Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.4.25.1 (proteasome)
28,817 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We studied the effect of inhibition of microsomal triglyceride transfer protein (MTP) on apolipoprotein (apo) B100 translation and secretion using HepG2 cells. The MTP-mediated lipid transfer activity was reduced using a specific MTP inhibitor. ApoB100 translation was synchronized by treatment with puromycin prior to L-[35S]methionine pulse-chase labeling. During the first 4 min of chase, synthesis of apoB polypeptides the size of 100-200 kDa was insensitive to the inhibitor, suggesting that inhibition of MTP did not affect the initiation of apoB100 translation. After 15 min of chase, the 100-200-kDa species were chased into polypeptides larger than 320 kDa (i.e. apoB65 or 65% of full-length apoB100) in both control and inhibitor-treated cells. However, the amount of these polypeptides decreased (by 36% for apoB65-75, by 64% for apoB75-85, by 76% for apoB85-95, and by 77% for apoB100) upon MTP inhibition. No accumulation of smaller polypeptides was observed, but total immunoprecipitable apoB radioactivity was decreased suggesting that apoB could undergo co-translational degradation when MTP activity was reduced. Inhibitors of the multicatalytic proteinase complex (proteasome) such as lactacystin or MG-115 could prevent apoB co-translational degradation. Nevertheless, MG-115 could not avoid the MTP inhibitor decreasing apoB100 secretion but rather induced the accumulation of secretion-incompetent apoB100 in the cell. These results indicate that MTP activity is required during the elongation of apoB100 polypeptides, particularly at the sequences downstream of carboxyl terminus of apoB65. Co-translational degradation might constitute a more general mechanism of early quality control for large or complex proteins.
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PMID:Co-translational degradation of apolipoprotein B100 by the proteasome is prevented by microsomal triglyceride transfer protein. Synchronized translation studies on HepG2 cells treated with an inhibitor of microsomal triglyceride transfer protein. 925 52

Intracellular degradation of newly synthesized apolipoprotein (apo) B can occur at every stage of the secretory pathway, from the protein translation, polypeptide translocation across the membrane of endoplasmic reticulum (ER), to vesicular transport. The prevalence of apoB degradation at each stage varies in different hepatic cell systems examined. Proteolysis of nascent apoB can be catalyzed by the ubiquitin-proteasome system in the cytosol, and probably by unidentified ER resident proteases as well. Cytosolic and ER lumenal molecular chaperones that facilitate apoB translocation and folding may also assist in the degradation of misfolded apoB proteins. Factors affecting the synthesis and mobilization of lipids during lipoprotein assembly exert important regulatory effects on apoB degradation in trans, and specific hydrophobic amino acid sequence elements within the apoB-100 molecule may play roles in apoB degradation in cis. This review summarizes the current understanding of the cellular and molecular mechanisms responsible for intracellular degradation of apoB in hepatocytes. The emphasis centers primarily on the topology of apoB with respect to the ER membrane during and after apoB translation and its relationship to proteolytic mechanisms potentially involved in apoB degradation.
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PMID:Intracellular degradation of newly synthesized apolipoprotein B. 937 17

Plasma levels of atherogenic lipoprotein [a] (Lp[a]) vary over a 1000-fold range and are largely determined by the gene for its unique glycoprotein, apolipoprotein [a] (apo[a]). The apo[a] locus comprises more than 100 alleles, encoding proteins from <300 to >800 kDa. Using primary baboon hepatocyte cultures, we previously demonstrated that differences in the secretion efficiency of apo[a] allelic variants contribute to the variation in plasma Lp[a] levels. In the current study, we investigated the mechanism of apo[a] presecretory degradation. The proteasome inhibitors, acetyl-leucyl-leucyl-norleucinal and lactacystin, prevented apo[a] degradation and increased apo[a] secretion. Transfection with an HA-tagged ubiquitin construct demonstrated the accumulation of ubiquitinated apo[a] in the presence of lactacystin. These results suggest a role for the cytoplasmic proteasome in apo[a] proteolysis. Apo[a] that accumulated intracellularly in the presence of lactacystin remained sensitive to endo-B-N-glucosaminidase H, and apo[a] degradation was reversibly inhibited by brefeldin A, suggesting that transport to a post-endoplasmic reticulum (ER) pre-medial Golgi compartment is required for apo[a] degradation. Newly synthesized apo[a] bound to the ER chaperone calnexin and conditions that enhanced this interaction prevented apo[a] degradation, suggesting that calnexin can protect apo[a] from proteolysis. These studies provide further support for the role of the proteasome in endoplasmic reticulum quality control, and expand this role to one that influences plasma levels of the atherogenic lipoprotein Lp[a].-White, A. L., B. Guerra, J. Wang, and R. E. Lanford. Presecretory degradation of apolipoprotein[a] is mediated by the proteasome pathway.
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PMID:Presecretory degradation of apolipoprotein [a] is mediated by the proteasome pathway. 992 57

Studies in different liver-derived cells in culture indicate that apolipoprotein (apo) B-100 production is regulated largely by intracellular degradation and the ubiquitin-proteasome pathway is a major mechanism for the degradation. The proteasomal degradation of apoB-100 was postulated to be an intrinsic property of the protein that occurs even in the presence of optimal amounts of lipids supplied to the cell. We examined apoB-100 and apoB-48 biogenesis in CaCo2, a human colon carcinoma cell line. To our surprise, apoB-100 and apoB-48 were quantitatively secreted by CaCo2 cells; essentially none of the newly synthesized apoB was degraded before secretion in a 2-h period whether the cells were cultured on filter or on plastic. Furthermore, although ubiquitin immunoreactivity was readily detected in the intracellular apoB isolated from HepG2 cells, little or no ubiquitin was detectable in the intracellular apoB from CaCo2 cells. The amounts of free ubiquitin and total and non-apoB ubiquitinated proteins were comparable in HepG2 and CaCo2 cells, indicating that CaCo2 cells have the necessary machinery for tagging ubiquitin chains onto cellular proteins for proteasomal degradation. Incubation in lipoprotein-deficient serum did not induce apoB degradation, but the addition of a microsomal triglyceride transfer protein inhibitor led to apoB degradation in CaCo2 cells. Finally, similar proportions of apoB polypeptide in isolated microsomes from CaCo2 and HepG2 cells were accessible to exogenously added trypsin, indicating that the mere exposure of apoB nascent chains to the cytosolic compartment is insufficient to cause the proteasomal degradation. Therefore, the intracellular degradation of apoB is not an intrinsic property of the protein, and the phenomenon is neither universal nor inevitable. The unconditional use of apoB as a paradigm for intracellular protein degradation is not warranted.
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PMID:Apolipoprotein B, a paradigm for proteins regulated by intracellular degradation, does not undergo intracellular degradation in CaCo2 cells. 1066 May 49

Highly active antiretroviral therapy, which includes a combination of protease inhibitors, is highly successful in controlling human immunodeficiency virus (HIV) infection and reducing the morbidity and mortality of autoimmune deficiency syndrome (AIDS). However, the benefits of HIV protease inhibitors are compromised by numerous undesirable side effects. These include peripheral fat wasting and excessive central fat deposition (lipodystrophy), overt hyperlipidemia, and insulin resistance. The mechanism associated with protease inhibitor-induced metabolic abnormalities is multifactorial. One major effect of the protease inhibitor is its suppression of the breakdown of the nuclear form of sterol regulatory element binding proteins (nSREBP) in the liver and adipose tissues. Hepatic accumulation of nSREBP results in increased fatty acid and cholesterol biosynthesis, whereas nSREBP accumulation in adipose tissue causes lipodystrophy, reduces leptin expression, and promotes insulin resistance. The HIV protease inhibitors also suppress proteasome-mediated breakdown of nascent apolipoprotein (apo) B, thus resulting in the overproduction and secretion of triglyceride-rich lipoproteins. Finally, protease inhibitor also suppresses the inhibition of the glucose transporter GLUT-4 activity in adipose and muscle. This latter effect also contributes directly to insulin resistance and diabetes. These adverse effects need to be alleviated for long-term use of protease inhibitor therapy in treatment of HIV infection.
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PMID:Effects of HIV protease inhibitor therapy on lipid metabolism. 1254 52

The aim of our study was to monitor the protein expression profile in pituitary glands of healthy C57BL/6J mice during aging. Pituitary glands of 4-week old (immature), 3-month old (mature), and >25-month old mice were analysed by proteomic tools such as two-dimensional electrophoresis and N-terminal micro-sequencing. A change was detected in the expression of growth hormone after sexual maturation. Our particular interest, however, was directed against up-regulated proteins in the old pituitary glands, which are proposed to be involved in the process of neuroendocrine aging. Among these proteins, the expression of glutathione-S-transferase (GST) and apolipoprotein A-1 were increased in old pituitaries. Furthermore, ubiquitin carboxyl-terminal hydrolase (UCH-L1) was significantly up-regulated in senescent C57BL/6J mouse pituitaries. Since only the rat homologue was known, we isolated and analysed the mouse UCH-L1 sequence. Since GST is involved in antioxidative defence and UCH-L1 is part of the ubiquitin/proteasome system, which is responsible for the removal of damaged proteins, these results suggest increased oxidative burden and an increased activity of the ubiquitin system.
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PMID:Age-related alterations in the protein expression profile of C57BL/6J mouse pituitaries. 1255 14

Studies in hepatocyte cultures indicate that apolipoprotein (apo) B-100 production is regulated largely by intracellular degradation and the proteasome pathway is a major mechanism for the degradation. In the present study, we have examined the detailed itinerary of apoB degradation through its secretory pathway in HepG2 cells. We found that ubiquitin-dependent proteasomal degradation of apoB largely occurred on the cytosolic surface of rough and smooth endoplasmic reticulum (ER) and that a small proportion of apoB was dislodged from the secretory organelles into the cytosolic compartment where it underwent ubiquitination for proteasomal degradation. The transmembrane conformation of apoB persisted as the protein was transported through the Golgi apparatus. We further demonstrated that proteasomal degradation of apoB was associated the Golgi apparatus but Golgi-associated apoB was not ubiquitinated, indicating an ubiquitin-independent proteasomal degradation of apoB is associated with this organelle. We conclude that apoB undergoes proteasomal degradation while going through different compartments of the secretory pathway; further, ER-associated proteasomal degradation of apoB in the ER is ubiquitin-dependent whereas that occurring in the Golgi is ubiquitin-independent.
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PMID:Ubiquitin-dependent and -independent proteasomal degradation of apoB associated with endoplasmic reticulum and Golgi apparatus, respectively, in HepG2 cells. 1287 35

By comparative proteome analysis we searched for characteristic alterations of human stomach adenocarcinoma tissue and paired surrounding normal tissue. Selected differential protein spots were identified with peptide mass fingerprinting based on matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF) and database searching. We identified protein alterations in 18 stomach cancer tissues compared with normal controls, comprising elevated levels of eight proteins, including 14-3-3 zeta, calcyclin, keratin, apolipoprotein A-1 precursor, proteasome activator complex subunit, nucleoside diphosphate kinase, nicotinamide N-methyltransferase, and pyridoxal kinase. Five proteins (CA11, prohibitin, peroxiredoxin 4, serum amyloid P component, and NADH-ubiquinone oxidoreductase 23 kDa subunit) were decreased. These data are valuable for identification of differentially expressed proteins involved in stomach cancer carcinogenesis, providing biomarker candidates to develop diagnostic and therapeutic tools.
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PMID:The differential proteome profile of stomach cancer: identification of the biomarker candidates. 1555 63

Genetic deficiency of the plasma phospholipid transfer protein (PLTP) in mice unexpectedly causes a substantial impairment in liver secretion of apolipoprotein-B (apoB), the major protein of atherogenic lipoproteins. To explore the mechanism, we examined the three known pathways for hepatic apoB secretory control, namely endoplasmic reticulum (ER)/proteasome-associated degradation (ERAD), post-ER pre-secretory proteolysis (PERPP), and receptor-mediated degradation, also known as re-uptake. First, we found that ERAD and cell surface re-uptake were not active in PLTP-null hepatocytes. Moreover, ER-to-Golgi blockade by brefeldin A, which enhances ERAD, equalized total apoB recovery from PLTP-null and wild-type cells, indicating that the relevant process occurs post-ER. Second, because PERPP can be stimulated by intracellular reactive oxygen species (ROS), we examined hepatic redox status. Although we found previously that PLTP-null mice exhibit elevated plasma concentrations of vitamin E, a lipid anti-oxidant, we now discovered that their livers contain significantly less vitamin E and significantly more lipid peroxides than do livers of wild-type mice. Third, to establish a causal connection, the addition of vitamin E or treatment with an inhibitor of intracellular iron-dependent peroxidation, desferrioxamine, abolished the elevation in cellular ROS as well as the defect in apoB secretion from PLTP-null hepatocytes. Overall, we conclude that PLTP deficiency decreases liver vitamin E content, increases hepatic oxidant tone, and substantially enhances ROS-dependent destruction of newly synthesized apoB via a post-ER process. These findings are likely to be broadly relevant to hepatic apoB secretory control in vivo.
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PMID:Phospholipid transfer protein deficiency impairs apolipoprotein-B secretion from hepatocytes by stimulating a proteolytic pathway through a relative deficiency of vitamin E and an increase in intracellular oxidants. 1573 42

Fibrate drugs improve cardiovascular health by lowering plasma triglycerides, normalize low density lipoprotein levels, and raise high density lipoprotein (HDL) levels in patients with dyslipidemias. The HDL-raising effect of fibrates has been shown to be due in part to an increase in human apolipoprotein AI gene expression. However, it has recently been shown that fibrates can affect HDL metabolism in mouse by significantly decreasing hepatic levels of the HDL receptor scavenger receptor B-I (SR-BI) and the PDZ domain containing protein PDZK1. PDZK1 is essential for maintaining hepatic SR-BI levels. Therefore, decreased SR-BI might be secondary to decreased PDZK1, but the mechanism by which fibrates lower SR-BI has not been elucidated. Here we show that feeding PDZK1-deficient mice fenofibrate resulted in the near absence of SR-BI in liver, definitively demonstrating that the effect of fenofibrate on SR-BI is PDZK1-independent. Metabolic labeling experiments in primary hepatocytes from fenofibrate-fed mice demonstrated that fenofibrate enhanced the degradation of SR-BI in a post-endoplasmic reticulum compartment. Moreover, fenofibrate-induced degradation of SR-BI was independent of the proteasome, calpain protease, or the lysosome, and antioxidants did not inhibit fenofibrate-induced degradation of SR-BI. Using metabolic labeling coupled with cell surface biotinylation assays, fenofibrate did not inhibit SR-BI trafficking to the plasma membrane. Together, the data support a model in which fenofibrate enhances the degradation of SR-BI in a post-ER, post-plasma membrane compartment. The further elucidation of this novel degradation pathway may provide new insights into the physiological and pathophysiological regulation of hepatic SR-BI.
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PMID:Fenofibrate induces a novel degradation pathway for scavenger receptor B-I independent of PDZK1. 1583 86


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