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The Prohibitin Proteins: Doc

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Added on  2019-10-18

The Prohibitin Proteins: Doc

   Added on 2019-10-18

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Please do paraphrase and do not change in-text referenceThe prohibitinThe prohibitin proteins are a ubiquitously expressed pair of proteins, prohibitin 1 and prohibitin 2 [PHB2, also referred to as repressor of estrogen receptor activity (REA) or B-cell receptor associate protein (BAP)-37], belonging to the stomatin, prohibitin, flotillin, and HflK/C superfamily (Mishra, Murphy & Murphy, 2017). It was originally observed that transfection of PHB1 cDNA resulted in cell cycle arrest; hence the designation “prohibitin” (Mishra, Murphy & Murphy, 2006), ("Isolation of a cDNA that hybrid selects antiproliferative mRNA from rat liver - ScienceDirect", 2006). Both PHB1 and PHB2 have been shown to be present in the nucleus, mitochondria and cytosol, as well as associated with certain cell membrane receptors ("Prohibitin Ligands in Cell Death and Survival: Mode of Action and Therapeutic Potential - ScienceDirect", 1989). The PHBs are currently one of the best examples presenting clear and distinctive functions depending on intercellular localization. In the mitochondria, PHB1 and PHB2 form an alternating heterodimeric ring-like complex required for mitochondrial stability. In contrast, in the nucleus both PHBs result in the transcriptional suppression of target genes, but independently from one another (Mishra, Murphy & Murphy, 2006), ("Prohibitin Ligands in Cell Death and Survival: Mode of Action and Therapeutic Potential - ScienceDirect", 1989). Although nuclear PHBs have been demonstrated to influence multiple transcription factors and the cell cycle, the majority of cellular effects observed following the loss of either prohibitin can be attributed to their function in the mitochondria (Merkwirth et al., 2008).Expression of the PHBs is correlated to the level of reactive oxygen species (ROS) and inflammation; therefore, diseases with an inflammatory component (cancer, diabetes, and neuromuscular degenerative disorders) likely present with alterations in PHB1/2 expression and/or localization. In fact, experimental alteration of PHB expression in diverse model systems mimics several inflammatory pathologies (Merkwirth et al., 2012),(Supale et al., 2013),(Theiss et al., 2007),(Kasashima, Ohta, Kagawa & Endo, 2006). While most of the knowledge to date concerning the PHBs has been gleaned from studies with PHB1, this review focuses on PHB2 and the recent advances in the field that implicate the PHBs as intercellular communicators between the nucleus and mitochondria.(Bavelloni, Piazzi, Raffini, Faenza & Blalock, 2015).Figure 1: PHB1 and PHB2 gene structure and coding region and the resulting full-length protein. The gene structure including exons (boxes) and introns for PHB1 and PHB2 is shown. Shaded boxes indicated exon protein coding sequences while unshaded boxes indicate exon sequence that do not code for protein. The major domains of the resulting full-length PHB1 and PHB2 proteins and their approximate amino acid start and stop are indicated,(Bavelloni, Piazzi, Raffini, Faenza & Blalock, 2015).Figure 2: PHB2 post-translational modification sites. Post-translational modification sites (PTMs) obtained from the PhosphositePlus database are subdivided into serine/threonine phosphorylation, tyrosine phosphorylation, andlysine modifications (acetylation/ubiquitylation). Those sites in bold red indicate that biochemical evidence has validated the modification at this site. Sites in bold black indicate those sites in which significant and specific evidence exists (e.g. targeted mass spectrometry) that the site is modified, but no biochemical validation has been
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conducted. Sites in small black indicate sites which have been identified by high-throughput techniques in a non-specific manner and no biochemical evidence exists, (Bavelloni, Piazzi, Raffini, Faenza & Blalock, 2015).Figure 3: Signal transduction mediated by PHB2. (A) Diagram of the canonical role PHB2 maintains in mitochondrial protection and in regulating estrogen receptor (ER)-α induced transcription. (B) The transcriptional regulating role of PHB2 in myogenic differentiation and its regulation by AKT2 binding and CaMK IV-dependent phosphorylation at S91. (C) The transcriptional regulating and anti-apoptotic role of PHB2 in myeloid differentiation. Left panel, erythroid differentiation involving the PHB2 binding of the E3 ligase RNF2. Right panel, the role of the hierarchical AKT-dependent phosphorylation of nuclear PHB2 during all-trans retinoic acid mediated differentiation of promyelocytic leukemia cells, (Bavelloni, Piazzi, Raffini, Faenza & Blalock, 2015).Gene Structure and ExpressionThe phb2 gene, which was mapped to chromosome 12p13.31, covers 5.47 kb, encodes 10 exons, and is expressed in virtually all tested tissues (Fig. 1), (Bavelloni, Piazzi, Raffini, Faenza & Blalock, 2015). As a gene highly conserved through evolution, the human phb2 coding sequence maintains 91%, 69%, and 58% homology with phb2 in mouse, fruit fly, and yeast, respectively. While transcriptive regulation of phb2 is poorly understood, the phb2 promoter is predicted to contain binding sites for approximately 130 diversetranscription factors, including GATA-1,-2,-3, Fork head box protein (FoxO), CAAT-enhancer binding protein (C/EBP)-a, signal transducer and activator of transcription (STAT)-1, -3, and -5, peroxisome proliferating-activated receptor (PPAR)-a/-c, nuclear factor (NF)-jB, Sp1, and homeobox (HOX) factors (The Champion ChiP Transcription Factor Search Portal). Constitutive transcription factors such as Sp1 are likely responsible for the basal expression of phb2, and the presence of many of the other transcriptionfactors may explain why PHB2 expression is tightly linked to metabolic tissues and inflammation. Interestingly, transcription factors known to be negatively regulated by PHB2 (ERa, MyoD, and MEF-2) all have consensus binding sites within the phb2 promoter, suggesting a feedback loop involving PHB2 expression. Transcription of phb2 results in the expression of an ~1,505 bpmRNA, encoding a 299-aminoacid protein (Fig. 1), (Bavelloni, Piazzi, Raffini, Faenza & Blalock, 2015). In addition to this transcript, 13 splice variants have been documented; 6 of these do not result in protein expression, while 7 transcripts code for either identified or putative PHB2 isoforms (Bavelloni, Piazzi, Raffini, Faenza & Blalock, 2015).PHB2 Protein and Post-translational ModificationFull-length PHB2 has a molecular weight of 33.3 kDa with an amino acid sequence similarity between human and mouse,fruit fly, or yeast at 100%, 71%, and 56%, respectively (Mishra, Murphy & Murphy, 2006). While the amino acid sequence similarity between PHB1 and PHB2 is only 54%, the PHB domains are 74% identical (Mishra, Murphy & Murphy, 2006). Similar to PHB1, PHB2 contains a transmembrane domain (amino acids 1–36) required for mitochondrial localization, a central prohibitin domain (amino acids 36–201) followed by an overlapping coiled-coil domain (amino acids 188–264) (Fig. 1), (Bavelloni, Piazzi, Raffini, Faenza & Blalock, 2015). In addition to these, PHB2 also possesses an ER-binding domainwithin the prohibitin domain. In contrast to PHB1,which contains a putative nuclear exclusion sequence, PHB2 contains a putative nuclear import sequence (NIS) located within the ER-binding domain,
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suggesting that posttranslational modifications (PTMs) or association with other proteins dictates PHB2 sub-cellular localization (Mishra, Murphy & Murphy, 2006),(Theiss et al., 2011) (Fig. 1).A search of the PhosphoSite database reveals at least 32 potential post translational modification sites (ubiquitylation, acetylation, and phosphorylation); the majority of which have not been validated (Hornbecket al., 2011). Table 1 shows the most noted PTMs and kinase(s) that likely phosphorylate the respective site. Approximately 15 Ser/Thr phosphorylation sites have been detected in a variety of tissues and cell lines (Fig. 2), (Bavelloni, Piazzi, Raffini, Faenza & Blalock, 2015). Of these, only three (S91, S176, and S243) have been biochemically validated,while an additional three (S105, S151, and S286) are considered highly probable as they have been identified in a large number of independent studies (Bavelloni et al., 2014),(Zhou et al., 2013).Similarly, six potential Tyr sites have been identified, but with the exception of Y248, none of these sites has been validated biochemically (Ross, Nagy & Kirken, 2008),(Gu et al., 2006), (Knowlton et al., 2010) (Fig. 2). Moreover, modification of 11 Lys residues has been identified using high-throughput proteomics (Kim et al., 2011),(Wagner et al., 2011), (Weinert et al., 2013)(Fig. 2). Many of these result from ubiquitylation, while others result from acetylation. It is evident by comparing mouse and human PHB2 data that both Lys modifications can occur at these sites, but the consequences of these modifications remain unknown (Wagner et al., 2013).Of the biochemically validated phosphorylation sites in PHB2, the best studied is S91. Serine 91 was demonstrated to be phosphorylated by both Ca11/calmodulin-dependent kinase (CaMK) IV and AKT1/2 (Bavelloni et al., 2014. Sun et al. demonstrated that CaMK IV associates with PHB2 and phosphorylates S91 during myocytic differentiation of C2C12 myoblasts. The introduction of a non-phosphorylated S91A mutant in this model system resulted in the inability of CaMK IV to relieve PHB2-mediated repression of MEF2-dependent transcription and myocyte differentiation ("CaMK IV phosphorylates prohibitin 2 and regulates prohibitin 2-mediated repression of MEF2 transcription - ScienceDirect", 2011). More recently, Bavelloni et al. demonstrated that nuclear PHB2 is phosphorylated by AKT1/2 during alltrans retinoic acid (ATRA)-mediated differentiation of promyelocytic leukemia cells (Bavelloni et al., 2014). In addition to S91,S176 was also phosphorylated by AKT in this model system, although S176 phosphorylation was secondary to S91, suggesting a hierarchical phosphorylation. Interestingly, in this study, while exogenous expression of a S176A mutant had little effect on cell viability, exogenous expression of a S91A phospho-mutant resulted in a rapid and complete apoptosis of NB4 cells within 24 h after transfection (Bavelloni et al., 2014). Such apoptosis is a hallmark of mitochondria catastrophe and this phenotype matched those observed in embryonic stem cells in which the PHBs were knocked-out and cell lines where PHBs were knocked-down using siRNA (Merkwirth et al., 2008),(Kasashima, Ohta, Kagawa & Endo, 2006),(Ross, Nagy & Kirken, 2008),26,27) (Fig. 3),(Bavelloni, Piazzi, Raffini, Faenza & Blalock, 2015). Using human derived T-cells and T-cell lines, Ross et al. found that PHB1 and PHB2 expression and phosphorylation increased following T-cell activation. PHB2 phosphorylation was subsequently mapped toS243 and Y248 (Ross, Nagy & Kirken, 2008). While the authors did not follow-up on the effects of S243
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phosphorylation, they did demonstrate through the use of a Y248F PHB2 mutant, that phosphorylation of Y248 was not essential for cell survival or association with PHB1 (Ross, Nagy & Kirken, 2008).Protein-Protein Interactions and Complex FormationThe prohibitins have been identified in complex with proteins with diverse cellular functions. While some associations at the plasma membrane and in the mitochondria require both PHB1 and PHB2, the majority of protein-protein interactions are specific to each prohibitin(Bavelloni, Piazzi, Raffini, Faenza & Blalock, 2015).In the nucleus, many PHB2 interacting proteins are both global and specific transcription factors, including: the cAMPdependent transcription factor (ATF)-2, b-catenin, COUP-TF1, COUP-TF2, the estrogen receptor (ER)-a, interleukinenhancer binding factor (ILF)-3, MEF2A, MYOD1, Runt-related transcription factor (RUNX3), and transcription factor (TF)-E3 (Lau et al., 2012), (Ewing et al., 2007), (Sun,2004). Others are DNA modifying enzymes, including: the histone deacetylases (HDAC1, HDAC2, HDAC3, and HDAC5), breast-related carcinoma antigen (BRCA)-1, cyclin-dependent kinase (CDK)-2, polycomb-related proteins of the PRC2/EEDEZH2 complex, DNA repair associated enzymes, and cell cycle associated proteins (Ewing et al., 2007), (Sun, 2004), (Neganova et al., 2011),(Kim et al., 2009). Several RNA-binding proteins required for RNA processing (ATP-dependent RNA helicase DDX20), stability [Epiplakin (EPPK1) and the basophile leukemia expressed protein (Bles03)] and transport (Staufen) also associate with nuclear PHB2 (Ewing et al., 2007), (Milev, Ravichandran, Khan, Schriemer &Mouland, 2012),(Bavelloni, Piazzi, Raffini, Faenza & Blalock, 2015).In the cytosol, most PHB2 interacting proteins are associated with the cytoskeleton and cytoskeletal transport [coatomer subunit gamma 1 (COPG1)], cellular signaling [MDM2 and receptor interacting S/T kinase (RIPK)-2], and ubiquitylation (Kim et al., 2011), (Wagner et al., 2011), (Ewing et al., 2007), (Xu, Cai, Yang, Huang & Ye, 2012). Other cytoplasmic proteins that interact with PHB2 are associated with integral cell membrane proteins and cellular receptors, such as IGFR1 and integrins like VCAM1 (Humphries et al., 2009).As the PHBs also function in the mitochondria, many critical mitochondrial proteins associate with PHB2. These proteins consist of resident proteins belonging to the mitochondrial respiratory chain as well as mitochondrial transporters and membrane translocases (Richter-Dennerlein et al., 2014). Some associated proteins are involved in cristae formation and maintenance of mitochondrial structure, while others are involved in mitochondrial-mediated translation. Prominent mitochondrial apoptosis and autophagy regulating proteins have also been found associated with PHB2; among these are the apoptotic related proteins SCaMC-1 (SLC25A24) and growth hormoneinducible transmembrane protein (GHITM), and the autophagy promoting E3-ligase RNF185 (Richter-Dennerlein et al., 2014). As many of these PHB2: protein interactions are just coming to light, the functional outcome of PHB2 association with the majority of these proteins is still unclear(Bavelloni, Piazzi, Raffini, Faenza & Blalock, 2015).
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