Defining the proteolytic landscape during enterovirus infection.

Viruses cleave cellular proteins to remodel the host proteome. The study of these cleavages has revealed mechanisms of immune evasion, resource exploitation, and pathogenesis. However, the full extent of virus-induced proteolysis in infected cells is unknown, mainly because until recently the technology for a global view of proteolysis within cells was lacking. Here, we report the first comprehensive catalog of proteins cleaved upon enterovirus infection and identify the sites within proteins where the cleavages occur. We employed multiple strategies to confirm protein cleavages and assigned them to one of the two enteroviral proteases. Detailed characterization of one substrate, LSM14A, a p body protein with a role in antiviral immunity, showed that cleavage of this protein disrupts its antiviral function. This study yields a new depth of information about the host interface with a group of viruses that are both important biological tools and significant agents of disease

Quantitative proteome analysis of an Escherichia coli exposed to tetracycline reveals multiple affected cytoplasmic metabolic processes

Background: Many studies have already focused on the evaluation of variations in protein expression caused by antibiotic exposure in tetracycline resistant microorganisms. However, little is known about the metabolic response of genetically resistant bacterial populations to antibiotic exposure. In this study we used a liquid chromatography-mass spectrometry-based (LC-MS/MS) proteomics to evaluate the global cytoplasmic metabolic changes of a resistant Escherichia coli isolate, when challenged with tetracycline. Material/methods: IsolateEcAamb278 was recovered from a soil sample, used for the intensive farming of tomato plants and showed nonsusceptibility to all categories of β-lactams except carbapanems and cefoxitin, and tetracycline. Previous genomic characterization revealed the presence of TEM-1, CTX-M-1, Sul2,TetA and TetB resistance determinants. Single colonies of E. coliEcAamb278 were used to extract crude cytoplasmic protein fraction using non detergent cell lysis buffer. The proteomic profile of E. coli EcAmb278 was determined by label-free LC-MS/MS upon tetracycline stress in a comparative study, using as referencethe proteome of the same strain nonexposed to antibiotics. The samples were analyzed by nano LC-MS/MS, using a Q-Exactive mass spectrometer. All data was searched with VEMS. Proteins were quantified by spectral counting and mziXIC, followed by iBAQ estimation. Results: The complete proteome yielded about 1484 proteins, using a 1% FDR as cut off. The comparison of the proteome profiles of the two E. coli EcAamb278 samples pointed to several proteins with altered expression under tetracycline stress conditions. The twelve most significant (FDRcorrected p<0.05) proteins differentially regulated by more than two-fold were involved in DNA metabolism, transcription, virulence, intracellular trafficking and secretion, biosynthesis of vitamins, other processes, and unknown functions. For instance, (1) acyl-CoA thioester hydrolase, involved in biosynthesis of coenzyme A; (2) bacterial NAD+-dependent DNA ligase, plays a critical role in DNA replication, recombination and repair in all living organisms; and (3) adhesion protein FimH, which confirmed the pathogenicity of this environmental-borne E. coli. Overall, several other detected proteins were associated with the process of antibiotic resistance, virulence and acquisition and transfer of foreign DNA (AcrA, TolC, MdtE, Omps, TnsE, Colicin peptides, among others). Conclusions: These results indicate that E. coli responses to tetracycline are related to protein translation as well as metabolic regulation. We observed differentially regulation of several metabolic proteins though not belonging to the canonical antibiotic resistance pathway, suggesting an integrated metabolic bacteria response to antibiotic exposure, which can potentially be explored for drug development.info:eu-repo/semantics/publishedVersio

Convergent Akt activation drives acquired EGFR inhibitor resistance in lung cancer

EGFR-mutant non-small cell lung cancer are often resistant to EGFR tyrosine kinase inhibitor treatment. In this study, the authors show that resistant tumors display high Akt activation and that a combined treatment with AKT inhibitors causes synergistic tumour growth inhibition in vitro and in vivo

Generation and characterization of a novel knockin minipig model of Hutchinson-Gilford progeria syndrome

Hutchinson-Gilford progeria syndrome (HGPS) is an extremely rare genetic disorder for which no cure exists. The disease is characterized by premature aging and inevitable death in adolescence due to cardiovascular complications. Most HGPS patients carry a heterozygous de novo LMNA c.1824C > T mutation, which provokes the expression of a dominant-negative mutant protein called progerin. Therapies proven effective in HGPS-like mouse models have yielded only modest benefit in HGPS clinical trials. To overcome the gap between HGPS mouse models and patients, we have generated by CRISPR-Cas9 gene editing the first large animal model for HGPS, a knockin heterozygous LMNA c.1824C > T Yucatan minipig. Like HGPS patients, HGPS minipigs endogenously co-express progerin and normal lamin A/C, and exhibit severe growth retardation, lipodystrophy, skin and bone alterations, cardiovascular disease, and die around puberty. Remarkably, the HGPS minipigs recapitulate critical cardiovascular alterations seen in patients, such as left ventricular diastolic dysfunction, altered cardiac electrical activity, and loss of vascular smooth muscle cells. Our analysis also revealed reduced myocardial perfusion due to microvascular damage and myocardial interstitial fibrosis, previously undescribed readouts potentially useful for monitoring disease progression in patients. The HGPS minipigs provide an appropriate preclinical model in which to test human-size interventional devices and optimize candidate therapies before advancing to clinical trials, thus accelerating the development of effective applications for HGPS patients.This project was mainly supported by an Established Investigator Award from the Progeria Research Foundation (2014-52), and from the Spanish Ministerio de Ciencia, Innovación y Universidades (MCIU), and the European Regional Development Fund (FEDER, “A way to build Europe”) (SAF2016-79490-R, CB16/11/00405). Ana Barettino has a predoctoral contract from MCIU (BES-2017-079705). Work at Universidad de Murcia is supported by Fundación Seneca-Agencia de Ciencia y Tecnología de la Región de Murcia (20040/GERM/16). The CNIC is supported by the MCIU and the Pro-CNIC Foundation and is a Severo Ochoa Center of Excellence (SEV-2015-0505).S

Actin remodeling by ADF/cofilin is required for cargo sorting at the trans-Golgi network

Sorting of both soluble and integral membrane proteins is disrupted by loss of ADF/cofilin, suggesting that actin severing controls expansion of a sorting domain within the TGN