting similarities between two profiles into an upper bound on the distance between the mutated residues. We determine the structure of the yeast histone H3-H4 complex based on similar to 500,000 genetic interactions of 350 mutants. We then apply the method to subunits Rpb1-Rpb2 of yeast RNA polymerase II and subunits RpoB-RpoC of bacterial RNA polymerase. The accuracy is comparable to that based on chemical cross-links; using restraints from both genetic interactions and cross-links further improves model accuracy and precision. The approach provides an efficient means to augment integrative structure determination with in vivo observations

Some pioneers of European human genetics

Some of the pioneers of human genetics across Europe are described, based on a series of 100 recorded interviews made by the author. These interviews, and the memories of earlier workers in the field recalled by interviewees, provide a vivid picture, albeit incomplete, of the early years of human and medical genetics. From small beginnings in the immediate post-World War 2 years, human genetics grew rapidly across many European countries, a powerful factor being the development of human cytogenetics, stimulated by concerns over the risks of radiation exposure. Medical applications soon followed, with the recognition of human chromosome abnormalities, the need for genetic counselling, the possibility of prenatal diagnosis and later, the applications of human molecular genetics. The evolution of the field has been strongly influenced by the characters and interests of the relatively small number of founding workers in different European countries, as well as by wider social, medical and scientific factors in the individual countries

Chk2 and p53 Are Haploinsufficient with Dependent and Independent Functions to Eliminate Cells after Telomere Loss

The mechanisms that cells use to monitor telomere integrity, and the array of responses that may be induced, are not fully defined. To date there have been no studies in animals describing the ability of cells to survive and contribute to adult organs following telomere loss. We developed assays to monitor the ability of somatic cells to proliferate and differentiate after telomere loss. Here we show that p53 and Chk2 limit the growth and differentiation of cells that lose a telomere. Furthermore, our results show that two copies of the genes encoding p53 and Chk2 are required for the cell to mount a rapid wildtype response to a missing telomere. Finally, our results show that, while Chk2 functions by activating the p53-dependent apoptotic cascade, Chk2 also functions independently of p53 to limit survival. In spite of these mechanisms to eliminate cells that have lost a telomere, we find that such cells can make a substantial contribution to differentiated adult tissues

Polo kinase recruitment via the constitutive centromere-associated network at the kinetochore elevates centromeric RNA

The kinetochore, a multi-protein complex assembled on centromeres, is essential to segregate chromosomes during cell division. Deficiencies in kinetochore function can lead to chromosomal instability and aneuploidy-a hallmark of cancer cells. Kinetochore function is controlled by recruitment of regulatory proteins, many of which have been documented, however their function often remains uncharacterized and many are yet to be identified. To identify candidates of kinetochore regulation we used a proteome-wide protein association strategy in budding yeast and detected many proteins that are involved in post-translational modifications such as kinases, phosphatases and histone modifiers. We focused on the Polo-like kinase, Cdc5, and interrogated which cellular components were sensitive to constitutive Cdc5 localization. The kinetochore is particularly sensitive to constitutive Cdc5 kinase activity. Targeting Cdc5 to different kinetochore subcomplexes produced diverse phenotypes, consistent with multiple distinct functions at the kinetochore. We show that targeting Cdc5 to the inner kinetochore, the constitutive centromere-associated network (CCAN), increases the levels of centromeric RNA via an SPT4 dependent mechanism

Global overview of the management of acute cholecystitis during the COVID-19 pandemic (CHOLECOVID study)

Background: This study provides a global overview of the management of patients with acute cholecystitis during the initial phase of the COVID-19 pandemic. Methods: CHOLECOVID is an international, multicentre, observational comparative study of patients admitted to hospital with acute cholecystitis during the COVID-19 pandemic. Data on management were collected for a 2-month study interval coincident with the WHO declaration of the SARS-CoV-2 pandemic and compared with an equivalent pre-pandemic time interval. Mediation analysis examined the influence of SARS-COV-2 infection on 30-day mortality. Results: This study collected data on 9783 patients with acute cholecystitis admitted to 247 hospitals across the world. The pandemic was associated with reduced availability of surgical workforce and operating facilities globally, a significant shift to worse severity of disease, and increased use of conservative management. There was a reduction (both absolute and proportionate) in the number of patients undergoing cholecystectomy from 3095 patients (56.2 per cent) pre-pandemic to 1998 patients (46.2 per cent) during the pandemic but there was no difference in 30-day all-cause mortality after cholecystectomy comparing the pre-pandemic interval with the pandemic (13 patients (0.4 per cent) pre-pandemic to 13 patients (0.6 per cent) pandemic; P = 0.355). In mediation analysis, an admission with acute cholecystitis during the pandemic was associated with a non-significant increased risk of death (OR 1.29, 95 per cent c.i. 0.93 to 1.79, P = 0.121). Conclusion: CHOLECOVID provides a unique overview of the treatment of patients with cholecystitis across the globe during the first months of the SARS-CoV-2 pandemic. The study highlights the need for system resilience in retention of elective surgical activity. Cholecystectomy was associated with a low risk of mortality and deferral of treatment results in an increase in avoidable morbidity that represents the non-COVID cost of this pandemic