SMAC/DIABLO and HTRA2 are mitochondrial proteins whose amino-terminal sequences, known as inhibitor of apoptosis binding motifs (IBMs), bind activate ubiquitin ligases (IAPs), unleashing a cell's apoptotic potential. IBMs comprise four-residue, loose consensus sequence, to IAPs requires an unmodified amino terminus. Closely related, IBM-like N termini present in approximately 5% human proteins. We show that suppression the N-alpha-acetyltransferase NatA turns these cryptic sequences into...

Abstract CD93 is a highly glycosylated transmembrane protein expressed on monocytes, neutrophils, endothelial cells, and stem cells. Antibodies directed at modulate phagocytosis, CD93-deficient mice are defective in the clearance of apoptotic cells from inflamed peritoneum. In this study we observe that CD93, human monocytes susceptible to phorbol dibutyrate-induced ectodomain shedding time- dose-dependent manner. The soluble fragment found culture supernatant retains N-terminal carbohydrate...

Enolase is secreted by C. albicans and present in its biofilms although extracellular function unknown. Here we show that enolase mediates the colonization of small intestine mucosa albicans. Assays using intestinal disks adhesion inhibited, a dose dependent mode, either pretreatment epithelium with recombinant (70% at 0.5 mg/ml enolase) or yeasts anti-enolase antibodies (48% 20 µg antiserum). Also flow cytometry, immunoblots conditioned media confocal microscopy demonstrate not an artifact...

Genetic and pharmacological interventions in yeast mammalian cells have suggested a cross-talk between the actin cytoskeleton protein synthesis. Regulation of activity translation initiation factor 2 (eIF2) is paramount mechanism for to rapidly adjust rate synthesis trigger reprogramming gene expression response internal external cues. Here, we show that disruption F-actin inhibits GCN2-dependent manner, correlating with increased levels uncharged tRNA. GCN2 activation phosphorylation its...

Studies of cell transplantation therapeutics in animal models traumatic spinal cord injury (SCI) are often hampered by partial or complete rejection the graft host. Pharmacological immunosuppression is rarely sufficient to prevent rejection. Further, immunological niche created both host immune response and immunosuppressant drugs could hypothetically influence proliferation, differentiation, fate transplanted progenitor/stem cells. To avoid these confounds, we have previously used...

Background Diversity of T. cruzi strains is a central problem in Chagas disease research because its correlation with the wide range clinical manifestations and biogeographical parasite distribution. The role played by microdiversity epidemiology still debatable. Also awaits clarification whether such diversity associated outcome oral infection, responsible for frequent outbreaks acute disease. Methods Findings We addressed impact comparative analysis two strains, Y30 Y82, both derived from...

Abstract The Nucleosome Remodeling and Deacetylation (NuRD) complex is a crucial regulator of cellular differentiation. Two members the Methyl-CpG-binding domain (MBD) protein family, MBD2 MBD3, are known to be integral, but mutually exclusive subunits NuRD complex. Several MBD3 isoforms present in mammalian cells, resulting distinct MBD-NuRD complexes. Whether these different complexes serve functional activities during differentiation not fully explored. Based on essential role lineage...

The General Control Nonderepressible 2 (GCN2) kinase is a conserved member of the integrated stress response (ISR) pathway that represses protein translation and helps cells to adapt conditions nutrient shortage. As such, GCN2 required for longevity resistance induced by dietary restriction (DR). IMPACT an ancient inhibits GCN2. Here, we tested whether down-regulation mimics effects DR in C. elegans. Knockdown elegans homolog impt-1 activated ISR increased lifespan worms gcn-2-dependent...

The recent SARS-CoV-2 pandemic and associated COVID19 disease illustrates the important role of viral defence mechanisms in ensuring survival recovery host or patient. Viruses absolutely depend on host's protein synthesis machinery to replicate, meaning that impeding translation is a powerful way counteract viruses. One major approach used by cells obstruct phosphorylate alpha subunit eukaryotic initiation factor 2 (eIF2α). Mammals possess four different eIF2α-kinases: PKR, HRI, PEK/PERK,...

The Saccharomyces cerevisiae protein Yih1, when overexpressed, inhibits the eIF2 alpha kinase Gcn2 by competing for Gcn1 binding. However, deletion of YIH1 has no detectable effect on activity, suggesting that Yih1 is not a general inhibitor Gcn2, and phenotypic defect identified so far. Thus, its physiological role largely unknown. Here, we show involved in cell cycle. Yeast lacking displays morphological patterns DNA content indicative delay G2/M phases cycle, this phenotype independent...

Abstract The unicellular protozoa Giardia lamblia is a food‐ and waterborne parasite that causes giardiasis. This illness manifested as acute self‐limited diarrhea can evolve to long‐term complications. Successful establishment of infection by trophozoites requires adhesion host cells colonization the small intestine, where parasites multiply mitotic division. tight binding occurs means ventral adhesive disc, spiral array microtubules associated proteins such giardins. In this work we show...

Many studies require the detection and relative quantitation of proteins from cell culture samples using immunoblotting. Limiting factors are cost protease inhibitors, time required to break cells generate samples, as well high risk protein loss during breakage procedures. In addition, a common problem is viscosity lysed due released genomic DNA. As consequence, DNA needs be broken down prior denaturing polyacrylamide gel electrophoresis (SDS-PAGE), e.g. by passing sample through syringe...

Pham B, Piard‐Ruster K, Silva R, Gallo A, Esquivel CO, Martinez OM, Krams SM. Changes in natural killer cell subsets pediatric liver transplant recipients. Pediatr Transplantation 2012: 16: 176–182. © 2012 John Wiley & Sons A/S. Abstract: NK cells are important the immune response against tumors and virally infected cells. A balance between inhibitory activating receptors controls effector functions of We examined fate circulating expression cell‐activating Blood specimens were collected...

The common method for liberating proteins from Saccharomyces cerevisiae cells involves mechanical cell disruption using glass beads and buffer containing inhibitors (protease, phosphatase and/or kinase inhibitors), followed by centrifugation to remove debris. This procedure requires the use of costly is laborious, in particular when many samples need be processed. Also, enzymatic reactions can still occur during harvesting breakage. As a result low-abundance labile may degraded, enzymes such...

Pch2 is an AAA+ protein that controls DNA break formation, recombination and checkpoint signaling during meiotic G2/prophase. Chromosomal association of linked to these processes, several factors influence the euchromatin specialized chromatin ribosomal (r)DNA array budding yeast. Here, we describe a comprehensive mapping localization across yeast genome Within non-rDNA chromatin, associates with subset actively RNA Polymerase II (RNAPII)-dependent transcribed genes. Chromatin...

Pch2 is a meiosis-specific AAA+ protein that controls several important chromosomal processes. We previously demonstrated Orc1, subunit of the ORC, functionally interacts with budding yeast Pch2. The ORC (Orc1-6) complex loads MCM helicase to origins replication, but whether and how collaborates remains unclear. Here, we show hexamer directly associates during meiotic G2/prophase. Biochemical analysis suggests uses its non-enzymatic NH 2 -terminal domain core likely engages interface also...

Usage of a semi-dry electrophoretic apparatus is fast and cost effective way to transfer proteins from polyacrylamide gels membranes for immunoblotting.However, its e ciency usually limited small mid-range molecular weight proteins, the time-consuming wet procedure preferred method high proteins.Here we describe protocol cient diverse sizes \(ranging 15 296 kDa) resolved in gradient SDS-PolyAcrylamide Gel Electrophoresis \(PAGE) gel.This was successfully employed detect protein Rps22 \(15...

Abstract DNA damage is a constant threat to genome integrity and function. Diminished capacity for repair linked many human diseases, therefore understanding the molecular pathways responding key developing novel therapies. Lack of unbiased probes report dynamics associated proteins in living cells animals limit our current efforts completely understand processes. In this study we overcome these limitations by engineering protein containing tandem-BRCT domain MCPH1, which show have specific...

A cross-sectional study was done in students with univentricular heart after undergoing total cavopulmonary connection (Fontan procedure) to establish their baseline functional capacity based on the six-minute walk test and identify potential determining factors.Thirty patients were included; median age 12 years old.Eighteen males.The distance walked 551.3 meters, 84 % of theoretical for a healthy pediatric population.Height, pre-test systolic blood pressure, resting arterial oxygen...

Bacterial artificial chromosome (BAC)–based transgenes have emerged as a powerful tool for controlled and conditional interrogation of protein function in higher eukaryotes. Although homologous recombination-based recombineering methods streamlined the efficient integration tags onto BAC transgenes, generating precise point mutations has remained less time-consuming. Here, we present simplified method inserting into requiring single step followed by antibiotic selection. This technique,...

ABSTRACT ORC (Orc1-6) is an AAA+ complex that loads the MCM helicase to replication origins. Orc1, a subunit of ORC, functionally interacts with budding yeast Pch2, meiosis-specific protein. Pch2 regulates several chromosomal events gametogenesis, but mechanisms dictate function remain poorly understood. We demonstrate directly hexamer. The ORC-Pch2 assembly established without Cdc6, factor crucial for ORC-MCM binding. Biochemical analysis suggests utilizes ORC’s Cdc6-binding interface and...