Characterization of Phosphorylated G Protein Function and Membrane Culstering by Super Resolution Imaging

Heterotrimeric G proteins play crucial roles in various signal transduction pathways, where they act as molecular switches in transducing a signal from G protein coupled receptors (GPCRs) at the plasma membrane to downstream effectors. Although their mechanism of action is mostly concentrated at the plasma membrane, their dynamic membrane organization and how it is regulated are not understood. Due to the diffraction limited resolution of fluorescence microscopy, studying the precise organization of membrane proteins can be challenging. In this study, we took advantage of super-resolution fluorescence photoactivation localization microscopy (FPALM) to overcome this challenge. Dictyostelium discoideum was used as a cellular model to study G protein function and membrane organization. These cells rely on chemotaxis toward a secreted chemoattractant, cyclic adenosine monophosphate (cAMP) during the development phase of their life cycle. The Gα2 subunit of D. discoideum is required for the chemotactic response. Once activation occurs, Gα2 is known to be phosphorylated on serine 113; however, the role of this phosphorylation remains poorly defined. Exchange of serine residue 113 to alanine causes starved cells to begin the aggregation phase several hours sooner when compared to wild type, while exchanging this serine to aspartic acid (phosphorylation mimic) shows a dramatic decrease in plasma membrane surface localization. At the nanoscale level, images using FPALM show that activation and phosphorylation cause significant changes to Gα2 cluster density in the plasma membrane. Getting these first nanoscale images of G protein provided robust information, which adds to our understanding of the ligand-dependent reorganization and clustering of Gα2 required for precise signaling. Cell fractionation experiments supported this result. In addition, phosphorylation-dependent interaction between phosphorylated Gα2 and D. discoideum 14-3-3 protein was detected

Membrane Localization of Palmitoylated G Proteins: Implication of Lipid Microdomains

Chemotaxis is a complex process that relies on various integrated signal cascades. Many of these signal pathways incorporate by the heterotrimeric guanosine nucleotide-binding proteins (G proteins). Lipid modification including the post-translational modification, palmitoylation can be considered as a key factor for proper G protein function. Various models have been used to gain a deeper understanding of the role of G proteins in cell chemotaxis. The soil amoeba, Dictyostelium discoideum, is well-known for its use in chemotaxis studies. During the development phase of its life cycle, Dictyostelium cells rely on their ability to performed chemotaxis toward the chemoattractant cyclic adenosine monophosphate (cAMP). Although significant details have already been worked out for this signaling and response pathway, little is known about how the G protein signal platform (lipid raft) may be involved. To elucidate how this microdomain may be involved in the G-protein-mediated chemotaxis signal, a low-density membrane fraction was isolated using a detergent-free method. In basal state, Ga2 was enriched in the low-density membrane and this was dependent on Ga2 acylation. Blocking Ga2 activation had no effect on this association, while activation shifted Ga2 from the low-density fraction to higher density fractions. Shifting of the activated-Ga2 to the 20% gradient fraction was under the control of F-actin. Our findings suggest that lipid microdomains are the proper environment for the palmitoylated Ga2 and activation leads them to leave these low-density domains and to accumulate in the heavier fractions

Knowledge, perceptions, and readiness of telepharmacy among community pharmacists

Telepharmacy is a practical part of telemedicine that refers to providing pharmaceutical services within the scope of the pharmacist's obligations while maintaining a temporal and spatial distance between patients, users of health services, and healthcare professionals. The present study was a cross-sectional study conducted among community pharmacists in Saudi Arabia between March and May 2022 to assess their knowledge, perceptions, and readiness for telepharmacy. The survey was filled out by 404 respondents. The majority of respondents were male (59.90%) and the age of more than half of them was between 30 and 39 years old (54.46%). Most participants worked in urban areas (83.66%), and 42.57% had less than five years of experience in a pharmacy. Most participants agreed that telepharmacy is available in Saudi Arabia (82.67%). Approximately 70% of pharmacists felt that telepharmacy promotes patient medication adherence, and 77.72% agreed that telepharmacy increases patient access to pharmaceuticals in rural areas. More than 72% of pharmacists said they would work on telepharmacy initiatives in rural areas for free, and 74.26% said they would work outside of usual working hours if necessary. In the future, this research could aid in adopting full-fledged telepharmacy pharmaceutical care services in Saudi Arabia. It could also help academic initiatives by allowing telepharmacy practice models to be included as a topic course in the curriculum to prepare future pharmacists to deliver telepharmacy services

Global, regional, and national burden of low back pain, 1990–2020, its attributable risk factors, and projections to 2050: a systematic analysis of the Global Burden of Disease Study 2021

Background: Low back pain is highly prevalent and the main cause of years lived with disability (YLDs). We present the most up-to-date global, regional, and national data on prevalence and YLDs for low back pain from the Global Burden of Diseases, Injuries, and Risk Factors Study 2021. Methods: Population-based studies from 1980 to 2019 identified in a systematic review, international surveys, US medical claims data, and dataset contributions by collaborators were used to estimate the prevalence and YLDs for low back pain from 1990 to 2020, for 204 countries and territories. Low back pain was defined as pain between the 12th ribs and the gluteal folds that lasted a day or more; input data using alternative definitions were adjusted in a network meta-regression analysis. Nested Bayesian meta-regression models were used to estimate prevalence and YLDs by age, sex, year, and location. Prevalence was projected to 2050 by running a regression on prevalence rates using Socio-demographic Index as a predictor, then multiplying them by projected population estimates. Findings: In 2020, low back pain affected 619 million (95% uncertainty interval 554–694) people globally, with a projection of 843 million (759–933) prevalent cases by 2050. In 2020, the global age-standardised rate of YLDs was 832 per 100 000 (578–1070). Between 1990 and 2020, age-standardised rates of prevalence and YLDs decreased by 10·4% (10·9–10·0) and 10·5% (11·1–10·0), respectively. A total of 38·8% (28·7–47·0) of YLDs were attributed to occupational factors, smoking, and high BMI. Interpretation: Low back pain remains the leading cause of YLDs globally, and in 2020, there were more than half a billion prevalent cases of low back pain worldwide. While age-standardised rates have decreased modestly over the past three decades, it is projected that globally in 2050, more than 800 million people will have low back pain. Challenges persist in obtaining primary country-level data on low back pain, and there is an urgent need for more high-quality, primary, country-level data on both prevalence and severity distributions to improve accuracy and monitor change. Funding: Bill and Melinda Gates Foundation