Fission yeast 26S proteasome mutants are multi-drug resistant due to stabilization of the pap1 transcription factor

Here we report the result of a genetic screen for mutants resistant to the microtubule poison methyl benzimidazol-2-yl carbamate (MBC) that were also temperature sensitive for growth. In total the isolated mutants were distributed in ten complementation groups. Cloning experiments revealed that most of the mutants were in essential genes encoding various 26S proteasome subunits. We found that the proteasome mutants are multi-drug resistant due to stabilization of the stress-activated transcription factor Pap1. We show that the ubiquitylation and ultimately the degradation of Pap1 depend on the Rhp6/Ubc2 E2 ubiquitin conjugating enzyme and the Ubr1 E3 ubiquitin-protein ligase. Accordingly, mutants lacking Rhp6 or Ubr1 display drug-resistant phenotypes

Gene expression profiles derived from fine needle aspiration correlate with response to systemic chemotherapy in breast cancer

BACKGROUND: Drug resistance in breast cancer is a major obstacle to successful chemotherapy. In this study we used cDNA microarray technology to examine gene expression profiles obtained from fine needle aspiration (FNA) of primary breast tumors before and after systemic chemotherapy. Our goal was to determine the feasibility of obtaining representative expression array profiles from limited amounts of tissue and to identify those expression profiles that correlate with treatment response. METHODS: Repeat presurgical FNA samples were taken from six patients who were to undergo primary surgical treatment. Additionally, a group of 10 patients who were to receive neoadjuvant chemotherapy underwent two FNAs before chemotherapy (adriamycin 60 mg/m(2) and cyclophosphamide 600 mg/m(2)) followed by another FNA on day 21 after the first cycle. Total RNA was amplified with T7 Eberwine's procedure and labeled cDNA was hybridized onto a 7600-feature glass cDNA microarray. RESULTS: We identified candidate gene expression profiles that might distinguish tumors with complete response to chemotherapy from tumors that do not respond, and found that the number of genes that change after one cycle of chemotherapy was 10 times greater in the responding group than in the non-responding group. CONCLUSION: This study supports the suitability of FNA-derived cDNA microarray expression profiling of breast cancers as a comprehensive genomic approach for studying the mechanisms of drug resistance. Our findings also demonstrate the potential of monitoring post-chemotherapy changes in expression profiles as a measure of pharmacodynamic effect and suggests that these approaches might yield useful results when validated by larger studies

Biallelic null variants in PNPLA8 cause microcephaly by reducing the number of basal radial glia.

Patatin-like phospholipase domain-containing lipase 8 (PNPLA8), one of the calcium-independent phospholipase A2 enzymes, is involved in various physiological processes through the maintenance of membrane phospholipids. Biallelic variants in PNPLA8 have been associated with a range of paediatric neurodegenerative disorders. However, the phenotypic spectrum, genotype-phenotype correlations and the underlying mechanisms are poorly understood. Here, we newly identified 14 individuals from 12 unrelated families with biallelic ultra-rare variants in PNPLA8 presenting with a wide phenotypic spectrum of clinical features. Analysis of the clinical features of current and previously reported individuals (25 affected individuals across 20 families) showed that PNPLA8-related neurological diseases manifest as a continuum ranging from variable developmental and/or degenerative epileptic-dyskinetic encephalopathy to childhood-onset neurodegeneration. We found that complete loss of PNPLA8 was associated with the more profound end of the spectrum, with congenital microcephaly. Using cerebral organoids generated from human induced pluripotent stem cells, we found that loss of PNPLA8 led to developmental defects by reducing the number of basal radial glial cells and upper-layer neurons. Spatial transcriptomics revealed that loss of PNPLA8 altered the fate specification of apical radial glial cells, as reflected by the enrichment of gene sets related to the cell cycle, basal radial glial cells and neural differentiation. Neural progenitor cells lacking PNPLA8 showed a reduced amount of lysophosphatidic acid, lysophosphatidylethanolamine and phosphatidic acid. The reduced number of basal radial glial cells in patient-derived cerebral organoids was rescued, in part, by the addition of lysophosphatidic acid. Our data suggest that PNPLA8 is crucial to meet phospholipid synthetic needs and to produce abundant basal radial glial cells in human brain development

A study on the diffusion-induced grain boundary migration ahead of stress corrosion cracking crack tips through advanced characterization

Three austenitic alloys with different Ni content were stress corrosion cracking (SCC) tested in simulated pressurized water reactor (PWR) primary water at 320 and 360ºC. Diffusion-induced grain boundary migration (DIGM) associated with preferential intergranular oxidation (PIO) is observed ahead of all SCC crack tips. The occurrence of DIGM is revealed to be driven by PIO-induced diffusion of Cr, Fe, and Ni. The extent of DIGM is controlled by the lateral and in-depth elemental diffusion and PIO, with these processes interconnected. Temperature and alloy composition are revealed to affect the extent of DIGM by affecting these processes

A mechanistic study of SCC in Alloy 600 through high-resolution characterization

High-resolution characterization was used to understand the mechanisms controlling stress corrosion cracking (SCC) in Alloy 600 exposed to simulated PWR primary water conditions. Three potential active crack tips obtained from different types of grain boundaries were studied and compared. The results suggest that the dominant mechanism controlling SCC propagation is intergranular internal oxidation. The applied stress, pre-existent residual strain, the accumulation of defects around the crack tip, the formation of a Fe-Cr-depleted zone, and a porous intergranular oxide are acknowledged as necessary precursors to SCC. Based on the results obtained in this study, a model of SCC propagation is proposed

3D Atom-Probe Characterization of Stress and Cold-Work in Stress Corrosion Cracking of 304 Stainless Steel

Cold-worked 304 stainless steels (SS) are known to be susceptible to stress corrosion cracking (SCC). This study employs atom-probe tomography (APT) for local chemical analysis of the oxides formed. Autoclave experiments on a set of samples with/without cold-work prior to oxidation, and with/without stress applied during oxidation, were carried out under simulated pressurised water reactor (PWR) primary conditions. APT and analytical transmission electron microscopy (ATEM) were combined to investigate chemical and structural implications of surface and grain boundary oxidation in 304 SS. Focussed ion beam (FIB) milling was used to prepare specimens containing the same grain boundary for every analysis technique. Grain boundary and deformation band oxidation were observed in all but the unstressed and non-cold worked sample. Cavities were found ahead of the Cr-rich oxide in some of the samples. APT data suggests the presence of hydrogen in Nickel-rich regions