Comparative analysis of extra- and intracellular proteasomes from K562 cells

Zaikova, Yu. Ya.; Куличкова, В. А.; Ermolaeva, Yu. B.; Gause, L. N.; Цимоха, А. С.

doi:10.1134/s1990519x11050130

Cited by 5 publications

(4 citation statements)

References 21 publications

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“…Previously, we have shown that proteasomes are detectable in CM conditioned by human tumor cell lines [ 21 , 28 ]. It is known that expression of proteasomes is overall enhanced (compared to normal cells) but varies amongst different tumor cells [ 29 – 36 ].…”

Section: Resultsmentioning

confidence: 99%

Proteomic analysis of affinity-purified extracellular proteasomes reveals exclusively 20S complexes

et al. 2017

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Proteasome-mediated proteolysis is important for many basic cellular processes. In addition to their functions in the cell, proteasomes have been found in physiological fluids of both healthy and diseased humans including cancer patients. Higher levels of these proteasomes are associated with higher cancer burden and stage. The etiology and functions of these proteasomes, referred to as circulating, plasmatic, or extracellular proteasomes (ex-PSs), are unclear. Here we show that human cancer cell lines, as well as human endometrium-derived mesenchymal stem cells (hMESCs), release proteasome complexes into culture medium (CM). To define ex-PS composition, we have affinity purified them from CM conditioned by human leukemia cell line K562. Using matrix-assisted laser desorption/ionization (MALDI) Fourier transform ion cyclotron resonance (FT-ICR) mass spectrometry (MS), we have identified core 20S proteasome subunits and a set of 15 proteasome-interacting proteins (PIPs), all previously described as exosome cargo proteins. Three of them, PPIase A, aldolase A, and transferrin, have never been reported as PIPs. The study provides compelling arguments that ex-PSs do not contain 19S or PA200 regulatory particles and are represented exclusively by the 20S complex.

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Section: Resultsmentioning

confidence: 99%

Proteomic analysis of affinity-purified extracellular proteasomes reveals exclusively 20S complexes

et al. 2017

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“…Previously, we have shown that extracellular proteasomes (EPs) can be detected in the medium conditioned by cultivated human cells from different tumor cell lines (Kulichkova et al, ; Zaikova et al, ). Here we report a characterization of extracellular proteasomes by the iTRAQ‐based quantitative proteomics approach.…”

Section: Resultsmentioning

confidence: 99%

“…Recently, proteasomes have also been detected in the extracellular space, such in normal blood plasma, alveolar and cerebrospinal fluids (Sixt and Dahlmann, ; Sixt and Peters, ), and in the culture medium of human cancer cell lines (Kulichkova et al, ,; Zaikova et al, ). The concentrations of these extracellular proteasomes has been found elevated in patients with various pathological conditions, such hematologic malignancies, autoimmune diseases, or critical diseases.…”

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confidence: 99%

Extracellular Proteasomes Are Deficient in 19S Subunits as Revealed by iTRAQ Quantitative Proteomics

Цимоха

Zaykova

Bottrill³

et al. 2016

Journal Cellular Physiology

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Proteasome-mediated proteolysis is critical for regulation of vast majority of cellular processes. In addition to their well-documented functions in the nucleus and cytoplasm proteasomes have also been found in extracellular space. The origin and functions of these proteasomes, dubbed as circulating/plasmatic or extracellular proteasomes, are unclear. To gain insights into the molecular and functional differences between extracellular (EPs) and cellular proteasomes (CPs) we compared their subunit composition using iTRAQ-based quantitative proteomics (iTRAQ LC/MS-MS). Our analysis of purified from K562 cells or conditioned medium intact proteasome complexes led to an identification and quantification of 114 proteins, out of which 19 were 26S proteasome proteins (all subunits of the 20S proteasome and a small number of the 19S regulatory particle proteins), and 3 belonged to the ubiquitin system. Sixty-two of proteasome interacting proteins (PIPs) were differentially represented in CP versus EP, with folds difference ranging from 1.5 to 4.8. The bioinformatics analysis revealed that functionally most of EP-PIPs were associated with protein biosynthesis and, unlike CP-PIPs, were under represented by chaperon/ATP-binding proteins. Identities of some of the proteasome proteins and PIPs were verified by Western blotting. Importantly, we uncovered that the stoichiometry of the 20S versus 19S complexes in the extracellular proteasomes was different compared to the one calculated for the intracellular proteasomes. Specifically, the EP prep contained only three 19S subunits versus at least 18 in the CP one, suggesting that the extracellular proteasomes are deficient in 19S complexes, which may imply that they have special biological functions. J. Cell. Physiol. 232: 842-851, 2017. © 2016 Wiley Periodicals, Inc.

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“…Secondly, surrounding cells are likely to supply proteasomes to extracellular environment by exocytosis to hydrolyze matrix or other substrates and provide tumor invasion as matrix metalloproteinases do [35] . This viewpoint is based on the fact of existence of extracellular proteasome pool [36] .…”

Section: Discussionmentioning

confidence: 99%

Proteasome Functioning in Breast Cancer: Connection with Clinical-Pathological Factors

et al. 2014

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Breast cancer is one of four oncology diseases that are most widespread in the world. Moreover, breast cancer is one of leading causes of cancer-related deaths in female population within economically developed regions of the world. So far, detection of new mechanisms of breast cancer development is very important for discovery of novel areas in which therapy approaches may be elaborated. The objective of the present study is to investigate involvement of proteasomes, which cleave up to 90% of cellular proteins and regulate numerous cellular processes, in mechanisms of breast cancer development. Proteasome characteristics in 106 patient breast carcinomas and adjacent tissues, as well as relationships of detected proteasome parameters with clinical-pathological factors, were investigated. Proteasome chymotrypsin-like activity was evaluated by hydrolysis of fluorogenic peptide Suc-LLVY-AMC. The expression of proteasome subunits was studied by Western-blotting and immunohistochemistry. The wide range of chymotrypsin-like activity in tumors was detected. Activity in tumors was higher if compared to adjacent tissues in 76 from 106 patients. Multiple analysis of generalized linear models discovered that in estrogen α-receptor absence, tumor growth was connected with the enhanced expression of proteasome immune subunit LMP2 and proteasome activator PA700 in tumor (at 95% confidence interval). Besides, by this analysis we detected some phenomena in adjacent tissue, which are important for tumor growth and progression of lymph node metastasis in estrogen α-receptor absence. These phenomena are related to the enhanced expression of activator PA700 and immune subunit LMP7. Thus, breast cancer development is connected with functioning of immune proteasome forms and activator PA700 in patients without estrogen α-receptors in tumor cells. These results could indicate a field for search of new therapy approaches for this category of patients, which has the worst prognosis of health recovery.

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Comparative analysis of extra- and intracellular proteasomes from K562 cells

Cited by 5 publications

References 21 publications

Proteomic analysis of affinity-purified extracellular proteasomes reveals exclusively 20S complexes

Proteomic analysis of affinity-purified extracellular proteasomes reveals exclusively 20S complexes

Extracellular Proteasomes Are Deficient in 19S Subunits as Revealed by iTRAQ Quantitative Proteomics

Proteasome Functioning in Breast Cancer: Connection with Clinical-Pathological Factors

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