User:Ethan Kitt/Sandbox 1
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| - | =Human Poly(A) Binding Protein | + | ==Discussion of Human Poly(A) Binding Protein== |
== Background == | == Background == | ||
The Human Poly(A) Binding Protein [https://www.rcsb.org/structure/1cvj (PABP)] was discovered in 1973 by the use of a sedimentation profile detailing the RNase digestion differentiated the PABP protein. <ref name="A Protein of Molecular Weight 78,000">Blobel, Gunter. “A Protein of Molecular Weight 78,000 Bound to the Polyadenylate Region of Eukaryotic Messenger Rnas.” Proceedings of the National Academy of Sciences of the United States of America, vol. 70, no. 3, 1973, pp. 924–8.</ref> Attempts to purify the 75 kDa protein then followed. In 1983, then considered “poly(A)-organizing protein,” was determined and purified by molecular weight, ligand-binding affinity, and amounts found in cytoplasmic portions of cell with ability to bind to free poly(A). <ref name="Cytoplasmic Poly(A)">Baer, Bradford W. and Kornberg, Roger D. "The Protein Responsible for the Repeating Structure of Cytoplasmic Poly(A)-Ribonucleoprotein." The Journal of Cell Biology, vol. 96, no. 3, Mar. 1983, pp. 717-721. EBSCOhost. </ref> | The Human Poly(A) Binding Protein [https://www.rcsb.org/structure/1cvj (PABP)] was discovered in 1973 by the use of a sedimentation profile detailing the RNase digestion differentiated the PABP protein. <ref name="A Protein of Molecular Weight 78,000">Blobel, Gunter. “A Protein of Molecular Weight 78,000 Bound to the Polyadenylate Region of Eukaryotic Messenger Rnas.” Proceedings of the National Academy of Sciences of the United States of America, vol. 70, no. 3, 1973, pp. 924–8.</ref> Attempts to purify the 75 kDa protein then followed. In 1983, then considered “poly(A)-organizing protein,” was determined and purified by molecular weight, ligand-binding affinity, and amounts found in cytoplasmic portions of cell with ability to bind to free poly(A). <ref name="Cytoplasmic Poly(A)">Baer, Bradford W. and Kornberg, Roger D. "The Protein Responsible for the Repeating Structure of Cytoplasmic Poly(A)-Ribonucleoprotein." The Journal of Cell Biology, vol. 96, no. 3, Mar. 1983, pp. 717-721. EBSCOhost. </ref> | ||
| + | |||
| + | PABP is a mRNA binding protein that binds to the 3’ Poly(A) tail on mRNA. It is comprised of four [https://en.wikipedia.org/wiki/RNA_recognition_motif RNA recognition motifs] (RRMs), which are highly conserved RNA-binding domains.<ref name="Recognition of Polyadenylate RNA by the Poly(A)-Binding Protein">Deo, Rahul C, et al. “Recognition of Polyadenylate RNA by the Poly(A)-Binding Protein.” Cell 98:6. (1999) 835-845. Print. </ref> The RRM in PABP is found in over two hundred families of proteins across species, indicating that it is ancient.<ref name="Recognition of Polyadenylate RNA by the Poly(A)-Binding Protein">Deo, Rahul C, et al. “Recognition of Polyadenylate RNA by the Poly(A)-Binding Protein.” Cell 98:6. (1999) 835-845. Print. </ref> Through extensive Adenosine recognition by the RRMs of PABP, the protein is involved in three main functions: recognition of the 3’ Poly(A) tail, mRNA stabilization, and eukaryotic translation initiation. The contributions of controlling gene expression via different families of PABPs is not yet fully understood. PABP families are divided into nuclear and cytoplasmic. <ref name="Roles of Cytoplasmic Poly(A)-Binding Proteins">Gorgoni, Barbra, and Gray, Nicola. “The Roles of Cytoplasmic Poly(A)-Binding Proteins in Regulating Gene Expression: A Developmental Perspective.” Briefings in Functional Genomics and Proteomics, vol. 3, no. 2, 1 Aug. 2004, pp. 125–141., doi:10.1093/bfgp/3.2.125.</ref> PABP1, which is predominantly cytoplasmic, is often referred to as PABP because it is the only form of PABP that has been extensively studied in its role with mRNA translation and stability. <ref name="Roles of Cytoplasmic Poly(A)-Binding Proteins">Gorgoni, Barbra, and Gray, Nicola. “The Roles of Cytoplasmic Poly(A)-Binding Proteins in Regulating Gene Expression: A Developmental Perspective.” Briefings in Functional Genomics and Proteomics, vol. 3, no. 2, 1 Aug. 2004, pp. 125–141., doi:10.1093/bfgp/3.2.125.</ref> | ||
| - | ==Structure== | ||
| - | <StructureSection load='1B7F' size='350' frame='true' side='right' caption='PABP' scene='78/782614/Structure_scene_color_scheme/1'> The crystal structure PABP was derived from X-ray Diffraction at 2.6Å (R-value: 23%). The subunits of PABP, RRM1 and RRM2, are examined in this article as the ''in vivo'' form seen in biological assembly 1 (via PDB). The protein has a homopolymeric structure, containing four RNA recognition motifs (RRMs), which are conserved. <ref name="Structure and Function">Kühn, Uwe and Elmar, Wahle. “Structure and Function of Poly(a) Binding Proteins.” Bba - Gene Structure & Expression, vol. 1678, no. 2/3, 2004. </ref> <scene name='78/782616/Rrm1_only/2'>RRM1</scene> and <scene name='78/782616/Rrm2_only/2'>RRM2</scene> are N-terminal domains that are connected by a <scene name='78/782616/Linker/3'>linker</scene>.<ref name="Recognition of Polyadenylate RNA by the Poly(A)-Binding Protein">Deo, Rahul C, et al. “Recognition of Polyadenylate RNA by the Poly(A)-Binding Protein.” Cell 98:6. (1999) 835-845. Print. </ref> Opposed to their counterparts, RRM3 and RRM4 bind Poly (A) RNA less tightly than RRM1 and RRM2.<ref name="Recognition of Polyadenylate RNA by the Poly(A)-Binding Protein">Deo, Rahul C, et al. “Recognition of Polyadenylate RNA by the Poly(A)-Binding Protein.” Cell 98:6. (1999) 835-845. Print. </ref> | ||
| - | ===RRMs=== | ||
| - | [[Image:ElectroPOSITIVE.png|250 px|left|thumb|Figure 2. Electropositive binding cleft. Electropositive region shown in blue and electronegative region shown in red.]] The sex-lethal protein (Sxl) is 354 amino acid residues long. It is composed of two highly conserved regions called <scene name='78/782600/Rbd1_and_rbd2/4'>RNA recognition motifs</scene> (RRMs) that function as a monomeric unit. Each RRM is approximately 90 amino acids long with a four stranded sheet <scene name='78/782600/Opening_image/3'>β-pleated</scene> and two <scene name='78/782600/Alpha_helices/1'>α-helices</scene>. The β-pleated sheets from each RRM interact with the RNA ligand, while the α-helices interact with each other to shape the protein <ref name="Clery"/>. The RRMs interact at <scene name='78/782600/Inter-domain_interactions/5'>two places</scene>: between the side chain of Lys 197 and the main chain carbonyl of Val 238 and between the side chains of Tyr 131 and Gln 239<ref name="Handa" />. The two RRMs are connected via an interdomain linker. The linker often forms a short 3<sub>10</sub> helix from Gly205 to Thr211.The interaction between the RRMs and the ligand is facilitated by the v-shaped <scene name='78/782600/Binding_cleft/7'>binding cleft</scene> formed by the β-pleated sheet of each RRM. The v-shaped cleft is strongly electropositive which also assists in ligand binding <ref name="Handa" /> (Figure 2). The presence of two RRMs increases RNA binding specificity by allowing for an elongated and continuous binding site <ref name="Clery">DOI 10.1016/j.sbi.2008.04.002</ref>. | ||
| - | === The Ligand=== | ||
| - | [[Image:Stacking.png|200 px|right|thumb|Figure 3: Intramolecular ring stacking of U7 (top) and U8 (bottom)]] The RNA <scene name='78/782599/Ligand_only/4'>ligand</scene> bound by Sxl is 9 nucleotides long—UGUUUUUUU. This ligand lacks intramolecular base pairs—a characteristic that would typically assist in RNA recognition—and therefore presents with many unique features. Sxl fixes U3-U11 in a specific elongated conformation. <scene name='78/782599/U6-11/4'>U6-U11</scene> interact within the strongly electropositive v-shaped cleft, while <scene name='78/782600/U3-u5/7'>U3-G4-U5</scene> are bound to a positively charged surface on RRM2 <ref name="Handa" />. The kink in the middle of the ligand is created by hydrogen bonds between the 2’OH's of U5 and U6 and the phosphate group of U8, as well as between the 2’OH of U7 and the phosphate group of U5. This is kink is also facilitated by the only intramolecular stacking pair in the ligand, U7 and U8 (Figure 3). All of the nucleotides besides U8 are in the [http://x3dna.org/highlights/sugar-pucker-correlates-with-phosphorus-base-distance C2’-endo] conformation. This orientation allows the bases to be highly exposed to the protein and therefore increases specificity. The low number of intramolecular stacking regions and the large number of C2’-endo conformations deem this ligand unique<ref name="Handa" />. | ||
| - | === | + | == Structure == |
| - | 1. | + | <StructureSection load='1cvj' size='340' side='right' caption='PABP' scene='78/782614/Structure_scene_color_scheme/1'> __NoTOC__ |
| + | The crystal structure PABP was derived from X-ray Diffraction at 2.6Å (R-value: 23%). The subunits of PABP, RRM1 and RRM2, are examined in this article as the ''in vivo'' form seen in biological assembly 1 (via PDB). The protein has a homopolymeric structure, containing four RNA recognition motifs (RRMs), which are conserved. <ref name="Structure and Function">Kühn, Uwe and Elmar, Wahle. “Structure and Function of Poly(a) Binding Proteins.” Bba - Gene Structure & Expression, vol. 1678, no. 2/3, 2004. </ref> <scene name='78/782616/Rrm1_only/3'>RRM1</scene> and <scene name='78/782616/Rrm2_only/3'>RRM2</scene> are N-terminal domains that are connected by a <scene name='78/782616/Linker/4'>linker</scene>.<ref name="Recognition of Polyadenylate RNA by the Poly(A)-Binding Protein">Deo, Rahul C, et al. “Recognition of Polyadenylate RNA by the Poly(A)-Binding Protein.” Cell 98:6. (1999) 835-845. Print. </ref> Opposed to their counterparts, RRM3 and RRM4 bind Poly (A) RNA less tightly than RRM1 and RRM2.<ref name="Recognition of Polyadenylate RNA by the Poly(A)-Binding Protein">Deo, Rahul C, et al. “Recognition of Polyadenylate RNA by the Poly(A)-Binding Protein.” Cell 98:6. (1999) 835-845. Print. </ref> | ||
| - | + | ===RNA Recognition Motifs (RRMs)=== | |
| + | The <scene name='78/782616/Subunits_of_pabp/4'>components of PABP</scene> are categorized into two RRMs: the n-terminus RRM1 (red) and c-terminus RRM2 (blue) are shown accordingly. The two RRMs are linked via an alpha-helix linker (green) that maintains the RRM1/2 complex that is the biological assembly and active form of PABP. Each RRM has a four-stranded antiparallel beta sheet backed by two corresponding alpha helices. <ref name="Recognition of Polyadenylate RNA by the Poly(A)-Binding Protein">Deo, Rahul C, et al. “Recognition of Polyadenylate RNA by the Poly(A)-Binding Protein.” Cell 98:6. (1999) 835-845. Print. </ref> mRNA poly-adenosine recognition is due to the presence of the conserved residues within the beta-sheet surface <ref name="The Poly(A)-Binding Protein and an mRNA Stability Protein Jointly Regulate an Endoribonuclease Activity.">Wang, Zuoren and Kiledjian, Megerditch. “The Poly(A)-Binding Protein and an mRNA Stability Protein Jointly Regulate an Endoribonuclease Activity.” Molecular and Cellular Biology 20.17 (2000): 6334–6341. Print.</ref> , which forms a <scene name='78/782616/Trough2/3'>trough</scene>-like pocket for the mRNA to bind. The beta-sheet flooring present in PABP interacts with the 3’ mRNA tail via a combination of van der Waals, aromatic stacking, and Hydrogen bonding. Through these interactions, PABP binds to 3’ Poly (A) tail with a KD of 2-7 nM. <ref name="Roles of Cytoplasmic Poly(A)-Binding Proteins">Gorgoni, Barbra, and Gray, Nicola. “The Roles of Cytoplasmic Poly(A)-Binding Proteins in Regulating Gene Expression: A Developmental Perspective.” Briefings in Functional Genomics and Proteomics, vol. 3, no. 2, 1 Aug. 2004, pp. 125–141., doi:10.1093/bfgp/3.2.125.</ref> [[Image:Hydrophobicity (1).png|200px|right|thumb| "Figure 1:" Surface hydrophobicity shown in presence of mRNA]] | ||
| - | 2. Hydrogen bonding with the RNA backbone: | ||
| - | + | Further, the RRM1/2 complex interacts with the mRNA's sugar-phosphate backbone, where 4 of the 8 mRNA adenosines interact electrostatically.<ref name="Recognition of Polyadenylate RNA by the Poly(A)-Binding Protein">Deo, Rahul C, et al. “Recognition of Polyadenylate RNA by the Poly(A)-Binding Protein.” Cell 98:6. (1999) 835-845. Print. </ref> Upon closer examination of the PABP structure, the protein contains loop-like domains that form the walls of the beta-sheet trough. Although these <scene name='78/782616/Walls_of_trough/4'>loop walls</scene> are present, no interaction occurs between the mRNA and these regions. We propose that these loops only keep unwanted cellular elements out of the binding pocket via hydrophobic and hydrophilic interactions, maintaining the protein's selectivity for mRNA (Figure 1). The structural elements highlighted consist of the RRM1/2 subunits, the linker domain, and the Poly(A) mRNA binding trough. | |
| - | 3. Intermolecular stacking: | ||
| - | [https://en.wikipedia.org/wiki/Stacking_(chemistry) Intermolecular stacking] between the aromatic side chains and the nucleotide bases also contributes to RNA binding. In RRM2, U3-G4-U5 stack with <scene name='78/782600/Rrm2_residues_stacking/3'>V254, Y214, and F256</scene> respectively. In RRM1, residues <scene name='78/782600/Rrm1_intermolecular_stacking/3'>Y131, R195, N130, F170</scene> are involved in favorable intermolecular stacking of aromatic rings with nucleotides U6-U11<ref name="Handa" />. | ||
| + | ==Interactions== | ||
| + | |||
| + | ===Adenosine Recognition Interactions (table left) and mRNA Stabilization via Aromatic Stacking (table right)=== | ||
| + | <table align='right'><tr><td colspan='2'> | ||
| + | <tr id='Adenosine Number'><td class="sblockLbl"><b>Adenosine Number</b></td><td class="sblockDat">PABP residue</td><tr> | ||
| + | <tr id='A3'><td class="sblockLbl"><b>A3</b></td><td class="sblockDat"><scene name='78/782616/A3-phe102_fd/1'>Phe102</scene></td><tr> | ||
| + | <tr id='A6'><td class="sblockLbl"><b>A6</b></td><td class="sblockDat"><scene name='78/782616/A6-tyr14/1'>Tyr14</scene></td></tr> | ||
| + | <tr id='A8'><td class="sblockLbl"><b>A8</b></td><td class="sblockDat"><scene name='78/782616/Tyr56-a8/1'>Tyr56</scene></td></tr> | ||
| + | </table> | ||
| + | |||
| + | <table><tr><td colspan='2'> | ||
| + | <tr id='Nucleotide'><td class="sblockLbl"><b>Nucleotide</b></td><td class="sblockDat">PABP residue</td><td class="sblockDat">Atom on PABP residue</td><td class="sblockDat">Atom on RNA</td> | ||
| + | <tr id='A2'><td class="sblockLbl"><b>A2</b></td><td class="sblockDat"><scene name='78/782616/Asn105_a2/2'>Asn105</scene></td><td class="sblockDat">side chain amine</td><td class="sblockDat">N6</td> | ||
| + | <tr id='A3'><td class="sblockLbl"><b>A3</b></td><td class="sblockDat"><scene name='78/782616/Lys174-a3/2'>Lys174</scene></td><td class="sblockDat">amine</td><td class="sblockDat">N3</td> | ||
| + | <tr id='A4'><td class="sblockLbl"><b>A4</b></td><td class="sblockDat"><scene name='78/782616/Asn100-a4/2'>Asn100</scene></td><td class="sblockDat">side chain amine and carbonyl</td><td class="sblockDat">N7</td> | ||
| + | <tr id='A4'><td class="sblockLbl"><b>A4</b></td><td class="sblockDat"><scene name='78/782616/Ser127-a4/2'>Ser127</scene></td><td class="sblockDat">side chain and N-term amine</td><td class="sblockDat">N1</td> | ||
| + | <tr id='A5'><td class="sblockLbl"><b>A5</b></td><td class="sblockDat"><scene name='78/782616/His144-a5/2'>His144</scene></td><td class="sblockDat">pyrimidine π-amine </td><td class="sblockDat">N6</td> | ||
| + | <tr id='A6'><td class="sblockLbl"><b>A6</b></td><td class="sblockDat"><scene name='78/782616/Gln88-a6/2'>Gln88</scene></td><td class="sblockDat">N-term amine and C-term OH</td><td class="sblockDat">N1</td> | ||
| + | <tr id='A6'><td class="sblockLbl"><b>A6</b></td><td class="sblockDat"><scene name='78/782616/Trp86-a6/2'>Trp86</scene></td><td class="sblockDat">N-term amine</td><td class="sblockDat">N6</td> | ||
| + | <tr id='A6'><td class="sblockLbl"><b>A6</b></td><td class="sblockDat"><scene name='78/782616/A6-tyr14_fd/1'>Tyr14</scene></td><td class="sblockDat">phenol OH</td><td class="sblockDat">phosphate OH</td> | ||
| + | <tr id='A7'><td class="sblockLbl"><b>A7</b></td><td class="sblockDat"><scene name='78/782616/Tyr54-ribose_a7/1'>Trp54</scene></td><td class="sblockDat">side chain OH</td><td class="sblockDat">ribose 2 OH</td> | ||
| + | <tr id='A7'><td class="sblockLbl"><b>A7</b></td><td class="sblockDat"><scene name='78/782616/Asp45-a7/2'>Asn45</scene></td><td class="sblockDat">side chain OH</td><td class="sblockDat">N6</td> | ||
| + | <tr id='A7'><td class="sblockLbl"><b>A7</b></td><td class="sblockDat"><scene name='78/782616/Met46-a7/2'>Met46</scene></td><td class="sblockDat">C-term carbonyl</td><td class="sblockDat">N1</td> | ||
| + | <tr id='A8'><td class="sblockLbl"><b>A8</b></td><td class="sblockDat"><scene name='78/782616/Tyr54-a8/2'>Tyr54</scene></td><td class="sblockDat">side chain OH</td><td class="sblockDat">phosphate OH</td> | ||
| + | <tr id='A8'><td class="sblockLbl"><b>A8</b></td><td class="sblockDat"><scene name='78/782616/Tyr56-a8_fd/2'>Tyr56</scene></td><td class="sblockDat">side chain OH</td><td class="sblockDat">phosphate OH</td> | ||
| + | <tr id='A9'><td class="sblockLbl"><b>A9</b></td><td class="sblockDat"><scene name='78/782616/Arg44-a9/2'>Arg44</scene></td><td class="sblockDat">side chain τ-amine</td><td class="sblockDat">N1</td> | ||
| + | </table> | ||
| + | |||
| + | |||
| + | |||
| + | == Function == | ||
| + | In eukaryotic mRNA translation, PABP recognizes the 3' Poly(A) tail via trough interactions determined above. While associated with the Poly(A) region, the complex then works together to stabilize the mRNA by preventing exoribonucleolytic degradation,<ref name="Recognition of Polyadenylate RNA by the Poly(A)-Binding Protein">Deo, Rahul C, et al. “Recognition of Polyadenylate RNA by the Poly(A)-Binding Protein.” Cell 98:6. (1999) 835-845. Print. </ref> thereby guiding the mRNA molecule into the translation pathway via interactions with translation initiation factor gG. | ||
| + | |||
| + | ===Recognition of the Poly(A) Tail=== | ||
| + | Polyadenylation of an mRNA involves the recognition of the 5’-AAUAAA-3’ consensus site, the cleavage downstream of the consensus site, and then the addition of adenines by [https://en.wikipedia.org/wiki/Polynucleotide_adenylyltransferase Poly(A) Polymerase] to the 3’ end. The newly added poly(A) tail is associated with the PABP. PABP requires 11-12 adenosines in order to bind. PABP and the bound Poly(A) tail work together to stabilize mRNA by preventing exo-ribonucleolytic degradation,<ref name="Recognition of Polyadenylate RNA by the Poly(A)-Binding Protein">Deo, Rahul C, et al. “Recognition of Polyadenylate RNA by the Poly(A)-Binding Protein.” Cell 98:6. (1999) 835-845. Print. </ref> thereby guiding the mRNA molecule into the translation pathway. Upon mRNA poly(A) recognition, PABP and the bound mRNA stimulate the initiation of translation by interacting with initiation factor [https://en.wikipedia.org/wiki/EIF4G eIF4G]. | ||
| + | |||
| + | ===mRNA Stabilization=== | ||
| + | PABP prevents the deadenylation and decapping of the mRNA, serving as a source of stabilization. Poly(A) ribonuclease [https://en.wikipedia.org/wiki/Poly(A)-specific_ribonuclease (PARN)] work to deadenylate mRNA, but the presence of PABP prevents its activity. The PABP protein is able to protect mRNA degradation through the complex that it forms with the elongation initiation factors, which prevent deadenylation and decapping due to their presence.<ref name="Recognition of Polyadenylate RNA by the Poly(A)-Binding Protein">Deo, Rahul C, et al. “Recognition of Polyadenylate RNA by the Poly(A)-Binding Protein.” Cell 98:6. (1999) 835-845. Print. </ref> This has been verified by the presence of deadenylation products and the comparative size of PABP footprints. There is some evidence indicating that PABP is involved in the prevention of endonucleolytic cleavage; however, only a small amount of mRNA is degraded from endonucleolytic cleavage, so it is not widely researched.<ref name="Recognition of Polyadenylate RNA by the Poly(A)-Binding Protein">Deo, Rahul C, et al. “Recognition of Polyadenylate RNA by the Poly(A)-Binding Protein.” Cell 98:6. (1999) 835-845. Print. </ref> | ||
| + | |||
| + | |||
| + | ===Eukaryotic Translation Initiation=== | ||
| + | Upon mRNA Poly(A) recognition, PABP and the bound mRNA stimulate the initiation of translation by interacting with initiation factor eIF4G. Protein eIF4G actually interacts with PABP's dorsal side (Figure2) (under the trough) hydrophobic and acidic residues that stimulate the interaction between the two proteins. These specific residues are phylogenetically conserved among all PABPs, and therefore significant in the protein's function and interaction with eIF4G. [[Image:Dorsal side.jpg|200px|right|thumb| "Figure 2:"Dorsal side with green conserved residues that interact with eIF4.]] | ||
| + | |||
| + | PABP and mRNA complex aids in translation initiation under two proposed mechanisms. Within the two mechanisms, studies have highlighted the presence The “Closed Loop” Model entails the recognition of the 5’ 7-methyl-Guanosine cap by [https://en.wikipedia.org/wiki/Eukaryotic_initiation_factor_4F eIF4F], which is a ternary complex made up of a cap-binding protein [https://en.wikipedia.org/wiki/EIF4E (eIF4E)] and RNA helicase [https://en.wikipedia.org/wiki/EIF4A (eIF4A)] connected by the bridging protein (eIF4G) (Figure 3).¹ Translation initiation is stimulated by the PABP bound to the poly(A) tail and its association with eIF4G.<ref name="Recognition of Polyadenylate RNA by the Poly(A)-Binding Protein">Deo, Rahul C, et al. “Recognition of Polyadenylate RNA by the Poly(A)-Binding Protein.” Cell 98:6. (1999) 835-845. Print. </ref> The 5’ UTR is unwound by the elF4F complex, and ribosomes are recruited to create the initiation complex. The eIF4G protein then guides the 40S subunit to the start codon (AUG), which is followed by the binding 60S ribosomal subunit, creating the 80S initiation complex.<ref name="Recognition of Polyadenylate RNA by the Poly(A)-Binding Protein">Deo, Rahul C, et al. “Recognition of Polyadenylate RNA by the Poly(A)-Binding Protein.” Cell 98:6. (1999) 835-845. Print. </ref> The association of the PABP and eIF4G gave rise to the name “closed loop.”<ref name="Recognition of Polyadenylate RNA by the Poly(A)-Binding Protein">Deo, Rahul C, et al. “Recognition of Polyadenylate RNA by the Poly(A)-Binding Protein.” Cell 98:6. (1999) 835-845. Print. </ref> Mutations of Arg→Ala and Lys→Ala in human eIF4G and in yeast extracts decrease the rate of translation initiation and destabilizing the interactions with PABP, indicating that basic residues are essential to the interaction with PABP.<ref name="Recognition of Polyadenylate RNA by the Poly(A)-Binding Protein">Deo, Rahul C, et al. “Recognition of Polyadenylate RNA by the Poly(A)-Binding Protein.” Cell 98:6. (1999) 835-845. Print. </ref> | ||
| + | |||
| + | [[Image:closedlooper.png|300px|right|thumb| "Figure 3:" Closed loop model of the eIF4F complex and PABP creating a loop out of the mRNA ]] | ||
| + | |||
| + | In more complex eukaryotic organisms, PABP indirectly stimulates translation via [https://en.wikipedia.org/wiki/PAIP1 PAIP-1] (PABP interacting protein). A higher presence of PAIP-1 increases the rate of translation initiation, indicating another way to “close the loop.”¹ | ||
</StructureSection> | </StructureSection> | ||
| - | == | + | ==Disease and Medical Relevance== |
| + | |||
| + | ===Oculopharyngeal Muscular Dystrophy=== | ||
| + | |||
| + | Oculopharyngeal muscular dystrophy, or [https://rarediseases.info.nih.gov/diseases/7245/oculopharyngeal-muscular-dystrophy OPMD], is an autosomal dominant late-onset disease. <ref name="Oculopharyngeal Muscular Dystrophy">“Oculopharyngeal Muscular Dystrophy.” NORD (National Organization for Rare Disorders), rarediseases.org/rare-diseases/oculopharyngeal-muscular-dystrophy/.</ref> It’s characterized by the myopathy of the eyelids and the throat. The symptoms entail eye-drooping and difficulty swallowing. There are two types of OPMD: autosomal dominant and recessive, both originating from the mutation of the PABP nuclear 1 [https://en.wikipedia.org/wiki/PABPN1 (PABPN1)] gene located on the long arm of chromosome 14. <ref name="Oculopharyngeal Muscular Dystrophy">“Oculopharyngeal Muscular Dystrophy.” NORD (National Organization for Rare Disorders), rarediseases.org/rare-diseases/oculopharyngeal-muscular-dystrophy/.</ref> This mutation results in an abnormally long polyalanine tract, 11-18 alanines, opposed to the normal 10. <ref name="Oculopharyngeal Muscular Dystrophy">“Oculopharyngeal Muscular Dystrophy.” NORD (National Organization for Rare Disorders), rarediseases.org/rare-diseases/oculopharyngeal-muscular-dystrophy/.</ref> Patients with longer PABPN1 expansion (more alanines) are on average diagnosed at an earlier in life than patients with a shorter expansion; therefore, expansion size plays a role in OPMD severity and progression. <ref name="“Correlation between PABPN1 Genotype and Disease Severity in Oculopharyngeal Muscular Dystrophy"> Richard, Pascale, et al. “Correlation between PABPN1 Genotype and Disease Severity in Oculopharyngeal Muscular Dystrophy.” Neurology, vol. 88, no. 4, 2016, pp. 359–365., doi:10.1212/wnl.0000000000003554. </ref> | ||
| + | |||
| + | The mutation results in PABPN1 forming clumps in muscle cells that can’t be degraded. <ref name="Oculopharyngeal Muscular Dystrophy">“Oculopharyngeal Muscular Dystrophy.” NORD (National Organization for Rare Disorders), rarediseases.org/rare-diseases/oculopharyngeal-muscular-dystrophy/.</ref> It’s suspected that this is a source of cell death for effected cells, however, it has not been concluded why this mutation only affects certain muscle cells. | ||
| + | |||
| + | ===Studies on Mutations=== | ||
| + | |||
| + | Studies conducted on [https://en.wikipedia.org/wiki/Drosophila Drosophila] are common due to 75% conservation between human and Drosophila genomes. Drosophila only encode one cytoplasmic PABP, and its deletion results in embryonic lethality. <ref name="Roles of Cytoplasmic Poly(A)-Binding Proteins">Gorgoni, Barbra, and Gray, Nicola. “The Roles of Cytoplasmic Poly(A)-Binding Proteins in Regulating Gene Expression: A Developmental Perspective.” Briefings in Functional Genomics and Proteomics, vol. 3, no. 2, 1 Aug. 2004, pp. 125–141., doi:10.1093/bfgp/3.2.125.</ref> Similarly, in [https://en.wikipedia.org/wiki/Caenorhabditis_elegans Caenorhabditis elegans], which have two cytoplasmic PABPs, display 50-80% embryonic lethality with the introduction of an RNAi to one of these PABPs. <ref name="Roles of Cytoplasmic Poly(A)-Binding Proteins">Gorgoni, Barbra, and Gray, Nicola. “The Roles of Cytoplasmic Poly(A)-Binding Proteins in Regulating Gene Expression: A Developmental Perspective.” Briefings in Functional Genomics and Proteomics, vol. 3, no. 2, 1 Aug. 2004, pp. 125–141., doi:10.1093/bfgp/3.2.125.</ref> | ||
| - | Sxl controls its own levels of expression via positive and negative [https://en.wikipedia.org/wiki/Autoregulation autoregulation]. Sxl binds its own pre-mRNA transcript in a similar manner as its downstream targets, Tra and Msl2. Through binding to its recognition element, Sxl causes a 3’ splice site to be skipped. [https://en.wikipedia.org/wiki/Alternative_splicing Alternative splicing] occurs utilizing a 3’ splice site further downstream, cleaving out a premature stop codon within Exon 3 and preventing truncation and inactivation<ref name="Black" />. This is a pathway of positive autoregulation, as functional Sxl protein must be present to cause proper processing of the pre-mRNA. The negative autoregulation pathway of Sxl proceeds via repression of its own translation. The Sxl transcript contains the target polyuridine sequence within its 3’UTR. Sxl binds this target, and blocks translation<ref name="Black" />. Negative autoregulation allows maintenance of a stable and standard Sxl protein concentration. An excess of Sxl increases the degree of translation repression because more Sxl protein are present to potentially bind at the 3’UTR, while a shortage allows for more unrepressed translation. | ||
| - | == Mutation == | ||
| - | In fruit flies, [https://en.wikipedia.org/wiki/Dosage_compensation dosage compensation] is achieved by hyperexpression of the single X chromosome found in the male genome. The single X chromosome is hyperexpressed and regulated by a group of genes known as the Male-specific lethal genes. The genes involve work together to produce a complex known as the Male-specific lethal complex (Msl complex). One gene that is important in forming this complex is the gene that codes for the Msl-2 protein. Without this protein, the Msl complex cannot form <ref name="Penalva">Penalva L, Sanchez L. RNA Binding Protein Sex-Lethal (Sxl) and Control of Drosophila Sex Determination and Dosage Compensation. Microbiol Mol Biol Rev.;67(3):343-356. [http://mmbr.asm.org/content/67/3/343.short doi: 10.1128/MMBR.67.3.343–359.2003]</ref>. Males exclusively make the Msl-2 protein and therefore exclusively create the Msl complex. Female cannot make the complex because the Sxl protein prevents the production of the Msl-2 protein. It prevents the protein formation by alternatively splicing of an intron near the 5’ UTR. In males this intron is included and in females it is not. <ref name="Penalva" /> Mutations of the Sxl protein can affect the formation of this complex and lead to death for females—hence the name sex lethal. If the Sxl gene is mutated in females, the formation of the Msl-2 complex will not be inhibited. If there is a functional Msl-2 complex in females then both of the X chromosomes will be hyperexpressed <ref name="Penalva" /> The overexpression of genes on the X chromosome is a fatal phenomenon ultimately caused by a mutation of the Sxl protein. Although a mutation to the Sxl gene is harmful to females, it has no effect on males because the functional Sxl gene is not expressed—a mutation in males has no phenotypic effect. | ||
== References == | == References == | ||
<references/> | <references/> | ||
Current revision
Contents |
Discussion of Human Poly(A) Binding Protein
Background
The Human Poly(A) Binding Protein (PABP) was discovered in 1973 by the use of a sedimentation profile detailing the RNase digestion differentiated the PABP protein. [1] Attempts to purify the 75 kDa protein then followed. In 1983, then considered “poly(A)-organizing protein,” was determined and purified by molecular weight, ligand-binding affinity, and amounts found in cytoplasmic portions of cell with ability to bind to free poly(A). [2]
PABP is a mRNA binding protein that binds to the 3’ Poly(A) tail on mRNA. It is comprised of four RNA recognition motifs (RRMs), which are highly conserved RNA-binding domains.[3] The RRM in PABP is found in over two hundred families of proteins across species, indicating that it is ancient.[3] Through extensive Adenosine recognition by the RRMs of PABP, the protein is involved in three main functions: recognition of the 3’ Poly(A) tail, mRNA stabilization, and eukaryotic translation initiation. The contributions of controlling gene expression via different families of PABPs is not yet fully understood. PABP families are divided into nuclear and cytoplasmic. [4] PABP1, which is predominantly cytoplasmic, is often referred to as PABP because it is the only form of PABP that has been extensively studied in its role with mRNA translation and stability. [4]
Structure
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Disease and Medical Relevance
Oculopharyngeal Muscular Dystrophy
Oculopharyngeal muscular dystrophy, or OPMD, is an autosomal dominant late-onset disease. [7] It’s characterized by the myopathy of the eyelids and the throat. The symptoms entail eye-drooping and difficulty swallowing. There are two types of OPMD: autosomal dominant and recessive, both originating from the mutation of the PABP nuclear 1 (PABPN1) gene located on the long arm of chromosome 14. [7] This mutation results in an abnormally long polyalanine tract, 11-18 alanines, opposed to the normal 10. [7] Patients with longer PABPN1 expansion (more alanines) are on average diagnosed at an earlier in life than patients with a shorter expansion; therefore, expansion size plays a role in OPMD severity and progression. [8]
The mutation results in PABPN1 forming clumps in muscle cells that can’t be degraded. [7] It’s suspected that this is a source of cell death for effected cells, however, it has not been concluded why this mutation only affects certain muscle cells.
Studies on Mutations
Studies conducted on Drosophila are common due to 75% conservation between human and Drosophila genomes. Drosophila only encode one cytoplasmic PABP, and its deletion results in embryonic lethality. [4] Similarly, in Caenorhabditis elegans, which have two cytoplasmic PABPs, display 50-80% embryonic lethality with the introduction of an RNAi to one of these PABPs. [4]
References
- ↑ Blobel, Gunter. “A Protein of Molecular Weight 78,000 Bound to the Polyadenylate Region of Eukaryotic Messenger Rnas.” Proceedings of the National Academy of Sciences of the United States of America, vol. 70, no. 3, 1973, pp. 924–8.
- ↑ Baer, Bradford W. and Kornberg, Roger D. "The Protein Responsible for the Repeating Structure of Cytoplasmic Poly(A)-Ribonucleoprotein." The Journal of Cell Biology, vol. 96, no. 3, Mar. 1983, pp. 717-721. EBSCOhost.
- ↑ 3.00 3.01 3.02 3.03 3.04 3.05 3.06 3.07 3.08 3.09 3.10 3.11 3.12 3.13 Deo, Rahul C, et al. “Recognition of Polyadenylate RNA by the Poly(A)-Binding Protein.” Cell 98:6. (1999) 835-845. Print.
- ↑ 4.0 4.1 4.2 4.3 4.4 Gorgoni, Barbra, and Gray, Nicola. “The Roles of Cytoplasmic Poly(A)-Binding Proteins in Regulating Gene Expression: A Developmental Perspective.” Briefings in Functional Genomics and Proteomics, vol. 3, no. 2, 1 Aug. 2004, pp. 125–141., doi:10.1093/bfgp/3.2.125.
- ↑ Kühn, Uwe and Elmar, Wahle. “Structure and Function of Poly(a) Binding Proteins.” Bba - Gene Structure & Expression, vol. 1678, no. 2/3, 2004.
- ↑ Wang, Zuoren and Kiledjian, Megerditch. “The Poly(A)-Binding Protein and an mRNA Stability Protein Jointly Regulate an Endoribonuclease Activity.” Molecular and Cellular Biology 20.17 (2000): 6334–6341. Print.
- ↑ 7.0 7.1 7.2 7.3 “Oculopharyngeal Muscular Dystrophy.” NORD (National Organization for Rare Disorders), rarediseases.org/rare-diseases/oculopharyngeal-muscular-dystrophy/.
- ↑ Richard, Pascale, et al. “Correlation between PABPN1 Genotype and Disease Severity in Oculopharyngeal Muscular Dystrophy.” Neurology, vol. 88, no. 4, 2016, pp. 359–365., doi:10.1212/wnl.0000000000003554.
