Quantitative chemical proteomics of Leishmania mexicana
Ontology highlight
ABSTRACT:
INSTRUMENT(S): TripleTOF 5600
ORGANISM(S): Leishmania Mexicana Mexicana
SUBMITTER: 
Kalesh Karunakaran
LAB HEAD: Kalesh Karunakaran
PROVIDER: PXD022708 | Pride | 2021-04-23
REPOSITORIES: Pride
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| 17mADAG_e1.wiff | Wiff | |||
| 17mADAG_e2.wiff | Wiff | |||
| 17mADAG_e3.wiff | Wiff | |||
| Tanespi_1h_e1.wiff | Wiff | |||
| Tanespi_1h_e2.wiff | Wiff |
Items per page: 5 1 - 5 of 17 |
Similar Datasets
Project description:Leishmaniasis are a group of diseases caused by parasitic protozoa of the genus Leishmania. Current treatments are limited by difficult administration, high cost, poor efficacy, toxicity, and growing resistance. New agents, with new mechanisms of action, are urgently needed to treat the disease. Although extensively studied in other organisms, serine proteases (SPs) have not been widely explored as antileishmanial drug targets. Herein we report for the first time an activity-based protein profiling (ABPP) strategy to investigate new therapeutic targets within the SPs of the Leishmania parasites. Active site directed fluorophosphonate probes (rhodamine and biotin‐conjugated) were used for the detection and identification of active Leishmania serine hydrolases (SHs). Significant differences were observed in the SHs expression levels throughout the Leishmania life cycle and between different Leishmania species. Using iTRAQ-labelling-based quantitative proteomic mass spectrometry, we identified two targetable SPs in Leishmania mexicana: carboxypeptidase LmxM.18.0450 and prolyl oligopeptidase LmxM.36.6750. Druggability was ascertained by selective inhibition using the commercial serine protease inhibitors chymostatin, lactocystin and ZPP, which represent templates for future anti-leishmanial drug discovery programs. Collectively, the use of ABPP method complements existing genetic methods for target identification and validation in Leishmania.
2022-07-04 | PXD033616 | Pride
Project description:Using a combination of mass spectrometry-based quantitative proteomics, chemical and metabolic labelling, we provide the first protein-level evidence that the Hsp90 inhibition affects the expression of hundreds of proteins in Leishmania spp. Our study defines the Leishmania parasite’s response to Hsp90 inhibition at its global proteome and provides a rich resource for future studies on Leishmania biology and antileishmanial drug development.
2021-04-23 | PXD024764 | Pride
Project description:Using a combination of BONCAT technique and iTRAQ 3-plex quantitative proteomics MS, we profiled the de novo protein synthesis in the starving L. mexicana parasites in a temporally resolved manner. This approach uniquely integrates the high specificity of the BONCAT technique for the NSPs with the high sensitivity and multiplexed quantitation capability of the iTRAQ quantitative proteomics MS. Proof-of-concept experiments identified over 250 starvation-specific NSPs in the parasite and quantified the relative changes in abundance of these proteins as a function of duration of starvation.
2019-12-04 | PXD015827 | Pride
Project description:Using a combination of phosphoproteome enrichment and tandem mass tag (TMT) labelling-based quantitative proteomic mass spectrometry (MS), we robustly identified and quantified 1,833 phosphorylated proteins across three life cycle stages of Leishmania mexicana (L. mexicana) parasite. Protein kinase domain was the most enriched protein domain in the L. mexicana phosphoproteome. Additionally, this study systematically characterised the perturbing effect of HSP90 inhibition on the global phosphoproteome of the L. mexicana across its life cycle stages and showed that the inhibition causes substantially distinct molecular effects in the promastigotes and the amastigotes. While tanespimycin treatment decreased the phosphorylation of HSP90 and its co-chaperon HSP70 in the amastigotes, the opposite effect was observed in the promastigotes. Additionally, our results show that while kinase activity and microtubule motor activity are highly represented in the negatively affected phosphoproteins of the promastigotes, ribosomal proteins, protein folding, and proton channel activity are preferentially enriched in the perturbed amastigote phosphoproteome. Our results also show that RNA helicase domain was distinctively enriched among the positively affected RNA-binding amastigote phosphoproteome. This study reveals the dramatically different ways the HSP90 inhibition stress modulates the phosphoproteome of the pathogenic amastigotes and provides in-depth insight into the scope of selective molecular targeting in the therapeutically relevant amastigotes.
2023-10-04 | PXD043002 | Pride
Project description:The capacity to sense and transduce temperature signals pervades all aspects of biology, and temperature exerts powerful control over the development and virulence of diverse pathogens. In the leading fungal pathogen of humans, Candida albicans, temperature has a profound impact on morphogenesis, a key virulence trait. Many cues that induce the transition from yeast to filamentous growth are contingent on a minimum temperature of 37ºC, while further elevatation to 39ºC serves as an independent inducing cue. The molecular chaperone Hsp90 is a key regulator of C. albicans temperature-dependent morphogenesis, as induction of filamentous growth requires relief from Hsp90-mediated repression of the morphogenetic program. Compromise of Hsp90 function genetically, pharmacologically, or by elevated temperature induces filamentation in a manner that depends on protein kinase A (PKA) signaling, but is independent of the terminal transcription factor, Efg1. Here, we determine that despite morphological and regulatory differences, inhibition of Hsp90 induces a transcriptional profile similar to that induced by other filamentation cues, and does so in a manner that is independent of Efg1. Further, we identify Hms1 as a transcriptional regulator required for morphogenesis induced by elevated temperature or compromise of Hsp90 function. Hms1 functions downstream of the cyclin Pcl, and the cyclin-dependent kinase Pho85, both of which are required for temperature-dependent filamentation. Upon Hsp90 inhibition, Hms1 binds to DNA elements involved in filamentous growth, including UME6 and RBT5, and regulates their expression, providing a mechanism through which Pho85, Pcl1, and Hms1 govern morphogenesis. Consistent with the importance of morphogenetic flexibility with virulence, deletion of C. albicans HMS1 attenuates virulence in a metazoan model of infection. Thus, we establish a new mechanism through which Hsp90 orchestrates C. albicans morphogenesis, and define novel regulatory circuitry governing a temperature-dependent developmental program, with broad implications for temperature sensing and virulence of microbial pathogens. Two-color experimental design testing the effect of geldanamycin on wild type of delta-efg1 cells. RNA from each replicate came from independent cultures.
2012-03-20 | E-GEOD-34255 | biostudies-arrayexpress
Project description:Purpose: The growth and survival of NF2-deficient cells are enhanced by the activation of multiple signaling pathways including ErbB2/IGF-1R/Met, PI3K/Akt, and Ras/Raf/Mek/Erk1/2. The ubiquitously expressed chaperone protein HSP90 is known to be essential for the stabilization of these signaling molecules. We aim to evaluate the effect of the HSP90 inhibition on the signaling pathways activated in NF2-related tumors. We tested the efficacy of the newly synthesized small molecule NXD30001 which was shown to be more potent in HSP90 inhibition in vitro and less toxic in vivo than already existing HSP90 inhibitors. Experimental Design: The anti-proliferative activity of NXD30001 was tested in NF2-deficient mouse cells in vitro, and its anti-tumor efficacy was verified in an NF2-deficient allograft model in vivo. The underlying molecular alteration in vitro and in vivo was further characterized by a global transcriptome approach. Results: NXD30001 was found to induce degradation of client proteins and to suppress proliferation in NF2-deficient cells in vitro and in vivo. Differential expression analysis further identified subsets of genes implicated in cell proliferation, survival, vascularization, and Schwann cell differentiation, whose expression were altered by NXD30001 treatment. Conclusions: NXD30001 is a potent HSP90 inhibitor showing significant antitumor activity against NF2-related tumor cells in vitro and in vivo, and represents a promising option for novel NF2 therapies. Global RNA expression in NXD30001-treated and untreated matched NF2-deficiemt cultured cells (in vitro) and allograft tumors (in vivo) were compared by RNA-Seq.
2013-06-06 | E-GEOD-40187 | biostudies-arrayexpress
Project description:The BCL6 transcriptional repressor is a critical oncogene in diffuse large B-cell lymphomas (DLBCL). The specific BCL6 inhibitor RI-BPI potently kills DLBCL cells. We find that RI-BPI induces a particular gene expression signature in DLBCL. In order to identify classes of drugs that might synergize with RIBPI we examined the connectivity of this signature and found a strong association with HDAC and Hsp90 inhibitors. This was explained by the discovery that BCL6 directly represses the p300 lysine acetyltransferase and its co-factor BAT3. RI-BPI induced expression of p300 and BAT3, and p300 acetyltransferase activity, resulting in acetylation of p300 targets like p53 and Hsp90. As a consequence, RI-BPI could attenuate Hsp90 chaperone function, similar to the effect of Hsp90 and HDAC inhibitors. Induction of p300 and BAT3 was required for the anti-lymphoma effects of RI-BPI since specific blockade of either protein rescued DLBCL cells from the BCL6 inhibitor. RI-BPI synergistically killed DLBCL cells in combination with HDAC inhibitors (SAHA, TSA and VPA) and Hsp90 inhibitors (17-DMAG and PUH71). The combination of RI-BPI and SAHA, or RI-BPI and PU-H71 potently suppressed or even eradicated human DLBCL in mice. BCL6 repression of EP300 thus provides a basis for rational targeted combinatorial therapy for patients with DLBCL. Direct comparison of gene expression levels in DLBCL cell lines after 24hs of treatment with either a Bcl6 inhibitor peptide or control peptide
2010-09-21 | E-GEOD-24236 | biostudies-arrayexpress
Project description:HuntingtonM-bM-^@M-^Ys disease (HD) is a neurodegenerative disorder that is associated with the deposition of proteinaceous aggregates in the brains of HD patients and mouse models. Previous studies have suggested that wide-scale disruption of protein homeostasis occurs in protein folding diseases. Protein homeostasis can be maintained by activation of the heat shock response (HSR) via the transcription factor heat shock factor 1 (HSF1), the pharmacological activation of which can be achieved by Hsp90 inhibition and has been demonstrated to be beneficial in cell and invertebrate models of HD. Whether the HSR is functional in HD and whether its activation has therapeutic potential in mammalian HD models is currently unknown. To address these issues, we used a novel, brain penetrant Hsp90 inhibitor to activate the HSR in brain after systemic administration. Microarrays, quantitative PCR and western blotting showed that the HSR becomes impaired with disease progression in two mouse models of HD and that this originates at the level of transcription. HuntingtonM-bM-^@M-^Ys disease (HD) is a neurodegenerative disorder that is associated with the deposition of proteinaceous aggregates in the brains of HD patients and mouse models. Previous studies have suggested that wide-scale disruption of protein homeostasis occurs in protein folding diseases. Protein homeostasis can be maintained by activation of the heat shock response (HSR) via the transcription factor heat shock factor 1 (HSF1), the pharmacological activation of which can be achieved by Hsp90 inhibition and has been demonstrated to be beneficial in cell and invertebrate models of HD. Whether the HSR is functional in HD and whether its activation has therapeutic potential in mammalian HD models is currently unknown. To address these issues, we used a novel, brain penetrant Hsp90 inhibitor to activate the HSR in brain after systemic administration. Microarrays, quantitative PCR and western blotting showed that the HSR becomes impaired with disease progression in two mouse models of HD and that this originates at the level of transcription. mRNA expression analysis was performed by microarray in 12 week old R6/2 and WT mice treated with vehicle or HSP990 for 4h (12mg/kg).The following number of replicates was analysed for each genotype: 7 for WT+vehicle, 10 for WT+HSP990, 7 for R6/2+vehicle and 8 for R6/2+HSP990. Microarray quality control was performed using the software package provided on RACE (http://race.unil.ch).
2014-06-01 | E-GEOD-29681 | biostudies-arrayexpress
Project description:The recent development within high-throughput technologies for expression profiling has allowed for parallel analysis of transcriptomes and proteomes in biological systems such as comparative analysis of transcript and protein levels of tissue regulated genes. Until now, such studies of have only included microarray or short length sequence tags for transcript profiling. Furthermore, most comparisons of transcript and protein levels have been based on absolute expression values from within the same tissue and not relative expression values based on tissue ratios. Presented here is a novel study of two porcine tissues based on integrative analysis of data from expression profiling of identical samples using cDNA microarray, 454-sequencing and iTRAQ-based proteomics. Sequence homology identified 2.541 unique transcripts that are detectable by both microarray hybridizations and 454-sequencing of 1.2 million cDNA tags. Both transcript-based technologies showed high reproducibility between sample replicates of the same tissue, but the correlation across these two technologies was modest. Thousands of genes being differentially expressed were identified with microarray. Out of the 306 differentially expressed genes, identified by 454-sequencing, 198 (65 %) were also found by microarray. The relationship between the regulation of transcript and protein levels was analyzed by integrating iTRAQ-based proteomics data. Protein expression ratios were determined for 354 genes, of which 148 could be mapped to both microarray and 454-sequencing data. A comparison of the expression ratios from the three technologies revealed that differences in transcript and protein levels across heart and muscle tissues are positively correlated. We show that the reproducibility within cDNA microarray and 454-sequencing is high, but that the agreement across these two technologies is modest. We demonstrate that the regulation of transcript and protein levels across identical tissue samples is positively correlated when the tissue expression ratios are used for comparison. The results presented are of interest in systems biology research in terms of integration and analysis of high-throughput expression data from mammalian tissues. Keywords: tissue comparison, platform comparison Tissue samples of heart (HEA) and Longissimus dorsi (LDO) were prepared by pooling from five healthy Hampshire gilts at age four to six months and division into six sub-samples, three for each tissue named HEA1, HEA2, HEA3, LDO1, LDO2 and LDO3. The exact same six tissue samples were used for subsequent expression profiling with cDNA microarray, 454-sequencing and iTRAQ-based proteomics. A reference sample for the cDNA microarray experiment and iTRAQ-based proteomics was constructed by combining the six samples. In the microarray experiment, three cDNA microarray slides were used per sample.
2009-01-22 | E-GEOD-10122 | biostudies-arrayexpress
Project description:Non-poly(A) RNA molecules including noncoding RNAs (ncRNAs) comprise the major portion of the total transcribed molecules in the cell. In addition to the mRNAs the ncRNAs also function as ribonucleoprotein particles (RNPs) and carry out biological functions including synthesis of new proteins, RNA processing, genome remodelling and regulation of transcription. We therefore envisaged a comprehensive transcriptome-wide identification of coding and non-coding RNA-binding proteins (RBPs) in the Leishmania spp. Towards this we applied the recently reported orthogonal organic phase separation (OOPS) method in combination with tandem mass tag (TMT) labelling-based quantitative proteomic mass spectrometry (MS) and report herein the most comprehensive identification of RBPs in Leishmania mexicana (L. mexicana) parasites. This study identified novel RNA binding property of thousands of L. mexicana proteins, significantly expanding the RBP landscape of the parasite. Furthermore, we showed that the classical Hsp90 inhibitor tanespimycin differentially regulates the RNA-binding property of hundreds of L. mexicana RBPs, shedding light into hitherto unknown large-scale downstream molecular effects of the small molecule inhibitor in the parasite.
2022-01-14 | PXD028092 | Pride