The elucidation of the organization and maintenance of the plasma membrane has been sought due to its numerous roles in cellular function. In the budding yeast Saccharomyces cerevisiae, a novel paradigm has begun to emerge in the understanding of the distribution of plasma membrane microdomains and how they are regulated. We aimed to investigate the dynamic interdependence between the protein complexes eisosome and TORC2, representing microdomains MCC and MCT, respectively. In this study, we reveal that the eisosome organizer Pil1 colocalizes with the MCT marker Avo2. Furthermore, we provide evidence that the formation of MCT is dependent on both eisosome integrity and adequate levels of the plasma membrane phosphoinositide PI(4,5)P2. Taken together, our findings indicate that TORC2, eisosomes, and PI(4,5)P2 exist in an interconnected relationship, which supports the emerging model of the plasma membrane.

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Carbon nanomaterials (CNMs), which include carbon nanotubes (CNTs) and their derivatives, have diverse technological and biomedical applications. The potential toxicity of CNMs to cells and tissues has become an important emerging question in nanotechnology. To assess the toxicity of CNTs and fullerenol C60(OH)24, we in the present work used the budding yeast Saccharomyces cerevisiae, one of the simplest eukaryotic organisms that share fundamental aspects of eukaryotic cell biology. We found that treatment with CNMs, regardless of their physical shape, negatively affected the growth rates, end-point cell densities and doubling times of CNM-exposed yeast cells when compared to unexposed cells. To investigate potential mechanisms behind the CNMs-induced growth defects, we performed RNA-Seq dependent transcriptional analysis and constructed global gene expression profiles of fullerenol C60(OH)24- and CNT-treated cells. When compared to non-treated control cells, CNM-treated cells displayed differential expression of genes whose functions are implicated in membrane transporters and stress response, although differentially expressed genes were not consistent between CNT- and fullerenol C60(OH)24-treated groups, leading to our conclusion that CNMs could serve as environmental toxic factors to eukaryotic cells.

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Background Influenza (flu) infections affect a large subset of the population every year and have significant impacts on the health of patients, especially those with weak or compromised immune systems such as the elderly, children, cancer patients, and transplant recipients. Baloxavir marboxil was approved in October 2018 as a novel antiviral therapeutic for treating flu. During clinical trials, mutations were identified at the I28 codon of the polymerase acidic (PA) protein that greatly increased the resistance of a flu strain to this novel drug. In this study, a qPCR was developed and validated to identify these resistance mutations, allowing for guided therapy based on the resistance profile of the strain. Methods Flu A sequences (6,175) of the PA gene from the NCBI Influenza Virus Database collected over the last 5 years were compiled and aligned. Primers and probes were designed to target the I38 codon of the PA gene, and specific probes for each codon yielding a resistant amino acid mutation (I38T, -M, and -F) were designed. Locked nucleic acid (LNA) bases were used to increase the specificity of the probes. A combination of clinical flu specimens, laboratory strains, and synthetic constructs of each potential resistance mutation were used to validate the precision, sensitivity, and accuracy of the assay in nasopharyngeal swabs. Results Precision of the cycle threshold (Ct) values for each detector was determined to have a standard deviation of less than 3 for inter-assay and less than 2 for intra-assay replicates. Sensitivity was determined to be 800 copies/mL in nasopharyngeal swabs. Accuracy was found to be 92.3%. A single laboratory strain from the H1N1 2009 epidemic showed cross-reactivity with both wild-type and resistant probes, but no circulating clinical H1N1 samples tested showed this response. Conclusion The precision, sensitivity, and accuracy of a qPCR for resistance mutations to baloxavir marboxil support this assay’s utility as an aid in the treatment of flu in at-risk patient groups. This assay allows for rapid detection (<24 hours) of resistance markers to aid clinicians in improving flu case outcomes. Disclosures All authors: No reported disclosures.