scholarly journals The C-terminal Tail of Presenilin Regulates Omi/HtrA2 Protease Activity

2004 ◽  
Vol 279 (44) ◽  
pp. 45844-45854 ◽  
Author(s):  
Sanjeev Gupta ◽  
Rajesh Singh ◽  
Pinaki Datta ◽  
ZhiJia Zhang ◽  
Christopher Orr ◽  
...  
Keyword(s):  
Cell Death ◽  
Proteolytic Activity ◽  
Protease Activity ◽  
Molecular Mechanisms ◽  
Pdz Domain ◽  
Ectopic Expression ◽  
Amyloid Β ◽  
Peptide Ligand ◽  
Binding Motif ◽  
Dependent Manner

Presenilin mutations are responsible for most cases of autosomal dominant inherited forms of early onset Alzheimer disease. Presenilins play an important role in amyloid β-precursor processing, NOTCH receptor signaling, and apoptosis. However, the molecular mechanisms by which presenilins regulate apoptosis are not fully understood. Here, we report that presenilin-1 (PS1) regulates the proteolytic activity of the serine protease Omi/HtrA2 through direct interaction with its regulatory PDZ domain. We show that a peptide corresponding to the cytoplasmic C-terminal tail of PS1 dramatically increases the proteolytic activity of Omi/HtrA2 toward the inhibitor of apoptosis proteins and β-casein and induces cell death in an Omi/HtrA2-dependent manner. Consistent with these results, ectopic expression of full-length PS1, but not PS1 lacking the C-terminal PDZ binding motif, potentiated Omi/HtrA2-induced cell death. Our results suggest that the C terminus of PS1 is an activation peptide ligand for the PDZ domain of Omi/HtrA2 and may regulate the protease activity of Omi/HtrA2 after its release from the mitochondria during apoptosis. This mechanism of Omi/HtrA2 activation is similar to the mechanism of activation of the related bacterial DegS protease by the outer-membrane porins.

scholarly journals The SARS Coronavirus E Protein Interacts with PALS1 and Alters Tight Junction Formation and Epithelial Morphogenesis

2010 ◽  
Vol 21 (22) ◽  
pp. 3838-3852 ◽  
Author(s):  
Kim-Tat Teoh ◽  
Yu-Lam Siu ◽  
Wing-Lim Chan ◽  
Marc A. Schlüter ◽  
Chia-Jen Liu ◽  
...  
Keyword(s):  
Tight Junction ◽  
Tight Junctions ◽  
Mammalian Cells ◽  
Pdz Domain ◽  
Ectopic Expression ◽  
Binding Motif ◽  
Dependent Manner ◽  
Tight Junction Formation ◽  
Golgi Region ◽  
Carboxy Terminal

Intercellular tight junctions define epithelial apicobasal polarity and form a physical fence which protects underlying tissues from pathogen invasions. PALS1, a tight junction-associated protein, is a member of the CRUMBS3-PALS1-PATJ polarity complex, which is crucial for the establishment and maintenance of epithelial polarity in mammals. Here we report that the carboxy-terminal domain of the SARS-CoV E small envelope protein (E) binds to human PALS1. Using coimmunoprecipitation and pull-down assays, we show that E interacts with PALS1 in mammalian cells and further demonstrate that the last four carboxy-terminal amino acids of E form a novel PDZ-binding motif that binds to PALS1 PDZ domain. PALS1 redistributes to the ERGIC/Golgi region, where E accumulates, in SARS-CoV–infected Vero E6 cells. Ectopic expression of E in MDCKII epithelial cells significantly alters cyst morphogenesis and, furthermore, delays formation of tight junctions, affects polarity, and modifies the subcellular distribution of PALS1, in a PDZ-binding motif-dependent manner. We speculate that hijacking of PALS1 by SARS-CoV E plays a determinant role in the disruption of the lung epithelium in SARS patients.

scholarly journals Downregulation of miR-17-92 cluster by PERK fine-tunes unfolded protein response mediated apoptosis

2020 ◽  
Author(s):  
Danielle E. Read ◽  
Ananya Gupta ◽  
Karen Cawley ◽  
Laura Fontana ◽  
Patrizia Agostinis ◽  
...  
Keyword(s):  
Cell Death ◽  
Er Stress ◽  
Unfolded Protein Response ◽  
Molecular Mechanisms ◽  
Ectopic Expression ◽  
Proximal Region ◽  
Dependent Manner ◽  
Unfolded Protein ◽  
Promoter Deletion Analysis ◽  
Protein Response

AbstractAn important event in the unfolded protein response (UPR) is the activation of the endoplasmic reticulum kinase PERK (EIF2AK3). The PERK signalling branch first mediates a prosurvival response, which switches into a proapoptotic response upon prolonged ER stress. However, the molecular mechanisms of PERK-mediated cell death are not well understood. Here we show that expression of the primary miR-17-92 transcript and mature miRNAs belonging to miR-17-92 cluster is decreased during UPR. We found that activity of miR-17-92 promoter reporter was reduced during UPR in a PERK-dependent manner. We show that activity of miR-17-92 promoter is repressed by ectopic expression of ATF4 and NRF2. The promoter deletion analysis and ChIP assays mapped the region responding to UPR-mediated repression to site in the proximal region of the miR-17-92 promoter. Hypericin-mediated photo-oxidative ER damage reduced the expression of miRNAs belonging to miR-17-92 cluster in wild-type but not in PERK-deficient cells. Importantly, ER stress-induced apoptosis was inhibited upon miR-17-92 overexpression in SH-SY5Y and H9c2 cells. Our results reveal a novel function for NRF2, where repression of miR-17-92 cluster by NRF2 plays an important role in ER stress-mediated apoptosis. The data presented here provides mechanistic details how sustained PERK signalling via NRF2 mediated repression of miR-17-92 cluster can potentiate cell death.

scholarly journals Downregulation of miR-17-92 Cluster by PERK Fine-Tunes Unfolded Protein Response Mediated Apoptosis

Life ◽  
2021 ◽  
Vol 11 (1) ◽  
pp. 30
Author(s):  
Danielle E. Read ◽  
Ananya Gupta ◽  
Karen Cawley ◽  
Laura Fontana ◽  
Patrizia Agostinis ◽  
...  
Keyword(s):  
Cell Death ◽  
Er Stress ◽  
Unfolded Protein Response ◽  
Molecular Mechanisms ◽  
Ectopic Expression ◽  
Proximal Region ◽  
Dependent Manner ◽  
Unfolded Protein ◽  
Promoter Deletion Analysis ◽  
Protein Response

An important event in the unfolded protein response (UPR) is activation of the endoplasmic reticulum (ER) kinase PERK. The PERK signalling branch initially mediates a prosurvival response, which progresses to a proapoptotic response upon prolonged ER stress. However, the molecular mechanisms of PERK-mediated cell death are not well understood. Here we show that expression of the primary miR-17-92 transcript and mature miRNAs belonging to the miR-17-92 cluster are decreased during UPR. We found that miR-17-92 promoter reporter activity was reduced during UPR in a PERK-dependent manner. Furthermore, we show that activity of the miR-17-92 promoter is repressed by ectopic expression of ATF4 and NRF2. Promoter deletion analysis mapped the region responding to UPR-mediated repression to a site in the proximal region of the miR-17-92 promoter. Hypericin-mediated photo-oxidative ER damage reduced the expression of miRNAs belonging to the miR-17-92 cluster in wild-type but not in PERK-deficient cells. Importantly, ER stress-induced apoptosis was inhibited upon miR-17-92 overexpression in SH-SY5Y and H9c2 cells. Our results reveal a novel function for ATF4 and NRF2, where repression of the miR-17-92 cluster plays an important role in ER stress-mediated apoptosis. Mechanistic details are provided for the potentiation of cell death via sustained PERK signalling mediated repression of the miR-17-92 cluster.

scholarly journals MARCKS domain phosphorylation regulates the differential interaction of Diacylglycerol Kinase ζ with Rac1, RhoA and Syntrophin

2020 ◽  
Author(s):  
Ryan Ard ◽  
Jean-Christian Maillet ◽  
Elias Daher ◽  
Michael Phan ◽  
Radoslav Zinoviev ◽  
...  
Keyword(s):  
Catalytic Activity ◽  
Molecular Mechanisms ◽  
Rho Gtpase ◽  
Pdz Domain ◽  
Diacylglycerol Kinase ◽  
Binding Motif ◽  
Membrane Blebbing ◽  
Rhoa Activity ◽  
C Terminus ◽  
Mechanistic Basis

AbstractCells can switch between Rac1, lamellipodia-based and RhoA, blebbing-based migration modes but the molecular mechanisms regulating this choice are not fully understood. Diacylglycerol kinase ζ (DGKζ), which phosphorylates diacylglycerol to yield phosphatidic acid, forms independent complexes with Rac1 and RhoA, selectively dissociating each from RhoGDI. DGKζ catalytic activity is required for Rac1 dissociation but is dispensable for RhoA dissociation. Instead, DGKζ functions as a scaffold that stimulates RhoA release by enhancing RhoGDI phosphorylation by protein kinase Cα (PKCα). Here, PKCα-mediated phosphorylation of the DGKζ MARCKS domain increased DGKζ association with RhoA and decreased its interaction with Rac1. The same modification increased binding of the DGKζ C-terminus to the α1-syntrophin PDZ domain. Expression of a phosphomimetic DGKζ mutant stimulated membrane blebbing in mouse embryonic fibroblasts and C2C12 myoblasts, which was augmented by inhibition of endogenous Rac1. DGKζ expression in differentiated C2 myotubes, which have low endogenous Rac1 levels, also induced substantial membrane blebbing via the Rho-ROCK pathway. These events were independent of DGKζ catalytic activity, but dependent upon a functional C-terminal PDZ-binding motif. Rescue of RhoA activity in DGKζ-null cells required the PDZ-binding motif, suggesting syntrophin interaction is necessary for optimal RhoA activation. Collectively, our results define a switch-like mechanism involving DGKζ phosphorylation by PKCα that favours RhoA-driven blebbing over Rac1-driven lamellipodia formation and macropinocytosis. These findings provide a mechanistic basis for the effect of PKCα signaling on Rho GTPase activity and suggest PKCα activity plays a role in the interconversion between Rac1 and RhoA signaling that underlies different migration modes.

scholarly journals Human Papillomavirus Type 16 E6 Activates NF-κB, Induces cIAP-2 Expression, and Protects against Apoptosis in a PDZ Binding Motif-Dependent Manner

2006 ◽  
Vol 80 (11) ◽  
pp. 5301-5307 ◽  
Author(s):  
Michael A. James ◽  
John H. Lee ◽  
Aloysius J. Klingelhutz
Keyword(s):  
Human Papillomavirus ◽  
Epithelial Cells ◽  
Transcriptional Activation ◽  
Pdz Domain ◽  
Airway Epithelial Cells ◽  
Binding Motif ◽  
Dependent Manner ◽  
Airway Epithelial ◽  
Transcript Accumulation ◽  
Human Telomerase Reverse Transcriptase

ABSTRACT Infection with human papillomavirus (HPV) is a critical factor in the pathogenesis of most cervical cancers and some aerodigestive cancers. The HPV E6 oncoprotein from high-risk HPV types contributes to the immortalization and transformation of cells by multiple mechanisms, including degradation of p53, transcriptional activation of human telomerase reverse transcriptase (hTERT), and degradation of several proteins containing PDZ domains. The ability of E6 to bind PDZ domain-containing proteins is independent of p53 degradation or hTERT activation but does correlate with oncogenic potential (R. A. Watson, M. Thomas, L. Banks, and S. Roberts, J. Cell Sci. 116:4925-4934, 2003) and is essential for induction of epithelial hyperplasia in vivo (M. L. Nguyen, M. M. Nguyen, D. Lee, A. E. Griep, and P. F. Lambert, J. Virol. 77:6957-6964, 2003). In this study, we found that HPV type 16 E6 was able to activate NF-κB in airway epithelial cells through the induction of nuclear binding activity of p52-containing NF-κB complexes in a PDZ binding motif-dependent manner. Transcript accumulation for the NF-κB-responsive antiapoptotic gene encoding cIAP-2 and binding of nuclear factors to the proximal NF-κB binding site of the cIAP-2 gene promoter are induced by E6 expression. Furthermore, E6 is able to protect cells from TNF-induced apoptosis. All of these E6-dependent phenotypes are dependent on the presence of the PDZ binding motif of E6. Our results imply a role for targeting of PDZ proteins by E6 in NF-κB activation and protection from apoptosis in airway epithelial cells.

scholarly journals Kalirin, a Key Player in Synapse Formation, Is Implicated in Human Diseases

2012 ◽  
Vol 2012 ◽  
pp. 1-9 ◽  
Author(s):  
Prashant Mandela ◽  
Xin-Ming Ma
Keyword(s):  
Learning And Memory ◽  
Cortical Neurons ◽  
Molecular Mechanisms ◽  
Synapse Formation ◽  
Pdz Domain ◽  
Long Term Potentiation ◽  
Human Diseases ◽  
Binding Motif ◽  
Spine Density ◽  
Morphological Alterations

Synapse formation is considered to be crucial for learning and memory. Understanding the underlying molecular mechanisms of synapse formation is a key to understanding learning and memory. Kalirin-7, a major isoform of Kalirin in adult rodent brain, is an essential component of mature excitatory synapses. Kalirin-7 interacts with multiple PDZ-domain-containing proteins including PSD95, spinophilin, and GluR1 through its PDZ-binding motif. In cultured hippocampal/cortical neurons, overexpression of Kalirin-7 increases spine density and spine size whereas reduction of endogenous Kalirin-7 expression decreases synapse number, and spine density. In Kalirin-7 knockout mice, spine length, synapse number, and postsynaptic density (PSD) size are decreased in hippocampal CA1 pyramidal neurons; these morphological alterations are accompanied by a deficiency in long-term potentiation (LTP) and a decreased spontaneous excitatory postsynaptic current (sEPSC) frequency. Human Kalirin-7, also known as Duo or Huntingtin-associated protein-interacting protein (HAPIP), is equivalent to rat Kalirin-7. Recent studies show that Kalirin is relevant to many human diseases such as Huntington’s Disease, Alzheimer’s Disease, ischemic stroke, schizophrenia, depression, and cocaine addiction. This paper summarizes our recent understanding of Kalirin function.

scholarly journals Debcl, a Proapoptotic Bcl-2 Homologue, Is a Component of the Drosophila melanogaster Cell Death Machinery

2000 ◽  
Vol 148 (4) ◽  
pp. 703-714 ◽  
Author(s):  
Paul A. Colussi ◽  
Leonie M. Quinn ◽  
David C.S. Huang ◽  
Michelle Coombe ◽  
Stuart H. Read ◽  
...  
Keyword(s):  
Cell Death ◽  
Drosophila Melanogaster ◽  
Mammalian Cells ◽  
Cultured Cells ◽  
Gene Dosage ◽  
Family Members ◽  
Ectopic Expression ◽  
Dependent Manner ◽  
Proapoptotic Protein ◽  
Main Components

Bcl-2 family of proteins are key regulators of apoptosis. Both proapoptotic and antiapoptotic members of this family are found in mammalian cells, but no such proteins have been described in insects. Here, we report the identification and characterization of Debcl, the first Bcl-2 homologue in Drosophila melanogaster. Structurally, Debcl is similar to Bax-like proapoptotic Bcl-2 family members. Ectopic expression of Debcl in cultured cells and in transgenic flies causes apoptosis, which is inhibited by coexpression of the baculovirus caspase inhibitor P35, indicating that Debcl is a proapoptotic protein that functions in a caspase-dependent manner. debcl expression correlates with developmental cell death in specific Drosophila tissues. We also show that debcl genetically interacts with diap1 and dark, and that debcl-mediated apoptosis is not affected by gene dosage of rpr, hid, and grim. Biochemically, Debcl can interact with several mammalian and viral prosurvival Bcl-2 family members, but not with the proapoptotic members, suggesting that it may regulate apoptosis by antagonizing prosurvival Bcl-2 proteins. RNA interference studies indicate that Debcl is required for developmental apoptosis in Drosophila embryos. These results suggest that the main components of the mammalian apoptosis machinery are conserved in insects.

scholarly journals α-Synuclein binds and sequesters PIKE-L into Lewy bodies, triggering dopaminergic cell death via AMPK hyperactivation

2017 ◽  
Vol 114 (5) ◽  
pp. 1183-1188 ◽  
Author(s):  
Seong Su Kang ◽  
Zhentao Zhang ◽  
Xia Liu ◽  
Fredric P. Manfredsson ◽  
Li He ◽  
...  
Keyword(s):  
Cell Death ◽  
Molecular Mechanisms ◽  
Critical Role ◽  
Lewy Bodies ◽  
Cell Loss ◽  
Dominant Negative ◽  
Dependent Manner ◽  
Ph Domain ◽  
Dopaminergic Cell ◽  
Neuroprotective Actions

The abnormal aggregation of fibrillar α-synuclein in Lewy bodies plays a critical role in the pathogenesis of Parkinson’s disease. However, the molecular mechanisms regulating α-synuclein pathological effects are incompletely understood. Here we show that α-synuclein binds phosphoinositide-3 kinase enhancer L (PIKE-L) in a phosphorylation-dependent manner and sequesters it in Lewy bodies, leading to dopaminergic cell death via AMP-activated protein kinase (AMPK) hyperactivation. α-Synuclein interacts with PIKE-L, an AMPK inhibitory binding partner, and this action is increased by S129 phosphorylation through AMPK and is decreased by Y125 phosphorylation via Src family kinase Fyn. A pleckstrin homology (PH) domain in PIKE-L directly binds α-synuclein and antagonizes its aggregation. Accordingly, PIKE-L overexpression decreases dopaminergic cell death elicited by 1-methyl-4-phenylpyridinium (MPP+), whereas PIKE-L knockdown elevates α-synuclein oligomerization and cell death. The overexpression of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) or α-synuclein induces greater dopaminergic cell loss and more severe motor defects in PIKE-KO and Fyn-KO mice than in wild-type mice, and these effects are attenuated by the expression of dominant-negative AMPK. Hence, our findings demonstrate that α-synuclein neutralizes PIKE-L’s neuroprotective actions in synucleinopathies, triggering dopaminergic neuronal death by hyperactivating AMPK.

scholarly journals The Serine Protease HhoA from Synechocystis sp. Strain PCC 6803: Substrate Specificity and Formation of a Hexameric Complex Are Regulated by the PDZ Domain

2007 ◽  
Vol 189 (18) ◽  
pp. 6611-6618 ◽  
Author(s):  
Pitter F. Huesgen ◽  
Philipp Scholz ◽  
Iwona Adamska
Keyword(s):  
Membrane Proteins ◽  
Substrate Specificity ◽  
Proteolytic Activity ◽  
Protease Activity ◽  
Elevated Temperatures ◽  
Critical Role ◽  
Pdz Domain ◽  
General Role ◽  
Pcc 6803

ABSTRACT Enzymes of the ATP-independent Deg serine endopeptidase family are very flexible with regard to their substrate specificity. Some family members cleave only one substrate, while others act as general proteases on unfolded substrates. The proteolytic activity of Deg proteases is regulated by PDZ protein interaction domains. Here we characterized the HhoA protease from Synechocystis sp. strain PCC 6803 in vitro using several recombinant protein constructs. The proteolytic activity of HhoA was found to increase with temperature and basic pH and was stimulated by the addition of Mg2+ or Ca2+. We found that the single PDZ domain of HhoA played a critical role in regulating protease activity and in the assembly of a hexameric complex. Deletion of the PDZ domain strongly reduced proteolysis of a sterically challenging resorufin-labeled casein substrate, but unlabeled β-casein was still degraded. Reconstitution of the purified HhoA with total membrane proteins isolated from Synechocystis sp. wild-type strain PCC 6803 and a ΔhhoA mutant resulted in specific degradation of selected proteins at elevated temperatures. We concluded that a single PDZ domain of HhoA plays a critical role in defining the protease activity and oligomerization state, combining the functions that are attributed to two PDZ domains in the homologous DegP protease from Escherichia coli. Based on this first enzymatic study of a Deg protease from cyanobacteria, we propose a general role for HhoA in the quality control of extracytoplasmic proteins, including membrane proteins, in Synechocystis sp. strain PCC 6803.

Ectopic expression of cyclin D1 prevents activation of gene transcription by myogenic basic helix-loop-helix regulators

1994 ◽  
Vol 14 (8) ◽  
pp. 5259-5267
Author(s):  
S S Rao ◽  
C Chu ◽  
D S Kohtz
Keyword(s):  
Cell Cycle ◽  
Cyclin D1 ◽  
Gene Transcription ◽  
Transcriptional Activation ◽  
Ectopic Expression ◽  
Binding Motif ◽  
Dependent Manner ◽  
Basic Helix Loop Helix ◽  
Helix Loop Helix ◽  
Muscle Gene

Activation of muscle gene transcription in differentiating skeletal myoblasts requires their withdrawal from the cell cycle. The effects of ectopic cyclin expression on activation of muscle gene transcription by myogenic basic helix-loop-helix (bHLH) regulators were investigated. Ectopic expression of cyclin D1, but not cyclins A, B1, B2, C, D3, and E, inhibited transcriptional activation of muscle gene reporter constructs by myogenic bHLH regulators in a dose-dependent manner. Ectopic expression of cyclin D1 inhibited the activity of a myogenic bHLH regulator mutant lacking the basic region protein kinase C site, indicating that phosphorylation of this site is not relevant to the mechanism of inhibition. Analysis of cyclin D1 mutants revealed that the C-terminal acidic region was required for inhibition of myogenic bHLH regulator activity, whereas an intact N-terminal pRb binding motif was not essential. Together, these results implicate expression of cyclin D1 as a central determinant of a putatively novel mechanism that links positive control of cell cycle progression to negative regulation of genes expressed in differentiated myocytes.

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