3,100 research outputs found
Energy- and temperature-dependent transport of integral proteins to the inner nuclear membrane via the nuclear pore
Resident integral proteins of the inner nuclear membrane (INM) are synthesized as membrane-integrated proteins on the peripheral endoplasmic reticulum (ER) and are transported to the INM throughout interphase using an unknown trafficking mechanism. To study this transport, we developed a live cell assay that measures the movement of transmembrane reporters from the ER to the INM by rapamycin-mediated trapping at the nuclear lamina. Reporter constructs with small (<30 kD) cytosolic and lumenal domains rapidly accumulated at the INM. However, increasing the size of either domain by 47 kD strongly inhibited movement. Reduced temperature and ATP depletion also inhibited movement, which is characteristic of membrane fusion mechanisms, but pharmacological inhibition of vesicular trafficking had no effect. Because reporter accumulation at the INM was inhibited by antibodies to the nuclear pore membrane protein gp210, our results support a model wherein transport of integral proteins to the INM involves lateral diffusion in the lipid bilayer around the nuclear pore membrane, coupled with active restructuring of the nuclear pore complex
Dominant gain-of-function mutations in Hsp104p reveal crucial roles for the middle region
Heat-shock protein 104 (Hsp104p) is a protein-remodeling factor that promotes survival after extreme stress by disassembling aggregated proteins and can either promote or prevent the propagation of prions (protein-based genetic elements). Hsp104p can be greatly overexpressed without slowing growth, suggesting tight control of its powerful protein-remodeling activities. We isolated point mutations in Hsp104p that interfere with this control and block cell growth. Each mutant contained alterations in the middle region (MR). Each of the three MR point mutations analyzed in detail had distinct phenotypes. In combination with nucleotide binding site mutations, Hsp104p(T499I) altered bud morphology and caused septin mislocalization, colocalizing with the misplaced septins. Point mutations in the septin Cdc12p suppressed this phenotype, suggesting that it is due to direct Hsp104p–septin interactions. Hsp104p(A503V) did not perturb morphology but stopped cell growth. Remarkably, when expressed transiently, the mutant protein promoted survival after extreme stress as effectively as did wild-type Hsp104p. Hsp104p(A509D) had no deleterious effects on growth or morphology but had a greatly reduced ability to promote thermotolerance. That mutations in an 11-amino acid stretch of the MR have such profound and diverse effects suggests the MR plays a central role in regulating Hsp104p function
Degrees of controllability for quantum systems and applications to atomic systems
Precise definitions for different degrees of controllability for quantum
systems are given, and necessary and sufficient conditions are discussed. The
results are applied to determine the degree of controllability for various
atomic systems with degenerate energy levels and transition frequencies.Comment: 20 pages, IoP LaTeX, revised and expanded versio
Quantum System Identification by Bayesian Analysis of Noisy Data: Beyond Hamiltonian Tomography
We consider how to characterize the dynamics of a quantum system from a
restricted set of initial states and measurements using Bayesian analysis.
Previous work has shown that Hamiltonian systems can be well estimated from
analysis of noisy data. Here we show how to generalize this approach to systems
with moderate dephasing in the eigenbasis of the Hamiltonian. We illustrate the
process for a range of three-level quantum systems. The results suggest that
the Bayesian estimation of the frequencies and dephasing rates is generally
highly accurate and the main source of errors are errors in the reconstructed
Hamiltonian basis.Comment: 6 pages, 3 figure
Clinical and functional characterisation of a novel TNFRSF1A c.605T > A/V173D cleavage site mutation associated with tumour necrosis factor receptor-associated periodic fever syndrome (TRAPS), cardiovascular complications and excellent response to etanercept treatment.
Objectives: To study the clinical outcome, treatment
response, T-cell subsets and functional consequences of a
novel tumour necrosis factor (TNF) receptor type 1
(TNFRSF1A) mutation affecting the receptor
cleavage site.
Methods: Patients with symptoms suggestive of tumour
necrosis factor receptor-associated periodic syndrome
(TRAPS) and 22 healthy controls (HC) were screened for
mutations in the TNFRSF1A gene. Soluble TNFRSF1A and
inflammatory cytokines were measured by ELISAs.
TNFRSF1A shedding was examined by stimulation of
peripheral blood mononuclear cells (PBMCs) with phorbol
12-myristate 13-acetate followed by flow cytometric
analysis (FACS). Apoptosis of PBMCs was studied by
stimulation with TNFa in the presence of cycloheximide
and annexin V staining. T cell phenotypes were monitored
by FACS.
Results: TNFRSF1A sequencing disclosed a novel V173D/
p.Val202Asp substitution encoded by exon 6 in one
family, the c.194–14G.A splice variant in another and
the R92Q/p.Arg121Gln substitution in two families.
Cardiovascular complications (lethal heart attack and
peripheral arterial thrombosis) developed in two V173D
patients. Subsequent etanercept treatment of the V173D
carriers was highly effective over an 18-month follow-up
period. Serum TNFRSF1A levels did not differ between
TRAPS patients and HC, while TNFRSF1A cleavage from
monocytes was significantly reduced in V173D and R92Q
patients. TNFa-induced apoptosis of PBMCs and T-cell
senescence were comparable between V173D patients
and HC.
Conclusions: The TNFRSF1A V173D cleavage site
mutation may be associated with an increased risk for
cardiovascular complications and shows a strong
response to etanercept. T-cell senescence does not seem
to have a pathogenetic role in affected patients
Organellar proteomics: the prizes and pitfalls of opening the nuclear envelope
Proteomic studies have the potential to comprehensively define the composition of organelles but are limited by the organellar cross-contamination that arises during subcellular fractionation. Comparative proteomics of organellar subfractions can mitigate these problems, as demonstrated by a recent study involving the nuclear envelope
Enhancers on the edge — how the nuclear envelope controls gene regulatory elements
Precise temporal and sequential control of gene expression during development and in response to environmental stimuli requires tight regulation of the physical contact between gene regulatory elements and promoters. Current models describing how the genome folds in 3D space to establish these interactions often ignore the role of the most stable structural nuclear feature — the nuclear envelope. While contributions of 3D folding within/between topologically associated domains (TADs) have been extensively described, mechanical contributions from the nuclear envelope can impact enhancer–promoter interactions both directly and indirectly through influencing intra/inter-TAD interactions. Importantly, these nuclear envelope contributions clearly link this mechanism to development and, when defective, to human disease. Here, we discuss evidence for nuclear envelope regulation of tissue-specific enhancer–promoter pairings, potential mechanisms for this regulation, exciting recent findings that other regulatory elements such as microRNAs and long noncoding RNAs are under nuclear envelope regulation, the possible involvement of condensates, and how disruption of this regulation can lead to disease
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