42 research outputs found
Acetylation-induced TDP-43 pathology is suppressed by an HSF1-dependent chaperone program
TDP-43 pathology marks a spectrum of multisystem proteinopathies including amyotrophic lateral sclerosis, frontotemporal lobar degeneration, and sporadic inclusion body myositis. Surprisingly, it has been challenging to recapitulate this pathology, highlighting an incomplete understanding of TDP-43 regulatory mechanisms. Here we provide evidence supporting TDP-43 acetylation as a trigger for disease pathology. Using cultured cells and mouse skeletal muscle, we show that TDP-43 acetylation-mimics promote TDP-43 phosphorylation and ubiquitination, perturb mitochondria, and initiate degenerative inflammatory responses that resemble sporadic inclusion body myositis pathology. Analysis of functionally linked amyotrophic lateral sclerosis proteins revealed recruitment of p62, ubiquilin-2, and optineurin to TDP-43 aggregates. We demonstrate that TDP-43 acetylation-mimic pathology is potently suppressed by an HSF1-dependent mechanism that disaggregates TDP-43. Our study illustrates bidirectional TDP-43 processing in which TDP-43 aggregation is targeted by a coordinated chaperone response. Thus, activation or restoration of refolding mechanisms may alleviate TDP-43 aggregation in tissues that are uniquely susceptible to TDP-43 proteinopathies.TDP-43 aggregation is linked to various diseases including amyotrophic lateral sclerosis. Here the authors show that acetylation of the protein triggers TDP-43 pathology in cultured cells and mouse skeletal muscle, which can be cleared through an HSF1-dependent chaperone mechanism that disaggregates the protein
Desorption electrospray ionization mass spectrometry for monitoring the kinetics of Baeyer-Villiger solid-state organic reactions
Remote Mass Spectrometric Sampling of Electrospray- and Desorption Electrospray-Generated Ions Using an Air Ejector
A commercial air ejector was coupled to an electrospray ionization linear ion trap mass spectrometer (LTQ) to transport remotely generated ions from both electrospray (ESI) and desorption electrospray ionization (DESI) sources. We demonstrate the remote analysis of a series of analyte ions that range from small molecules and polymers to polypeptides using the AE-LTQ interface. The details of the ESI-AE-LTQ and DESI-AE-LTQ experimental configurations are described and preliminary mass spectrometric data are presented
An Automated Pipeline to Monitor System Performance in Liquid Chromatography–Tandem Mass Spectrometry Proteomic Experiments
Learning Score Function Parameters for Improved Spectrum Identification in Tandem Mass Spectrometry Experiments
The identification of proteins from spectra derived from
a tandem
mass spectrometry experiment involves several challenges: matching
each observed spectrum to a peptide sequence, ranking the resulting
collection of peptide-spectrum matches, assigning statistical confidence
estimates to the matches, and identifying the proteins. The present
work addresses algorithms to rank peptide–spectrum matches.
Many of these algorithms, such as PeptideProphet, IDPicker, or Q-ranker,
follow a similar methodology that includes representing peptide-spectrum
matches as feature vectors and using optimization techniques to rank
them. We propose a richer and more flexible feature set representation
that is based on the parametrization of the SEQUEST XCorr score and
that can be used by all of these algorithms. This extended feature
set allows a more effective ranking of the peptide-spectrum matches
based on the target-decoy strategy, in comparison to a baseline feature
set devoid of these XCorr-based features. Ranking using the extended
feature set gives 10–40% improvement in the number of distinct
peptide identifications relative to a range of <i>q</i>-value
thresholds. While this work is inspired by the model of the theoretical
spectrum and the similarity measure between spectra used specifically
by SEQUEST, the method itself can be applied to the output of any
database search. Further, our approach can be trivially extended beyond
XCorr to any linear operator that can serve as similarity score between
experimental spectra and peptide sequences
Reagent for Evaluating Liquid Chromatography–Tandem Mass Spectrometry (LC-MS/MS) Performance in Bottom-Up Proteomic Experiments
We
present a novel proteomic standard for assessing liquid chromatography–tandem
mass spectrometry (LC-MS/MS) instrument performance, in terms of chromatographic
reproducibility and dynamic range within a single LC-MS/MS injection.
The peptide mixture standard consists of six peptides that were specifically
synthesized to cover a wide range of hydrophobicities (grand average
hydropathy (GRAVY) scores of −0.6 to 1.9). A combination of
stable isotope labeled amino acids (<sup>13</sup>C and <sup>15</sup>N) were inserted to create five isotopologues. By combining these
isotopologues at different ratios, they span four orders of magnitude
within each distinct peptide sequence. Each peptide, from lightest
to heaviest, increases in abundance by a factor of 10. We evaluate
several metrics on our quadrupole orbitrap instrument using the 6
× 5 LC-MS/MS reference mixture spiked into a complex lysate background
as a function of dynamic range, including mass measurement accuracy
(MMA) and the linear range of quantitation of MS1 and parallel reaction
monitoring experiments. Detection and linearity of the instrument
routinely spanned three orders of magnitude across the gradient (500
fmol to 0.5 fmol on column) and no systematic trend was observed for
MMA of targeted peptides as a function of abundance by analysis of
variance analysis (<i>p</i> = 0.17). Detection and linearity
of the fifth isotopologue (i.e., 0.05 fmol on column) was dependent
on the peptide and instrument scan type (MS1 vs PRM). We foresee that
this standard will serve as a powerful method to conduct both intra-instrument
performance monitoring/evaluation, technology development, and inter-instrument
comparisons
A method to determine the kinetics of multiple proteins in human infants with respiratory distress syndrome
We report a method to measure in vivo turnover of four proteins from sequential tracheal aspirates obtained from human newborn infants with respiratory distress syndrome using targeted proteomics. We detected enrichment for all targeted proteins approximately 3 h from the start of infusion of [5,5,5-2H3] leucine, secretion times that varied from 1.2 to 2.5 h, and half lives that ranged between 10 and 21 h. Complement factor B, a component of the alternative pathway of complement activation, had an approximately twofold-longer half-life than the other three proteins. In addition, the kinetics of mature and carboxy-terminal tryptic peptides from the same protein (surfactant protein B) were not statistically different (p\u2009=\u20090.49)