MPN+, a putative catalytic motif found in a subset of MPN domain proteins from eukaryotes and prokaryotes, is critical for Rpn11 function

BACKGROUND: Three macromolecular assemblages, the lid complex of the proteasome, the COP9-Signalosome (CSN) and the eIF3 complex, all consist of multiple proteins harboring MPN and PCI domains. Up to now, no specific function for any of these proteins has been defined, nor has the importance of these motifs been elucidated. In particular Rpn11, a lid subunit, serves as the paradigm for MPN-containing proteins as it is highly conserved and important for proteasome function. RESULTS: We have identified a sequence motif, termed the MPN+ motif, which is highly conserved in a subset of MPN domain proteins such as Rpn11 and Csn5/Jab1, but is not present outside of this subfamily. The MPN+ motif consists of five polar residues that resemble the active site residues of hydrolytic enzyme classes, particularly that of metalloproteases. By using site-directed mutagenesis, we show that the MPN+ residues are important for the function of Rpn11, while a highly conserved Cys residue outside of the MPN+ motif is not essential. Single amino acid substitutions in MPN+ residues all show similar phenotypes, including slow growth, sensitivity to temperature and amino acid analogs, and general proteasome-dependent proteolysis defects. CONCLUSIONS: The MPN+ motif is abundant in certain MPN-domain proteins, including newly identified proteins of eukaryotes, bacteria and archaea thought to act outside of the traditional large PCI/MPN complexes. The putative catalytic nature of the MPN+ motif makes it a good candidate for a pivotal enzymatic function, possibly a proteasome-associated deubiquitinating activity and a CSN-associated Nedd8/Rub1-removing activity

Gender bias in the evaluation of interns in different medical specialties: An archival study

Introduction The field of medicine is characterized by within-field gender segregation: Gender ratios vary systematically by subdisciplines. This segregation might be, in part, due to gender bias in the assessment of women and men medical doctors. Methods We examined whether the assessments, i.e. overall score, department scores and skills scores, interns receive by their superiors during their internship year, vary as a function of their gender and the representation of women in the field. We analyzed an archival data set from a large hospital in Israel which included 3326 assessments that were given to all interns who completed their internship year between 2015 and 2019. Results Women received lower department scores and skills scores in fields with a low (versus high) representation of women. Men received higher scores in fields with a high (versus low) representation of men, yet there was no difference in their skills scores. Conclusions Women are evaluated more negatively in fields with a low representation of women doctors. Similarly, men are evaluated more negatively in fields with a low representation of men, yet this cannot be explained by their skills. This pattern of results might point to a gender bias in assessments. A better understanding of these differences is important as assessments affect interns’ career choices and options

Rumen cellulosomics : divergent fiber-degrading strategies revealed by comparative genome-wide analysis of six ruminococcal strains

Peer reviewedPublisher PD

Robust nuclear lamina-based cell classification of aging and senescent cells

Changes in the shape of the nuclear lamina are exhibited in senescent cells, as well as in cells expressing mutations in lamina genes. To identify cells with defects in the nuclear lamina we developed an imaging method that quantifies the intensity and curvature of the nuclear lamina. We show that this method accurately describes changes in the nuclear lamina. Spatial changes in nuclear lamina coincide with redistribution of lamin A proteins and local reduction in protein mobility in senescent cell. We suggest that local accumulation of lamin A in the nuclear envelope leads to bending of the structure. A quantitative distinction of the nuclear lamina shape in cell populations was found between fresh and senescent cells, and between primary myoblasts from young and old donors. Moreover, with this method mutations in lamina genes were significantly distinct from cells with wild-type genes. We suggest that this method can be applied to identify abnormal cells during aging, in in vitro propagation, and in lamina disorders

Molecular Image Analysis: Quantitative Description and Classification of the Nuclear Lamina in Human Mesenchymal Stem Cells

The nuclear lamina is an intermediate filament network that provides a structural framework for the cell nucleus. Changes in lamina structure are found during changes in cell fate such as cell division or cell death and are associated with human diseases. An unbiased method that quantifies changes in lamina shape can provide information on cells undergoing changes in cellular functions. We have developed an image processing methodology that finds and quantifies the 3D structure of the nuclear lamina. We show that measurements on such images can be used for cell classification and provide information concerning protein spatial localization in this structure. To demonstrate the efficacy of this method, we compared the lamina of unmanipulated human mesenchymal stem cells (hMSCs) at passage 4 to cells activated for apoptosis. A statistically significant classification was found between the two populations

Four-YearFollow-upof [F-18]Fluorodeoxyglucose Positron Emission Tomography-Based Parkinson's Disease-Related Pattern Expression in 20 Patients With Isolated Rapid Eye Movement Sleep Behavior Disorder Shows Prodromal Progression

Background: Isolated rapid eye movement sleep behavior disorder is known to be prodromal for alpha-synucleinopathies, such as Parkinson's disease (PD) and dementia with Lewy bodies. The [18F]fluorodeoxyglucose-positron emission tomography (PET)–based PD-related brain pattern can be used to monitor disease progression. Objective: We longitudinally investigated PD-related brain pattern expression changes in 20 subjects with isolated rapid eye movement sleep behavior disorder to investigate whether this may be a suitable technique to study prodromal PD progression in these patients and to identify potential phenoconverters. Methods: Subjects underwent two [18F]fluorodeoxyglucose-PET brain scans ~3.7 years apart, along with baseline and repeated motor, cognitive, and olfactory testing within roughly the same time frame. Results: At baseline, 8 of 20 (40%) subjects significantly expressed the PD-related brain pattern (with z scores above the receiver operating characteristic–determined threshold). At follow-up, six additional subjects exhibited significant PD-related brain pattern expression (70% in total). PD-related brain pattern expression increased in all subjects (P = 0.00008). Four subjects (20%), all with significant baseline PD-related brain pattern expression, phenoconverted to clinical PD. Conclusions: Suprathreshold PD-related brain pattern expression and greater score rate of change may signify greater shorter-term risk for phenoconversion. Our results support the use of serial PD-related brain pattern expression measurements as a prodromal PD progression biomarker in patients with isolated rapid eye movement sleep behavior disorder

Integrated metabolomics identifies CYP72A67 and CYP72A68 oxidases in the biosynthesis of Medicago truncatula oleanate sapogenins

Introduction: Triterpene saponins are important bioactive plant natural products found in many plant families including the Leguminosae. Objectives: We characterize two Medicago truncatula cytochrome P450 enzymes, MtCYP72A67 and MtCYP72A68, involved in saponin biosynthesis including both in vitro and in planta evidence. Methods: UHPLC-(-)ESI-QToF-MS was used to profile saponin accumulation across a collection of 106 M. truncatula ecotypes. The profiling results identified numerous ecotypes with high and low saponin accumulation in root and aerial tissues. Four ecotypes with significant differential saponin content in the root and/or aerial tissues were selected, and correlated gene expression profiling was performed. Results: Correlation analyses between gene expression and saponin accumulation revealed high correlations between saponin content with gene expression of -amyrin synthase, MtCYP716A12, and two cytochromes P450 genes, MtCYP72A67 and MtCYP72A68. In vivo and in vitro biochemical assays using yeast microsomes containing MtCYP72A67 revealed hydroxylase activity for carbon 2 of oleanolic acid and hederagenin. This finding was supported by functional characterization of MtCYP72A67 using RNAi-mediated gene silencing in M. truncatula hairy roots, which revealed a significant reduction of 2-hydroxylated sapogenins. In vivo and in vitro assays with MtCYP72A68 produced in yeast showed multifunctional oxidase activity for carbon 23 of oleanolic acid and hederagenin. These findings were supported by overexpression of MtCYP72A68 in M. truncatula hairy roots, which revealed significant increases of oleanolic acid, 2-hydroxyoleanolic acid, hederagenin and total saponin levels. Conclusions: The cumulative data support that MtCYP72A68 is a multisubstrate, multifunctional oxidase and MtCYP72A67 is a 2-hydroxylase, both of which function during the early steps of triterpene-oleanate sapogenin biosynthesis