Super-SILAC mix coupled with SIM/AIMS assays for targeted verification of phosphopeptides discovered in a large-scale phosphoproteome analysis of hepatocellular carcinoma

Lin, Yung Hsiang; Chien, Kun-Yi; Wu, Chia‐Chun; Chang, Wen‐Yu; Chu, Lichieh Julie; Chen, Min-Chi; Yeh, Chau‐Ting; Yu, Jau‐Song

doi:10.1016/j.jprot.2017.02.005

Cited by 8 publications

(4 citation statements)

References 53 publications

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“…β ‐casein and BSA/ β ‐casein mixture) and also human serum. Phosphoproteome analysis in biological fluids has attracted significant attention in recent years since a linkage between the phosphorylated forms of proteins/peptides and tumor formation has been shown for different tumor types (Gökmen et al, ; Hong et al, ; Labots et al, ; Lin et al, ; Zhao et al, ; Zhao et al, ). Moreover, in our case, the use of a capillary monolith with larger diameter and shorter length allowed us to work with relatively lower back pressures (i.e.…”

Section: Resultsmentioning

confidence: 99%

Ti (IV) attached‐phosphonic acid functionalized capillary monolith as a stationary phase for in‐syringe‐type fast and robust enrichment of phosphopeptides

Salimi

Kip

Çelikbıçak

et al. 2019

Biomedical Chromatography

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In this study, poly(vinylphosphonic acid-co-ethylene dimethacrylate), poly(VPA-co-EDMA) capillary monolith was synthesized as a starting material for obtaining a stationary phase for microscale enrichment of phosphopeptides. The chelation of active phosphonate groups with Ti (IV) ions gave a macroporous monolithic column with a mean pore size of 5.4 μm. The phosphopeptides from different sources were enriched on Ti (IV)-attached poly(VPA-co-EDMA) monolith using a syringe-pump. The monolithic capillary columns exhibited highly sensitive/selective enrichment performance with phosphoprotein concentrations as low as 1.0 fmol/mL. Six different phosphopeptides were detected with high intensity by the treatment of β-casein digest with the concentration of 1.0 fmol/mL, using Ti (IV)@poly(VPA-co-EDMA) monolith. Highly selective enrichment of phosphopeptides was also successfully carried out even at trace amounts, in a complex mixture of digested proteins (molar ratio of β-casein to bovine serum albumin, 1:1500) and three phosphopeptides were successfully detected. Four highly intense signals of phosphopeptides in human serum were also observed with high signal-to-noise ratio and a clear background after enrichment with Ti (IV)@poly(VPA-co-EDMA) monolith. It was concluded that the capillary microextraction system enabled fast, efficient and robust enrichment of phosphopeptides from microscale complex samples. The whole enrichment process was completed within 20 min, which was shorter than in the previously reported studies.KEYWORDS capillary monolith, human serum, immobilized metal affinity chromatography, phosphopeptide enrichment, syringe type microextration, vinylphosphonic acid

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Section: Resultsmentioning

confidence: 99%

Ti (IV) attached‐phosphonic acid functionalized capillary monolith as a stationary phase for in‐syringe‐type fast and robust enrichment of phosphopeptides

Salimi

Kip

Çelikbıçak

et al. 2019

Biomedical Chromatography

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“…Lin and colleagues combined stable isotope dimethylation labeling coupled with online 3D SCX-TiO 2 /RP using an LTQ-Orbitrap and super-SILAC mix coupled with selected ion monitoring/ accurate inclusion mass screening (SIM/AIMS), in order to develop a strategy for both the discovery and targeted verification of differentially phosphorylated proteins in three HCCs and their nearby tissue specimens. Their workflow led to the identification of 7868 phosphopeptides [ 34 ].…”

Section: Quantitative Phosphoproteomics and Glycoproteomics Stratementioning

confidence: 99%

Recent Advances of Functional Proteomics in Gastrointestinal Cancers- a Path towards the Identification of Candidate Diagnostic, Prognostic, and Therapeutic Molecular Biomarkers

Abyadeh

Meyfour

Gupta

et al. 2020

IJMS

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Gastrointestinal (GI) cancer remains one of the common causes of morbidity and mortality. A high number of cases are diagnosed at an advanced stage, leading to a poor survival rate. This is primarily attributed to the lack of reliable diagnostic biomarkers and limited treatment options. Therefore, more sensitive, specific biomarkers and curative treatments are desirable. Functional proteomics as a research area in the proteomic field aims to elucidate the biological function of unknown proteins and unravel the cellular mechanisms at the molecular level. Phosphoproteomic and glycoproteomic studies have emerged as two efficient functional proteomics approaches used to identify diagnostic biomarkers, therapeutic targets, the molecular basis of disease and mechanisms underlying drug resistance in GI cancers. In this review, we present an overview on how functional proteomics may contribute to the understanding of GI cancers, namely colorectal, gastric, hepatocellular carcinoma and pancreatic cancers. Moreover, we have summarized recent methodological developments in phosphoproteomics and glycoproteomics for GI cancer studies.

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“…To date, many reports about LMNA-related diseases are available. For example, some evidence supports the function of LMNA as an oncogene in gastrointestinal tract tumors [10], prostate cancer (PC) [11], HCC [12,13], and metastatic colorectal cancer (CRC) [14][15][16]. In addition, mutations in LMNA induce common laminopathies characterized by multiple rare clinical symptoms, including muscular dystrophy, lipodystrophy, diabetes, dermopathy, neuropathy, leukodystrophy, and progeria [17][18][19].…”

Section: Introductionmentioning

confidence: 99%

Abnormal in LMNA Triggers a Shift in Intracellular Metabolic Homeostasis Via AMPK Activation

Zhao¹,

Yang²,

Cheng³

et al. 2022

Preprint

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Laminopathies are a spectrum of diseases caused by LMNA mutations. In familial partial lipodystrophy of Dunnigan (FPLD), LMNA plays role in the differentiation and development of adipocytes and lipid metabolism. Changes in LMNA predict not only the differentiation of adipose-derived mesenchymal stem cells (AD-MSCs) but also the transformation of cancer cells. Hence, our in-depth study aimed to identify the molecular connection between disordered lipid metabolism and hepatic carcinogenesis. We first discovered significant positive correlations between pLMNA and two key rate-limiting enzymes in de novo fatty acid synthesis, acetyl-CoA-carboxylase 1 (ACC1) and fatty acid synthase (FASN), in the liver tissue but not in adipose tissue of obese model rats. Moreover, LMNA knockdown (KD) in rat AD-MSCs prevented the differentiation and maturation of adipocytes. To clarify the mechanistic relationship with lipogenesis, gain- and loss-of-function experiments in which functional changes and the related molecular pathways were investigated in a normal hepatocyte line (7701 cells). Adenosine 5'-monophosphate activated protein kinase α (AMPKα) was found to be activated by abnormalities in the LMNA structure under conditions of LMNA deletion, farnesyltransferase inhibitor (FTI) treatment and LMNA mutations associated with clinical FPLD pathogenic phenotype. Active AMPKα could directly phosphorylate ACC1 and thus inhibit lipid synthesis but induced glycolysis in both HCC cells and normal cells. The HCC cells could not survive with LMNA knockout (KO) or even KD. Lonafarnib (an FTI) combined with low-glucose conditions significantly decreased the proliferation of HepG2 and MHCC cells by inhibiting glycolysis and the maturation of prelamin A.

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Super-SILAC mix coupled with SIM/AIMS assays for targeted verification of phosphopeptides discovered in a large-scale phosphoproteome analysis of hepatocellular carcinoma

Cited by 8 publications

References 53 publications

Ti (IV) attached‐phosphonic acid functionalized capillary monolith as a stationary phase for in‐syringe‐type fast and robust enrichment of phosphopeptides

Ti (IV) attached‐phosphonic acid functionalized capillary monolith as a stationary phase for in‐syringe‐type fast and robust enrichment of phosphopeptides

Recent Advances of Functional Proteomics in Gastrointestinal Cancers- a Path towards the Identification of Candidate Diagnostic, Prognostic, and Therapeutic Molecular Biomarkers

Abnormal in LMNA Triggers a Shift in Intracellular Metabolic Homeostasis Via AMPK Activation

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