Dataset Information

Chronic lymphocytic leukaemia cells exhibit functional heterogeneity in the unfolded protein response: a proteomic study

ABSTRACT: Genomic, epigenomic, and transcriptomic studies have provided only a partial explanation of disease heterogeneity in chronic lymphocytic leukaemia (CLL). Since mRNA and protein expression do not always correlate, CLL heterogeneity was investigated using global cellular proteomics. Sequential Windowed Acquisition of All THeoretical fragments Mass Spectrometry (SWATH-MS) was applied to CLL samples obtained from patients undergoing initial treatment with fludarabine-containing chemoimmunotherapy. Two independent proteomic datasets were generated: one compared pre-treatment samples from 32 patients who achieved an optimal versus suboptimal therapy response; the other compared paired samples from 16 patients collected before treatment and at disease progression. eIF2 signalling, which is pivotal to the unfolded protein response (UPR), was identified by Ingenuity Pathway Analysis as the most enriched pathway in both datasets. eIF2 signalling was also identified as the top pathway in a fludarabine-resistant CLL cell line established from HG3 cells. Measurement of key components of the eIF2 signalling pathway by Western blotting showed that fludarabine-resistant HG3 cells expressed higher levels of PERK, which phosphorylates the regulatory eIF2α subunit, and lower levels of BiP, an HSP70 molecular chaperone that inactivates PERK but preferentially binds to misfolded proteins during ER stress. The PERK inhibitor, GSK2606414, sensitised resistant, but not sensitive, HG-3 cells to fludarabine without affecting background cell viability or cytotoxicity induced by venetoclax. These findings provide the first evidence of functionally important heterogeneity in the UPR of CLL cells and, in doing so, illustrate the value of global cellular proteomics as an investigative tool in assessing differential patient response to therapy.

INSTRUMENT(S):

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): B Cell

DISEASE(S): Chronic Lymphocytic Leukemia

SUBMITTER: Rosalind Jenkins

LAB HEAD: Rosalind Jenkins

PROVIDER: PXD058183 | Pride | 2025-08-04

REPOSITORIES: Pride

ACCESS DATA

Dataset's files

Source:

			Action	DRS
	Baseline_progression_study_DIANN_matrix.xlsx	Xlsx
	MRD_study_DIANN_matrix.xlsx	Xlsx
	Resistance_model_DIANN_matrix.xlsx	Xlsx
	UK_CLL_10_2_7_SWATH_050819.wiff	Wiff
	UK_CLL_10_2_7_SWATH_050819.wiff.scan	Wiff

Items per page:

1 - 5 of 251

Publications

The unfolded protein response influences therapy outcome and disease progression in chronic lymphocytic leukaemia.

Khan Umair Tahir UT Clarke Kim K Eagle Gina G Oates Melanie M Hillmen Peter P Jayne Sandrine S Dyer Martin J S MJS Phipps Alex A Falciani Francesco F Jenkins Rosalind E RE Pettitt Andrew R AR

Scientific reports 20250728 1

Since genomics, epigenomics and transcriptomics have provided only a partial explanation of chronic lymphocytic leukaemia (CLL) heterogeneity, and since concordance between mRNA and protein expression is incomplete, we related the CLL proteome to clinical outcome. CLL samples from patients who received fludarabine-containing chemoimmunotherapy were analysed by mass spectrometry (SWATH-MS). One dataset compared pre-treatment samples associated with an optimal versus suboptimal response, while ano ...[more]

PMID: 40721481

Similar Datasets

Project description:Disruption of protein folding in the endoplasmic reticulum triggers the Unfolded Protein Response (UPR), a transcriptional and translational control network designed to restore protein homeostasis. Central to the UPR is PERK phosphorylation of the alpha subunit of eIF2 (eIF2~P), which represses global translation coincident with preferential translation of mRNAs, such as ATF4 and CHOP, that serve to implement the UPR transcriptional regulation. In this study, we used sucrose gradient ultracentrifugation and a genome-wide microarray approach to measure changes in mRNA translation during ER stress. Our analysis suggests that translational efficiencies vary across a broad range during ER stress, with the majority of transcripts being either repressed or resistant to eIF2~P, while a notable cohort of key regulators are subject to preferential translation. From this latter group, we identify IBTKa as being subject to both translation and transcriptional induction during eIF2~P in both cell lines and a mouse model of ER stress. Translational regulation of IBTKalpha mRNA involves the stress-induced relief of two inhibitory uORFs in the 5'-leader of the transcript. Depletion of IBTKalpha by shRNA reduced viability of cultured cells coincident with increased caspase 3/7 cleavage, suggesting that IBTKalpha is a key regulator in determining cell fate during the UPR. We used a genome-wide microarray approach to determine how individual mRNAs were differentially translated during endoplasmic reticulum stress. A microarray analysis from our laboratory identified gene transcripts suggested to be under translation control in mouse embryonic fibroblast (MEF) cells following a 6 hour treatment with thapsigargin, a potent inducer of ER stress, or no stress. The mRNAs were separated by sucrose gradient analyses to yield three fractions, those transcripts associated with large polysomes (?4 ribosomes per mRNA), those associated with monosome, disomes, or trisomes, and those fractionated at the top of the gradient with free ribosomes. RNA was extracted from sucrose gradients corresponding to these fractions and hybridized on Affymetrix microarrays. In parallel, we also measured total levels for each gene transcript in the presence or absence of thapsigargin treatment to address transcription regulation coincident with translational control. Please note that the treatment plus fractionation based on association with different numbers of ribosomes did yield different populations of mRNAs, which resulted in considerable variation in normalized data across the samples.

Project description:A data-entrained computational model for testing the regulatory logic of the vertebrate unfolded protein response This model is described in the article: A data-entrained computational model for testing the regulatory logic of the vertebrate unfolded protein response. Diedrichs DR, Gomez JA, Huang CS, Rutkowski DT, Curtu R. Mol. Biol. Cell 2018 Apr; : mbcE17090565 Abstract: The vertebrate unfolded protein response (UPR) is characterized by multiple interacting nodes among its three pathways, yet the logic underlying this regulatory complexity is unclear. To begin to address this issue, we created a computational model of the vertebrate UPR that was entrained upon and then validated against experimental data. As part of this validation, the model successfully predicted the phenotypes of cells with lesions in UPR signaling, including a surprising and previously unreported differential role for the eIF2? phosphatase GADD34 in exacerbating severe stress but ameliorating mild stress. We then used the model to test the functional importance of a feed-forward circuit within the PERK/CHOP axis, and of cross-regulatory control of BiP and CHOP expression. We found that the wiring structure of the UPR appears to balance the ability of the response to remain sensitive to ER stress yet also to be rapidly deactivated by improved protein folding conditions. This model should serve as a valuable resource for further exploring the regulatory logic of the UPR. This model is hosted on BioModels Database and identified by: MODEL1803300000. To cite BioModels Database, please use: Chelliah V et al. BioModels: ten-year anniversary. Nucl. Acids Res. 2015, 43(Database issue):D542-8. To the extent possible under law, all copyright and related or neighbouring rights to this encoded model have been dedicated to the public domain worldwide. Please refer to CC0 Public Domain Dedication for more information.