Cell cortex composition and homeostasis resolved by integrating proteomics and quantitative imaging.

The cellular actin cortex is the cytoskeletal structure primarily responsible for the control of animal cell shape and as such plays a central role in cell division, migration, and tissue morphogenesis. Due to the lack of experimental systems where the cortex can be investigated independently from other organelles, little is known about its composition, assembly, and homeostasis. Here, we describe novel tools to resolve the composition and regulation of the cortex. We report and validate a protocol for cortex purification based on the separation of cellular blebs. Mass spectrometry analysis of purified cortices provides a first extensive list of cortical components. To assess the function of identified proteins, we design an automated imaging assay for precise quantification of cortical actomyosin assembly dynamics. We show subtle changes in cortex assembly dynamics upon depletion of the identified cortical component profilin. Our widely applicable integrated method paves the way for systems-level investigations of the actomyosin cortex and its regulation during morphogenesis. © 2013 Wiley Periodicals, Inc

Area deprivation and regional disparities in treatment and outcome quality of 29,284 pediatric patients with type 1 diabetes in Germany: A cross-sectional multicenter DPV analysis.

OBJECTIVEThis study analyzed whether area deprivation is associated with disparities in health care of pediatric type 1 diabetes in Germany.RESEARCH DESIGN AND METHODSWe selected patients <20 years of age with type 1 diabetes and German residence documented in the "diabetes patient follow-up" (Diabetes-Patienten-Verlaufsdokumentation [DPV]) registry for 2015/2016. Area deprivation was assessed by quintiles of the German Index of Multiple Deprivation (GIMD 2010) at the district level and was assigned to patients. To investigate associations between GIMD 2010 and indicators of diabetes care, we used multivariable regression models (linear, logistic, and Poisson) adjusting for sex, age, migration background, diabetes duration, and German federal state.RESULTSWe analyzed data from 29,284 patients. From the least to the most deprived quintile, use of continuous glucose monitoring systems (CGMS) decreased from 6.3 to 3.4% and use of long-acting insulin analogs from 80.8 to 64.3%, whereas use of rapid-acting insulin analogs increased from 74.7 to 79.0%; average HbA(1c) increased from 7.84 to 8.07% (62 to 65 mmol/mol), and the prevalence of overweight from 11.8 to 15.5%, but the rate of severe hypoglycemia decreased from 12.1 to 6.9 events/100 patient-years. Associations with other parameters showed a more complex pattern (use of continuous subcutaneous insulin infusion [CSII]) or were not significant.CONCLUSIONSArea deprivation was associated not only with key outcomes in pediatric type 1 diabetes but also with treatment modalities. Our results show, in particular, that the access to CGMS and CSII could be improved in the most deprived regions in Germany

Longitudinal trajectories of BMI z-score: an international comparison of 11,513 Australian, American and German/Austrian/Luxembourgian youth with type 1 diabetes

BACKGROUND:BMI fluctuations during puberty are common. Data on individual change in BMI from childhood to young adulthood are limited in youth with type 1 diabetes. OBJECTIVES:To compare longitudinal trajectories of body mass index z score (BMIz) from childhood to adolescence across three registries spanning five countries. METHODS:Data sources: T1DX (USA), DPV (Germany/Austria/Luxembourg) and ADDN (Australia). The analysis included 11,513 youth with type 1 diabetes, duration >1 year, at least one BMI measure at baseline (age 8-10 years) and >5 aggregated BMI measures by year of age during follow-up until age 17 years. BMIz was calculated based on WHO charts. Latent class growth modelling was used to identify subgroups following a similar trajectory of BMIz over time. RESULTS:Five distinct trajectories of BMIz were present in the T1DX and ADDN cohorts, while six trajectories were identified in the DPV cohort. Boys followed more often a low/near-normal pattern while elevated BMIz curves were more likely in girls (ADDN; DPV). For T1DX cohort, no sex differences were observed. Comparing the reference group (BMIz ~0) with the other groups during puberty, higher BMIz was significantly associated with older age at T1D onset, racial/ethnic minority and elevated HbA1c (all p<0.05). CONCLUSION:This multinational study presents unique BMIz trajectories in youth with T1D across three continents. The prevalence of overweight and the longitudinal persistence of overweight support the need for close monitoring of weight and nutrition in this population. The international and individual differences likely result from diverse genetic, environmental and therapeutic factors.Helen Phelan, Nicole C. Foster, Anke Schwandt, Jennifer J. Couper, Steven Willi, Peter Kroschwald ... et al

RNA Controls PolyQ Protein Phase Transitions

Compartmentalization in cells is central to the spatial and temporal control of biochemistry. In addition to membrane-bound organelles, membrane-less compartments form partitions in cells. Increasing evidence suggests that these compartments assemble through liquid-liquid phase separation. However the spatiotemporal control of their assembly, and how they maintain distinct functional and physical identities is poorly understood. We have previously shown an RNA-binding protein with a polyQ-expansion called Whi3 is essential for the spatial patterning of cyclin and formin transcripts in cytosol. Here, we show that specific mRNAs that are known physiological targets of Whi3 drive phase separation. mRNA can alter the viscosity of droplets, their propensity to fuse, and the exchange rates of components with bulk solution. Different mRNAs impart distinct biophysical properties of droplets indicating mRNA can bring individuality to assemblies. Our findings suggest that mRNAs can encode not only genetic information, but also the biophysical properties of phase-separated compartments

Coexisting Liquid Phases Underlie Nucleolar Subcompartments

The nucleolus and other ribonucleoprotein (RNP) bodies are membrane-less organelles that appear to assemble through phase separation of their molecular components. However, many such RNP bodies contain internal sub-compartments, and the mechanism of their formation remains unclear. Here, we combine in vivo and in vitro studies, together with computational modeling, to show that sub-compartments within the nucleolus represent distinct, coexisting liquid phases. Consistent with their in vivo immiscibility, purified nucleolar proteins phase separate into droplets containing distinct non-coalescing phases that are remarkably similar to nucleoli in vivo. This layered droplet organization is caused by differences in the biophysical properties of the phases – particularly droplet surface tension – which arises from sequence-encoded features of their macromolecular components. These results suggest that phase separation can give rise to multilayered liquids that may facilitate sequential RNA processing reactions in a variety of RNP bodies