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PubMed2004

The use of in vitro peptide-library screens in the analysis of phosphoserine/threonine-binding domain structure and function.

Phosphoserine/threonine-binding domains integrate intracellular signal transduction events by forming multiprotein complexes with substrates of protein serine/threonine kinases. These phosphorylation-dependent molecular recognition events are responsible for coordinating the precise temporal and spatial response of cells to a wide range of stimuli, particularly those involved in cell cycle control and the response to DNA damage. The known families of phosphoserine/threonine-binding modules include 14-3-3 proteins, WW domains, FHA domains, WD40 repeats, and the Polo-box domains of Polo-like kinases. Peptide-library experiments reveal the optimal sequence motifs recognized by these domains, and facilitate high-resolution structural studies elucidating the mechanisms of phospho-dependent binding and the molecular basis for domain function within intricate signaling networks. Information emerging from these studies is critical for the design of novel experimental and therapeutic tools aimed at altering signal transduction cascades in normal and diseased cells.

Amino Acid Motifs↗

PubMed2003

Structure, function, and regulation of budding yeast kinetochores.

Kinetochores are multiprotein complexes that assemble on centromeric DNA and mediate the attachment and movement of chromosomes along the microtubules (MTs) of the mitotic spindle. This review focuses on the simplest eukaryotic centromeres and kinetochores, those found in the budding yeast Saccharomyces cerevisiae. Research on kinetochore function and chromosome segregation is focused on four questions of general significance: what specifies the location of centromeres? What are the protein components of kinetochores, and how do they assemble a MT attachment site? How do MT attachments generate force? How do cells sense the state of attachment via the spindle assembly checkpoint?

Cell Division↗

PubMed2006

Tight junctions and cell polarity.

The tight junction is an intracellular junctional structure that mediates adhesion between epithelial cells and is required for epithelial cell function. Tight junctions control paracellular permeability across epithelial cell sheets and also serve as a barrier to intramembrane diffusion of components between a cell's apical and basolateral membrane domains. Recent genetic and biochemical studies in invertebrates and vertebrates indicate that tight junction proteins play an important role in the establishment and maintenance of apico-basal polarity. Proteins involved in epithelial cell polarization form evolutionarily conserved multiprotein complexes at the tight junction, and these protein complexes regulate the architecture of epithelia throughout the polarization process. Accumulating information regarding the regulation of these polarity proteins will lead to a better understanding of the molecular mechanisms whereby cell polarity is established.

Animals↗

PubMed1998

The mitochondrial death/life regulator in apoptosis and necrosis.

Both physiological cell death (apoptosis) and, in some cases, accidental cell death (necrosis) involve a two-step process. At a first level, numerous physiological and some pathological stimuli trigger an increase in mitochondrial membrane permeability. The mitochondria release apoptogenic factors through the outer membrane and dissipate the electrochemical gradient of the inner membrane. Mitochondrial permeability transition (PT) involves a dynamic multiprotein complex formed in the contact site between the inner and outer mitochondrial membranes. The PT complex can function as a sensor for stress and damage, as well as for certain signals connected to receptors. Inhibition of PT by pharmacological intervention on mitochondrial structures or mitochondrial expression of the apoptosis-inhibitory oncoprotein Bcl-2 prevents cell death, suggesting that PT is a rate-limiting event of the death process. At a second level, the consequences of mitochondrial dysfunction (collapse of the mitochondrial inner transmembrane potential, uncoupling of the respiratory chain, hyperproduction of superoxide anions, disruption of mitochondrial biogenesis, outflow of matrix calcium and glutathione, and release of soluble intermembrane proteins) entails a bioenergetic catastrophe culminating in the disruption of plasma membrane integrity (necrosis) and/or the activation of specific apoptogenic proteases (caspases) by mitochondrial proteins that leak into the cytosol (cytochrome c, apoptosis-inducing factor) with secondary endonuclease activation (apoptosis). The relative rate of these two processes (bioenergetic catastrophe versus protease and endonuclease activation) determines whether a cell will undergo primary necrosis or apoptosis. The acquisition of the biochemical and ultrastructural features of apoptosis critically relies on the liberation of apoptogenic proteases or protease activators from mitochondria. The fact that mitochondrial events control cell death has major implications for the development of cytoprotective and cytotoxic drugs.

Animals↗

PubMed2006

DPP10 is an inactivation modulatory protein of Kv4.3 and Kv1.4.

Voltage-gated K(+) channels exist in vivo as multiprotein complexes made up of pore-forming and ancillary subunits. To further our understanding of the role of a dipeptidyl peptidase-related ancillary subunit, DPP10, we expressed it with Kv4.3 and Kv1.4, two channels responsible for fast-inactivating K(+) currents. Previously, DPP10 has been shown to effect Kv4 channels. However, Kv1.4, when expressed with DPP10, showed many of the same effects as Kv4.3, such as faster time to peak current and negative shifts in the half-inactivation potential of steady-state activation and inactivation. The exception was recovery from inactivation, which is slowed by DPP10. DPP10 expressed with Kv4.3 caused negative shifts in both steady-state activation and inactivation of Kv4.3, but no significant shifts were detected when DPP10 was expressed with Kv4.3 + KChIP2b (Kv channel interacting protein). DPP10 and KChIP2b had different effects on closed-state inactivation. At -60 mV, KChIP2b nearly abolishes closed-state inactivation in Kv4.3, whereas it developed to a much greater extent in the presence of DPP10. Finally, expression of a DPP10 mutant consisting of its transmembrane and cytoplasmic 58 amino acids resulted in effects on Kv4.3 gating that were nearly identical to those of wild-type DPP10. These data show that DPP10 and KChIP2b both modulate Kv4.3 inactivation but that their primary effects are on different inactivation states. Thus DPP10 may be a general modulator of voltage-gated K(+) channel inactivation; understanding its mechanism of action may lead to deeper understanding of the inactivation of a broad range of K(+) channels.

Animals↗

PubMed2006

ApoB mRNA editing is mediated by a coordinated modulation of multiple apoB mRNA editing enzyme components.

Apolipoprotein (apo)B mRNA editing is accomplished by a large multiprotein complex. How these proteins interact to achieve the precise single-nucleotide change induced by this complex remains unclear. We investigated the relationship between altered apoB mRNA editing and changes in editing enzyme components to evaluate their roles in editing regulation. In the mouse fetal small intestine, we found that the dramatic developmental upregulation of apoB mRNA editing from approximately 3% to 88% begins with decreased levels of inhibitory CUG binding protein 2 (CUGBP2) expression followed by increased levels of apoB mRNA editing enzyme (apobec)-1 and apobec-1 complementation factor (ACF) (4- and 8-fold) and then by decreased levels of the inhibitory components glycine-arginine-tyrosine-rich RNA binding protein (GRY-RBP) and heterogeneous nuclear ribonucleoprotein (hnRNP)-C1 (75% and 56%). In contrast, the expression of KH-type splicing regulatory protein (KSRP), apobec-1 binding protein (ABBP)1, ABBP2, and Bcl-2-associated athanogene 4 (BAG4) were unaltered. In the human intestinal cell line Caco-2, the increase of apoB mRNA editing from approximately 1.7% to approximately 23% was associated with 6- and 3.2-fold increases of apobec-1 and CUGBP2, respectively. In the mouse large intestine, the editing was 48% and had a 2.7-fold relatively greater CUGBP2 level. Caco-2 and the large intestine thus have increased instead of decreased CUGBP2 and a lower level of editing, suggesting that inhibitory CUGBP2 may play a critical role in the magnitude of editing regulation. Short interfering RNA-mediated gene-specific knockdown of CUGBP2, GRY-RBP, and hnRNP-C1 resulted in increased editing in Caco-2 cells, consistent with their known inhibitory function. These data suggest that a coordinated expression of editing components determines the magnitude and specificity of apoB mRNA editing.

Animals↗

PubMed2006

The emerging role of PDZ adapter proteins for regulation of intestinal ion transport.

In the gastrointestinal tract, CFTR, in conjunction with one or several members of the SLC26 anion exchanger family, mediates electrogenic Cl- and HCO3- secretion. Na+/H+ exchanger isoform NHE3, on the other hand, coupled to one or several of the SLC26 isoforms, mediates electroneutral NaCl absorption. The agonist-induced activation of anion secretion and inhibition of salt absorption causes secretory diarrhea. Current dogma sees the formation of a multiprotein complex of transport proteins, postsynaptic density-95/discs large/zonula occludens-1 (PDZ) adapter proteins, anchoring proteins, the cytoskeleton, and the involved protein kinases as one crucial step in the regulation of these transport processes. Data obtained in heterologous expression studies suggest an important role of these PDZ adapter proteins in trafficking, endocytic recycling, and membrane retention of the respective transmembrane proteins. This article reviews recent advances in our understanding of the role of the PDZ adapter proteins NHERF, E3KARP, PDZK1, IKEPP (NHERF-1 to NHERF-4), CAL, and Shank-2 that bind to CFTR, NHE3, and the intestinal SLC26 members in the regulation of intestinal fluid transport. Current concepts are mostly derived from heterologous expression studies and studies on their role in organ physiology are still in infancy. Recently, however, PDZ adapter protein-deficient mice and organ-specific cell lines have become available, and the first results suggest a more cell-type and possibly signal-specific role of these adapter proteins. This opens the potential for drug development targeted to PDZ domain interactions, which is, in theory, one of the most efficient antidiarrheal strategies.

Animals↗

PubMed1999

Carbachol activates ERK2 in isolated gastric parietal cells via multiple signaling pathways.

We previously reported that both carbachol and epidermal growth factor (EGF) are potent inducers of the extracellular signal-regulated protein kinases (ERKs) in isolated gastric canine parietal cells and that induction of these kinases leads to acute inhibitory and chronic stimulatory effects on gastric acid secretion. In this study we investigated the molecular mechanisms responsible for these effects. Both carbachol (100 microM) and EGF (10 nM) induced Ras activation. The role of Ras in ERK2 induction was examined by transfecting parietal cells with a vector expressing hemoagglutinin (HA)-tagged ERK2 (HA-ERK2) together with a dominantly expressed mutant (inactive) ras gene. HA-ERK2 activity was quantitated by in-gel kinase assays. Dominant negative Ras reduced carbachol induction of HA-ERK2 activity by 60% and completely inhibited the stimulatory effect of EGF. Since Ras activation requires the assembly of a multiprotein complex, we examined the effect of carbachol and EGF on tyrosyl phosphorylation of Shc and its association with Grb2 and the guanine nucleotide exchange factor Sos. Western blot analysis of anti-Shc immunoprecipitates with an anti-phosphotyrosine antibody demonstrated that both carbachol and EGF induced tyrosyl phosphorylation of a major 52-kDa shc isoform. Grb2 association with Shc was demonstrated by blotting Grb2 immunoprecipitates with an anti-Shc antibody. Probing of anti-Sos immunoprecipitates with an anti-Grb2 antibody revealed that Sos was constitutively bound to Grb2. To examine the functional role of Sos in ERK2 activation, we transfected parietal cells with the HA-ERK2 vector together with a dominantly expressed mutant (inactive) sos gene. Dominant negative Sos did not affect carbachol stimulation of HA-ERK2 but inhibited the stimulatory effect of EGF by 60%. We then investigated the role of betagamma-subunits in carbachol induction of HA-ERK2. Parietal cells were transfected with the HA-ERK2 vector together with a vector expressing the carboxy terminus of the beta-adrenergic receptor kinase 1, known to block signaling mediated by betagamma-subunits. In the presence of this vector, carbachol induction of HA-ERK2 was inhibited by 40%. Together these data suggest that, in the gastric parietal cells, carbachol activates the ERKs through Ras- and betagamma-dependent mechanisms that require guanine nucleotide exchange factors other than Sos.

Adaptor Proteins, Signal Transducing↗

PubMed2000

Molecular physiology and pathophysiology of tight junctions. IV. Regulation of tight junctions by extracellular stimuli: nutrients, cytokines, and immune cells.

The epithelial lining of the gastrointestinal tract forms a regulated, selectively permeable barrier between luminal contents and the underlying tissue compartments. Permeability across the epithelium is, in part, determined by the rate-limiting barrier of the paracellular pathway-the most apical intercellular junction referred to as the tight junction (TJ). The TJ is composed of a multiprotein complex that affiliates with the underlying apical actomyosin ring. TJ structure and function, and therefore epithelial permeability, are influenced by diverse physiological and pathological stimuli; here we review examples of such stimuli that are detected at the cell surface. For example, luminal glucose induces an increase in paracellular permeability to small molecules. Similarly, but by other means, cytokines and leukocytes in the vicinity of the epithelium also regulate TJ structure and paracellular permeability by influencing the TJ protein complex and/or its association with the underlying actin cytoskeleton.

Cytokines↗

PubMed2006

L-type calcium channel alpha-subunit and protein kinase inhibitors modulate Rem-mediated regulation of current.

Cardiac voltage-gated L-type Ca channels (Ca(V)) are multiprotein complexes, including accessory subunits such as Ca(V)beta2 that increase current expression. Recently, members of the Rad and Gem/Kir-related family of small GTPases have been shown to decrease current, although the mechanism remains poorly defined. In this study, we evaluated the contribution of the L-type Ca channel alpha-subunit (Ca(V)1.2) to Ca(V)beta2-Rem inhibition of Ca channel current. Specifically, we addressed whether protein kinase A (PKA) modulation of the Ca channel modifies Ca(V)beta2-Rem inhibition of Ca channel current. We first tested the effect of Rem on Ca(V)1.2 in human embryonic kidney 293 (HEK-293) cells using the whole cell patch-clamp configuration. Rem coexpression with Ca(V)1.2 reduces Ba current expression under basal conditions, and Ca(V)beta2a coexpression enhances Rem block of Ca(V)1.2 current. Surprisingly, PKA inhibition by 133 nM H-89 or 50 microM Rp-cAMP-S partially relieved the Rem-mediated inhibition of current activity both with and without Ca(V)beta2a. To test whether the H-89 action was a consequence of the phosphorylation status of Ca(V)1.2, we examined Rem regulation of the PKA-insensitive Ca(V)1.2 serine 1928 (S1928) to alanine mutation (Ca(V)1.2-S1928A). Ca(V)1.2-S1928A current was not inhibited by Rem and when coexpression with Ca(V)beta2a was not completely blocked by Rem coexpression, suggesting that the phosphorylation of S1928 contributes to Rem-mediated Ca channel modulation. As a model for native Ca channel complexes, we tested the ability of Rem overexpression in HIT-T15 cells and embryonic ventricular myocytes to interfere with native current. We find that native current is also sensitive to Rem block and that H-89 pretreatment relieves the ability of Rem to regulate Ca current. We conclude that Rem is capable of regulating L-type current, that release of Rem block is modulated by cellular kinase pathways, and that the Ca(V)1.2 COOH terminus contributes to Rem-dependent channel inhibition.

Animals↗

PubMed2003

Protein kinase C modifications of VE-cadherin, p120, and beta-catenin contribute to endothelial barrier dysregulation induced by thrombin.

The adherens junction is a multiprotein complex consisting of the transmembrane vascular endothelial cadherin (VEC) and cytoplasmic catenins (p120, beta-catenin, plakoglobin, alpha-catenin) responsible for the maintenance of endothelial barrier function. Junctional disassembly and modifications in cadherin/catenin complex lead to increased paracellular permeability of the endothelial barrier. However, the mechanisms of junctional disassembly remain unclear. In this study, we used the proinflammatory mediator thrombin to compromise the barrier function and test the hypothesis that phosphorylation-induced alterations of VEC, beta-catenin, and p120 regulate junction disassembly and mediate the increased endothelial permeability response. The study showed that thrombin induced dephosphorylation of VEC, which is coupled to disassembly of cell-cell contacts, but VEC remained in aggregates at the plasma membrane. The cytoplasmic catenins dissociated from the VEC cytoplasmic domain in thin membrane projections formed in interendothelial gaps. We also showed that thrombin induced dephosphorylation of beta-catenin and phosphorylation of p120. Thrombin-induced interendothelial gap formation and increased endothelial permeability were blocked by protein kinase C inhibition using chelerythrine and Gö-6976 but not by LY-379196. Chelerythrine also prevented thrombin-induced phosphorylation changes of the cadherin/catenin complex. Thus the present study links posttranslational modifications of VEC, beta-catenin, and p120 to the mechanism of thrombin-induced increase in endothelial permeability.

Alkaloids↗

PubMed2001

Identification of a novel antigen on the apical surface of rat alveolar epithelial type II and Clara cells.

Here we describe a monoclonal antibody (MMC4) that recognizes a novel antigen on the apical surface of rat alveolar epithelial type II and Clara cells in the lung, proximal tubule epithelial cells in the kidney, and villus epithelial cells in the small intestine. Biochemical analysis showed that the MMC4 antigen was sensitive to heating and proteinase K digestion and that it is distributed in the detergent-rich phase after Triton X-114 phase separation. These data suggest that the MMC4 antigen is an integral membrane protein. Glycerol gradient sedimentation identified two forms of the MMC4 antigen: one with a sedimentation coefficient of 10.1 and one with a sedimentation coefficient of 1.66, suggesting that the antigen may be part of a multiprotein complex. During rat development (fetal day 16 to adult), the MMC4 antigen increased 12-fold in the lung and 200-fold in the kidney. In the intestine, the MMC4 antigen increased 150-fold by neonatal day 1 and then decreased to adult values. Our data demonstrate that the MMC4 antigen is unlike known type II cell- and Clara cell-associated proteins. The MMC4 monoclonal antibody will be useful as a marker of epithelial cell phenotype in development and injury studies.

Aging↗

PubMed1986

Protons, osmolytes, and fitness of internal milieu for protein function.

The composition of the intracellular milieu shows striking similarities among widely different species. Only certain values of intracellular pH, values that generally reflect alphastat regulation, and only narrow ranges of inorganic ion concentrations are found in the cytoplasm of the cells of most animals, plants, and microorganisms. In water-stressed organisms only a few types of low-molecular-weight organic molecules (osmolytes) are accumulated. These highly conserved characteristics of the intracellular fluids reflect the need to maintain critical features of macromolecules within narrow ranges optimal for life. For proteins these features include maintaining adequate rates of catalysis, a high level of regulatory responsiveness, and a precise balance between stability and lability of structure (tertiary conformation, subunit assembly, and multiprotein complexes). The optimal values for these functional and structural features of proteins often lie near the midrange of possible values for these properties, and only under specific conditions of intracellular pH, ionic strength, and osmolyte composition are these optimal midrange values conserved. In dormant cells the departure of solution conditions from values that are optimal for protein function and structure may be instrumental in reducing or shutting down metabolic functions. Seen from a broad evolutionary perspective, the evolution of the intracellular milieu is an important complement to macromolecular evolution. In certain instances appropriate modifications of the internal milieu may reduce the need for adaptive amino acid replacements in proteins.

Acid-Base Equilibrium↗

PubMed2001

NHERF: targeting and trafficking membrane proteins.

Vectorial ion transport initiated by Na+/H+ exchanger isoform 3 (NHE3) mediates the reabsorption of NaCl and NaHCO(3) in renal proximal tubule cells. NHE3 activity is modulated by numerous physiological stimuli. Biochemical and cellular experiments identified Na+/H+ exchanger regulatory factor (NHERF) as a protein cofactor essential for cAMP-mediated inhibition of NHE3 activity. Identification of numerous NHERF targets, including several transmembrane receptors and ion transporters, has broadened the role of this PSD-95/Dlg-1, Drososphila disk large/ZO-1 domain-containing adapter protein in membrane physiology. NHERF also associates with members of the ezrin/radixin/moesin family of actin-binding proteins and thus links NHE3 to the actin cytoskeleton. Formation of this multiprotein complex facilitates NHE3 phosphorylation and hormonal control of Na+/H+ exchange. NHERF also plays a critical role in targeting transport proteins to apical membranes. Moreover, the NHERF signaling complex functions as a regulatory unit to control endocytosis and internal trafficking of membrane proteins. This article reviews the new evidence that implicates NHERF in wider aspects of epithelial membrane biology.

Animals↗

PubMed2002

Playing Tag with HIF: The VHL Story.

Inactivation of the von Hippel-Lindau (VHL) tumour suppressor gene product pVHL is the cause of inherited VHL disease and is associated with sporadic kidney cancer. pVHL is found in a multiprotein complex with elongins B/C, Cul2, and Rbx1 forming an E3 ubiquitin ligase complex called VEC. This modular enzyme targets the alpha subunits of hypoxia-inducible factor (HIF) for ubiquitin-mediated destruction. Consequently, tumour cells lacking functional pVHL overproduce the products of HIF-target genes such as vascular endothelial growth factor (VEGF), which promotes angiogenesis. This likely accounts for the hypervascular nature of VHL-associated neoplasms. Although pVHL has been linked to the cell-cycle, differentiation, and the regulation of extracellular matrix assembly, microenvironment pH, and tissue invasiveness, this review will focus on the recent insights into the molecular mechanisms governing the E3 ubiquitin ligase function of VEC.

Journal Article↗

PubMed2005

Negative regulation of estrogen receptor alpha transactivation functions by LIM domain only 4 protein.

LIM domain only 4 (LMO4), a member of the LIM-only family of transcriptional coregulatory proteins, consists of two LIM protein-protein interaction domains that enable it to function as a linker protein in multiprotein complexes. Here, we have identified estrogen receptor alpha (ERalpha) and its corepressor, metastasis tumor antigen 1 (MTA1), as two novel binding partners of LMO4. Interestingly, LMO4 exhibited binding with both ERalpha and MTA1 and existed as a complex with ERalpha, MTA1, and histone deacetylases (HDAC), implying that LMO4 was a component of the MTA1 corepressor complex. Consistent with this notion, LMO4 overexpression repressed ERalpha transactivation functions in an HDAC-dependent manner. Accordingly, silencing of endogenous LMO4 expression resulted in a significant increased recruitment of ERalpha to target gene chromatin, stimulation of ERalpha transactivation activity, and enhanced expression of ERalpha-regulated genes. These findings suggested that LMO4 was an integral part of the molecular machinery involved in the negative regulation of ERalpha transactivation function in breast cells. Because LMO4 is up-regulated in human breast cancers, repression of ERalpha transactivation functions by LMO4 might contribute to the process of breast cancer progression by allowing the development of ERalpha-negative phenotypes, leading to increased aggressiveness of breast cancer cells.

Adaptor Proteins, Signal Transducing↗

PubMed2005

The -251T allele of the interleukin-8 promoter is associated with increased risk of gastric carcinoma featuring diffuse-type histopathology in Chinese population.

PURPOSE: Persistent interleukin-8 (IL-8) production contributes to chronic inflammation of the stomach. The proinflammatory IL-1beta polymorphisms, which enhance the cytokine production, are associated with increased risk of gastric cancer. The -251A/T polymorphism of the IL-8 promoter is involved in several human diseases. Particularly, the -251A is associated with decreased risk of colorectal cancer. We aimed to determine whether the -251 allele resulting in high IL-8 expression was associated with increased risk of gastric carcinoma. EXPERIMENTAL DESIGN: The -251A/T promoters were cloned and analyzed by luciferase assay. Binding of nuclear proteins to the -251A/T promoters was analyzed by electrophoretic mobility shift assay. The -251A/T promoters were differentiated by PCR-RFLP. Comparison of gastric cancer risk between the -251A/T promoters was done by a case-control study. RESULTS: The -251T allele possessed transcriptional activity 2- to 5-fold stronger than the -251A counterpart. Electrophoretic mobility shift assay showed that the -251A promoter had strong ability to bind to an unknown protein or multiprotein complex. The -251T allele was associated with increased risk of noncardia (P(trend) = 0.012) and cardia (P(trend) = 0.029) carcinomas. Gastric carcinoma patients with the low-risk AA genotype had a tendency to sustain intestinal-type carcinomas (chi(2) = 6.816; P = 0.033); however, the high-risk -251T allele was associated with >2-fold increased risk of diffuse-type (AA versus AT + TT: odds ratio, 2.52; 95% confidence interval, 1.16-5.49; P = 0.017) and mixed-type (AA versus AT + TT: odds ratio, 2.22; 95% confidence interval, 1.12-4.40; P = 0.019) carcinomas. CONCLUSIONS: The IL-8 -251T allele is significantly associated with increased risk of gastric carcinoma, particularly the diffuse and mixed types in Chinese population.

Adult↗

PubMed1999

Proteasome inhibitors induce p53/p21-independent apoptosis in human glioma cells.

The proteasome is a multiprotein complex involved in the degradation of ubiquitinated proteins. Three proteasome inhibitors, calpain inhibitor I, lactacystin and MG132, induced apoptosis in several human malignant glioma cell lines. Although proteasome inhibitors induced p53 accumulation in a cell line retaining wild-type p53 activity, p53 activity was dispensable for apoptosis since transdominant-negative p53 abrogated p53-dependent p21 induction but did not modulate apoptosis. Further, p21 was induced by higher concentrations of proteasome inhibitors in a p53-independent manner both in p53 wild-type and in p53 mutant cell lines. Although there was a strong G2/M arrest in response to proteasome inhibition in glioma cells, this G2/M arrest was also observed in p21(-/-) colon carcinoma cells, suggesting that p21 is dispensable for the G2/M arrest associated with proteasome inhibition. Interestingly, the p21(-/-) cells were more resistant to protease inhibitors than parental p21(+/+) cells. In summary, our data indicate that proteasome inhibition induces a p21-independent G2/M arrest and p53-independent apoptosis in human malignant glioma cells.

Acetylcysteine↗