Proteomics as Applied to Inherited Metabolic Diseases

Richard, E Davis; Gámez, Alejandra; Ruíz-Sala, Pedro; Pérez, Belén; Desviat, Lourdes R.; Ugarte, Magdalena

doi:10.2174/157016409789351897

Cited by 4 publications

(8 citation statements)

References 92 publications

(100 reference statements)

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“…Furthermore, certain genes such as SOD1, activated downstream of the OSR-associated transcription factor NRF2, 52 separate from the OSR cluster (purple in Figure 1C,D), reflecting the ability of this gene to be regulated by multiple stress-responsive signaling pathways apart from the OSR. 53 The PERK/ISR target MTHFR is also regulated by other UPR signaling pathways, as well as the OSR, 54,55 and is similarly found to separate from the larger cluster of PERK/ISR targets (green in Figure 1C,D). The S3 for the full ANOVA table.…”

Section: ■ Results and Discussionmentioning

confidence: 88%

“…1C-D), reflecting the ability for this gene to be regulated by multiple stress-responsive signaling pathways apart from the OSR 53 . The PERK/ISR target MTHFR is also regulated by other UPR signaling pathways, as well as the OSR [54][55] , and is similarly found to separate from the larger cluster of PERK/ISR targets (green in Fig. 1C-D).…”

Section: A Targeted Rnaseq Assay To Monitor Activation Of Stress-respmentioning

confidence: 83%

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Deconvoluting Stress-Responsive Proteostasis Signaling Pathways for Pharmacologic Activation Using Targeted RNA Sequencing

et al. 2019

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Cellular proteostasis is maintained by stress-responsive signaling pathways such as the heat shock response (HSR), the oxidative stress response (OSR), and the unfolded protein response (UPR). Activation of these pathways results in the transcriptional upregulation of select subsets of stressresponsive genes that restore proteostasis and adapt cellular physiology to promote recovery following various types of acute insult. The capacity for these pathways to regulate cellular proteostasis makes them attractive therapeutic targets to correct proteostasis defects associated with diverse diseases. High-throughput screening (HTS) using cell-based reporter assays is highly effective for identifying putative activators of stress-responsive signaling pathways. However, the development of these compounds is hampered by the lack of medium-throughput assays to define compound potency and selectivity for a given pathway. Here, we describe a targeted RNA sequencing (RNAseq) assay that allows cost effective, medium-throughput screening of stressresponsive signaling pathway activation. We demonstrate that this assay allows deconvolution of stress-responsive signaling activated by chemical genetic or pharmacologic agents. Furthermore, we use this assay to define the selectivity of putative OSR and HSR activating compounds previously identified by HTS. Our results demonstrate the potential for integrating this adaptable targeted RNAseq assay into screening programs focused on developing pharmacologic activators of stress-responsive signaling pathways.

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Section: ■ Results and Discussionmentioning

confidence: 88%

Section: A Targeted Rnaseq Assay To Monitor Activation Of Stress-respmentioning

confidence: 83%

Deconvoluting Stress-Responsive Proteostasis Signaling Pathways for Pharmacologic Activation Using Targeted RNA Sequencing

et al. 2019

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“…Structural proteomics is used to characterize the protein structure or the protein complexes present in a cell type or organelle. Finally, functional proteomics enables characterization of the biological function and cellular mechanisms of proteins [ 45 ]. Combined knowledge of protein function, activity, and post-translational modifications, as well as protein–protein interactions, provides valuable information about cell stage and therefore about the disease.…”

Section: Proteomics For the Study Of Imdmentioning

confidence: 99%

“…Combined knowledge of protein function, activity, and post-translational modifications, as well as protein–protein interactions, provides valuable information about cell stage and therefore about the disease. In the study of rare diseases, this information can facilitate the development of more diagnostically useful tools and identification of therapeutic targets, ultimately providing more information about the pathophysiology of the disease [ 45 ].…”

Section: Proteomics For the Study Of Imdmentioning

confidence: 99%

Proteomics in Inherited Metabolic Disorders

Chantada-Vázquez

Bravo

Gouveia

et al. 2022

IJMS

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Inherited metabolic disorders (IMD) are rare medical conditions caused by genetic defects that interfere with the body’s metabolism. The clinical phenotype is highly variable and can present at any age, although it more often manifests in childhood. The number of treatable IMDs has increased in recent years, making early diagnosis and a better understanding of the natural history of the disease more important than ever. In this review, we discuss the main challenges faced in applying proteomics to the study of IMDs, and the key advances achieved in this field using tandem mass spectrometry (MS/MS). This technology enables the analysis of large numbers of proteins in different body fluids (serum, plasma, urine, saliva, tears) with a single analysis of each sample, and can even be applied to dried samples. MS/MS has thus emerged as the tool of choice for proteome characterization and has provided new insights into many diseases and biological systems. In the last 10 years, sequential window acquisition of all theoretical fragmentation spectra mass spectrometry (SWATH-MS) has emerged as an accurate, high-resolution technique for the identification and quantification of proteins differentially expressed between healthy controls and IMD patients. Proteomics is a particularly promising approach to help obtain more information on rare genetic diseases, including identification of biomarkers to aid early diagnosis and better understanding of the underlying pathophysiology to guide the development of new therapies. Here, we summarize new and emerging proteomic technologies and discuss current uses and limitations of this approach to identify and quantify proteins. Moreover, we describe the use of proteomics to identify the mechanisms regulating complex IMD phenotypes; an area of research essential to better understand these rare disorders and many other human diseases.

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“…The parallel use of an antibody panel and multiple lectin probing provides discrete glycosylation profiles of multiple proteins in a given sample [18][19][20] . This method has been used successfully in multiple different labs 1,7,13,[19][20][21][22][23][24][25][26][27][28][29][30][31] . However, stability of MPBH and Cys-Gly, complicated and extended procedure in this method affect the reproducibility, effectiveness and efficiency of the method.…”

Section: Introductionmentioning

confidence: 99%

Chemically-blocked Antibody Microarray for Multiplexed High-throughput Profiling of Specific Protein Glycosylation in Complex Samples

Wonsidler

et al. 2012

JoVE

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In this study, we describe an effective protocol for use in a multiplexed high-throughput antibody microarray with glycan binding protein detection that allows for the glycosylation profiling of specific proteins. Glycosylation of proteins is the most prevalent post-translational modification found on proteins, and leads diversified modifications of the physical, chemical, and biological properties of proteins. Because the glycosylation machinery is particularly susceptible to disease progression and malignant transformation, aberrant glycosylation has been recognized as early detection biomarkers for cancer and other diseases. However, current methods to study protein glycosylation typically are too complicated or expensive for use in most normal laboratory or clinical settings and a more practical method to study protein glycosylation is needed. The new protocol described in this study makes use of a chemically blocked antibody microarray with glycan-binding protein (GBP) detection and significantly reduces the time, cost, and lab equipment requirements needed to study protein glycosylation. In this method, multiple immobilized glycoprotein-specific antibodies are printed directly onto the microarray slides and the N-glycans on the antibodies are blocked. The blocked, immobilized glycoprotein-specific antibodies are able to capture and isolate glycoproteins from a complex sample that is applied directly onto the microarray slides. Glycan detection then can be performed by the application of biotinylated lectins and other GBPs to the microarray slide, while binding levels can be determined using Dylight 549-Streptavidin. Through the use of an antibody panel and probing with multiple biotinylated lectins, this method allows for an effective glycosylation profile of the different proteins found in a given human or animal sample to be developed.

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Proteomics as Applied to Inherited Metabolic Diseases

Cited by 4 publications

References 92 publications

Deconvoluting Stress-Responsive Proteostasis Signaling Pathways for Pharmacologic Activation Using Targeted RNA Sequencing

Deconvoluting Stress-Responsive Proteostasis Signaling Pathways for Pharmacologic Activation Using Targeted RNA Sequencing

Proteomics in Inherited Metabolic Disorders

Chemically-blocked Antibody Microarray for Multiplexed High-throughput Profiling of Specific Protein Glycosylation in Complex Samples

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