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51-525: The progesterone receptor A ( PR-A ) is one of three known isoforms of the progesterone receptor (PR), the main biological target of the endogenous progestogen sex hormone progesterone . The other isoforms of the PR include the PR-B and PR-C . This article about a biochemical receptor is a stub . You can help Misplaced Pages by expanding it . Isoform The discovery of isoforms could explain

102-496: A DNA binding site called a promoter region before RNAP can initiate the DNA unwinding at that position. RNAP not only initiates RNA transcription, it also guides the nucleotides into position, facilitates attachment and elongation , has intrinsic proofreading and replacement capabilities, and termination recognition capability. In eukaryotes , RNAP can build chains as long as 2.4 million nucleotides. RNAP produces RNA that, functionally,

153-449: A beta (β) subunit of 150 kDa, a beta prime subunit (β′) of 155 kDa, and a small omega (ω) subunit. A sigma (σ) factor binds to the core, forming the holoenzyme. After transcription starts, the factor can unbind and let the core enzyme proceed with its work. The core RNA polymerase complex forms a "crab claw" or "clamp-jaw" structure with an internal channel running along the full length. Eukaryotic and archaeal RNA polymerases have

204-453: A distinct subset of RNA: The 2006 Nobel Prize in Chemistry was awarded to Roger D. Kornberg for creating detailed molecular images of RNA polymerase during various stages of the transcription process. In most prokaryotes , a single RNA polymerase species transcribes all types of RNA. RNA polymerase "core" from E. coli consists of five subunits: two alpha (α) subunits of 36  kDa ,

255-492: A few single-subunit RNA polymerases (ssRNAP) from phages and organelles. The other multi-subunit RNAP lineage formed all of the modern cellular RNA polymerases. In bacteria , the same enzyme catalyzes the synthesis of mRNA and non-coding RNA (ncRNA) . RNAP is a large molecule. The core enzyme has five subunits (~400 kDa ): In order to bind promoters, RNAP core associates with the transcription initiation factor sigma (σ) to form RNA polymerase holoenzyme. Sigma reduces

306-434: A gene that serves as an initial binding site—resulting in slightly modified transcripts and protein isoforms. Generally, one protein isoform is labeled as the canonical sequence based on criteria such as its prevalence and similarity to orthologous —or functionally analogous—sequences in other species. Isoforms are assumed to have similar functional properties, as most have similar sequences, and share some to most exons with

357-419: A group consisting of 10 PAPs was identified through biochemical methods, which was later extended to 12 PAPs. Chloroplast also contain a second, structurally and mechanistically unrelated, single-subunit RNAP ("nucleus-encoded polymerase, NEP"). Eukaryotic mitochondria use POLRMT (human), a nucleus-encoded single-subunit RNAP. Such phage-like polymerases are referred to as RpoT in plants. Archaea have

408-509: A multi-subunit RNAP ("PEP, plastid-encoded polymerase"). Due to its bacterial origin, the organization of PEP resembles that of current bacterial RNA polymerases: It is encoded by the RPOA, RPOB, RPOC1 and RPOC2 genes on the plastome, which as proteins form the core subunits of PEP, respectively named α, β, β′ and β″. Similar to the RNA polymerase in E. coli , PEP requires the presence of sigma (σ) factors for

459-556: A result, the 8 bp DNA-RNA hybrid in the transcription complex shifts to a 4 bp hybrid. These last 4 base pairs are weak A-U base pairs, and the entire RNA transcript will fall off the DNA. Transcription termination in eukaryotes is less well understood than in bacteria, but involves cleavage of the new transcript followed by template-independent addition of adenines at its new 3′ end, in a process called polyadenylation . Given that DNA and RNA polymerases both carry out template-dependent nucleotide polymerization, it might be expected that

510-497: A similar core structure and work in a similar manner, although they have many extra subunits. All RNAPs contain metal cofactors , in particular zinc and magnesium cations which aid in the transcription process. Control of the process of gene transcription affects patterns of gene expression and, thereby, allows a cell to adapt to a changing environment, perform specialized roles within an organism, and maintain basic metabolic processes necessary for survival. Therefore, it

561-472: A single type of RNAP, responsible for the synthesis of all RNA. Archaeal RNAP is structurally and mechanistically similar to bacterial RNAP and eukaryotic nuclear RNAP I-V, and is especially closely structurally and mechanistically related to eukaryotic nuclear RNAP II. The history of the discovery of the archaeal RNA polymerase is quite recent. The first analysis of the RNAP of an archaeon was performed in 1971, when

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612-522: A structural function. Archaeal RNAP subunit previously used an "RpoX" nomenclature where each subunit is assigned a letter in a way unrelated to any other systems. In 2009, a new nomenclature based on Eukaryotic Pol II subunit "Rpb" numbering was proposed. Orthopoxviruses and some other nucleocytoplasmic large DNA viruses synthesize RNA using a virally encoded multi-subunit RNAP. They are most similar to eukaryotic RNAPs, with some subunits minified or removed. Exactly which RNAP they are most similar to

663-462: A template instead of DNA). This occurs in negative strand RNA viruses and dsRNA viruses , both of which exist for a portion of their life cycle as double-stranded RNA. However, some positive strand RNA viruses , such as poliovirus , also contain RNA-dependent RNAP. RNAP was discovered independently by Charles Loe, Audrey Stevens , and Jerard Hurwitz in 1960. By this time, one half of

714-457: Is bacteriophage T7 RNA polymerase . ssRNAPs cannot proofread. B. subtilis prophage SPβ uses YonO, a homolog of the β+β′ subunits of msRNAPs to form a monomeric (both barrels on the same chain) RNAP distinct from the usual "right hand" ssRNAP. It probably diverged very long ago from the canonical five-unit msRNAP, before the time of the last universal common ancestor . Other viruses use an RNA-dependent RNAP (an RNAP that employs RNA as

765-592: Is a major molecular mechanism that may contribute to protein diversity. The spliceosome , a large ribonucleoprotein , is the molecular machine inside the nucleus responsible for RNA cleavage and ligation , removing non-protein coding segments ( introns ). Because splicing is a process that occurs between transcription and translation , its primary effects have mainly been studied through genomics techniques—for example, microarray analyses and RNA sequencing have been used to identify alternatively spliced transcripts and measure their abundances. Transcript abundance

816-417: Is a topic of debate. Most other viruses that synthesize RNA use unrelated mechanics. Many viruses use a single-subunit DNA-dependent RNAP (ssRNAP) that is structurally and mechanistically related to the single-subunit RNAP of eukaryotic chloroplasts (RpoT) and mitochondria ( POLRMT ) and, more distantly, to DNA polymerases and reverse transcriptases . Perhaps the most widely studied such single-subunit RNAP

867-402: Is an enzyme that catalyzes the chemical reactions that synthesize RNA from a DNA template. Using the enzyme helicase , RNAP locally opens the double-stranded DNA so that one strand of the exposed nucleotides can be used as a template for the synthesis of RNA, a process called transcription . A transcription factor and its associated transcription mediator complex must be attached to

918-519: Is an additional subunit dubbed Rpo13; together with Rpo5 it occupies a space filled by an insertion found in bacterial β′ subunits (1,377–1,420 in Taq ). An earlier, lower-resolution study on S. solfataricus structure did not find Rpo13 and only assigned the space to Rpo5/Rpb5. Rpo3 is notable in that it's an iron–sulfur protein . RNAP I/III subunit AC40 found in some eukaryotes share similar sequences, but does not bind iron. This domain, in either case, serves

969-422: Is derived by the protein's structure/function, as well as the cell type and developmental stage during which they are produced. Determining specificity becomes more complicated when a protein has multiple subunits and each subunit has multiple isoforms. For example, the 5' AMP-activated protein kinase (AMPK), an enzyme, which performs different roles in human cells, has 3 subunits: In human skeletal muscle,

1020-458: Is either for protein coding , i.e. messenger RNA (mRNA); or non-coding (so-called "RNA genes"). Examples of four functional types of RNA genes are: RNA polymerase is essential to life, and is found in all living organisms and many viruses . Depending on the organism, a RNA polymerase can be a protein complex (multi-subunit RNAP) or only consist of one subunit (single-subunit RNAP, ssRNAP), each representing an independent lineage. The former

1071-481: Is found in bacteria , archaea , and eukaryotes alike, sharing a similar core structure and mechanism. The latter is found in phages as well as eukaryotic chloroplasts and mitochondria , and is related to modern DNA polymerases . Eukaryotic and archaeal RNAPs have more subunits than bacterial ones do, and are controlled differently. Bacteria and archaea only have one RNA polymerase. Eukaryotes have multiple types of nuclear RNAP, each responsible for synthesis of

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1122-419: Is hardly surprising that the activity of RNAP is long, complex, and highly regulated. In Escherichia coli bacteria, more than 100 transcription factors have been identified, which modify the activity of RNAP. RNAP can initiate transcription at specific DNA sequences known as promoters . It then produces an RNA chain, which is complementary to the template DNA strand. The process of adding nucleotides to

1173-532: Is no conclusive evidence that it acts primarily by producing novel protein isoforms. Alternative splicing generally describes a tightly regulated process in which alternative transcripts are intentionally generated by the splicing machinery. However, such transcripts are also produced by splicing errors in a process called "noisy splicing," and are also potentially translated into protein isoforms. Although ~95% of multi-exonic genes are thought to be alternatively spliced, one study on noisy splicing observed that most of

1224-543: Is often used as a proxy for the abundance of protein isoforms, though proteomics experiments using gel electrophoresis and mass spectrometry have demonstrated that the correlation between transcript and protein counts is often low, and that one protein isoform is usually dominant. One 2015 study states that the cause of this discrepancy likely occurs after translation, though the mechanism is essentially unknown. Consequently, although alternative splicing has been implicated as an important link between variation and disease, there

1275-493: The blood proteins as orosomucoid , antitrypsin , and haptoglobin . An unusual glycoform variation is seen in neuronal cell adhesion molecule, NCAM involving polysialic acids, PSA . Monoamine oxidase , a family of enzymes that catalyze the oxidation of monoamines, exists in two isoforms, MAO-A and MAO-B. RNA polymerase In molecular biology , RNA polymerase (abbreviated RNAP or RNApol ), or more specifically DNA-directed/dependent RNA polymerase ( DdRP ),

1326-517: The rho factor , which destabilizes the DNA-RNA heteroduplex and causes RNA release. The latter, also known as intrinsic termination , relies on a palindromic region of DNA. Transcribing the region causes the formation of a "hairpin" structure from the RNA transcription looping and binding upon itself. This hairpin structure is often rich in G-C base-pairs, making it more stable than the DNA-RNA hybrid itself. As

1377-421: The " transcription bubble ". Supercoiling plays an important part in polymerase activity because of the unwinding and rewinding of DNA. Because regions of DNA in front of RNAP are unwound, there are compensatory positive supercoils. Regions behind RNAP are rewound and negative supercoils are present. RNA polymerase then starts to synthesize the initial DNA-RNA heteroduplex, with ribonucleotides base-paired to

1428-419: The DNA template strand. As transcription progresses, ribonucleotides are added to the 3′ end of the RNA transcript and the RNAP complex moves along the DNA. The characteristic elongation rates in prokaryotes and eukaryotes are about 10–100 nts/sec. Aspartyl ( asp ) residues in the RNAP will hold on to Mg ions, which will, in turn, coordinate the phosphates of the ribonucleotides. The first Mg will hold on to

1479-524: The DNA template. This pauses transcription. The polymerase then backtracks by one position and cleaves the dinucleotide that contains the mismatched nucleotide. In the RNA polymerase this occurs at the same active site used for polymerization and is therefore markedly different from the DNA polymerase where proofreading occurs at a distinct nuclease active site. The overall error rate is around 10 to 10 . In bacteria, termination of RNA transcription can be rho-dependent or rho-independent. The former relies on

1530-425: The RNA strand is known as elongation; in eukaryotes, RNAP can build chains as long as 2.4 million nucleotides (the full length of the dystrophin gene). RNAP will preferentially release its RNA transcript at specific DNA sequences encoded at the end of genes, which are known as terminators . Products of RNAP include: RNAP accomplishes de novo synthesis . It is able to do this because specific interactions with

1581-487: The RNAP from the extreme halophile Halobacterium cutirubrum was isolated and purified. Crystal structures of RNAPs from Sulfolobus solfataricus and Sulfolobus shibatae set the total number of identified archaeal subunits at thirteen. Archaea has the subunit corresponding to Eukaryotic Rpb1 split into two. There is no homolog to eukaryotic Rpb9 ( POLR2I ) in the S. shibatae complex, although TFS (TFIIS homolog) has been proposed as one based on similarity. There

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1632-425: The abundance of mRNA transcript isoforms does not necessarily correlate with the abundance of protein isoforms. Three-dimensional protein structure comparisons can be used to help determine which, if any, isoforms represent functional protein products, and the structure of most isoforms in the human proteome has been predicted by AlphaFold and publicly released at isoform.io . The specificity of translated isoforms

1683-523: The active center stabilizes the elongation complex. However, promoter escape is not the only outcome. RNA polymerase can also relieve the stress by releasing its downstream contacts, arresting transcription. The paused transcribing complex has two options: (1) release the nascent transcript and begin anew at the promoter or (2) reestablish a new 3′-OH on the nascent transcript at the active site via RNA polymerase's catalytic activity and recommence DNA scrunching to achieve promoter escape. Abortive initiation ,

1734-449: The affinity of RNAP for nonspecific DNA while increasing specificity for promoters, allowing transcription to initiate at correct sites. The complete holoenzyme therefore has 6 subunits: β′βα and α ωσ (~450 kDa). Eukaryotes have multiple types of nuclear RNAP, each responsible for synthesis of a distinct subset of RNA. All are structurally and mechanistically related to each other and to bacterial RNAP: Eukaryotic chloroplasts contain

1785-491: The canonical sequence. However, some isoforms show much greater divergence (for example, through trans-splicing ), and can share few to no exons with the canonical sequence. In addition, they can have different biological effects—for example, in an extreme case, the function of one isoform can promote cell survival, while another promotes cell death—or can have similar basic functions but differ in their sub-cellular localization. A 2016 study, however, functionally characterized all

1836-612: The core promoter region containing the −35 and −10 elements (located before the beginning of sequence to be transcribed) and also, at some promoters, the α subunit C-terminal domain recognizing promoter upstream elements. There are multiple interchangeable sigma factors, each of which recognizes a distinct set of promoters. For example, in E. coli , σ is expressed under normal conditions and recognizes promoters for genes required under normal conditions (" housekeeping genes "), while σ recognizes promoters for genes required at high temperatures (" heat-shock genes "). In archaea and eukaryotes,

1887-486: The different low-abundance transcripts are noise, and predicts that most alternative transcript and protein isoforms present in a cell are not functionally relevant. Other transcriptional and post-transcriptional regulatory steps can also produce different protein isoforms. Variable promoter usage occurs when the transcriptional machinery of a cell ( RNA polymerase , transcription factors , and other enzymes ) begin transcription at different promoters—the region of DNA near

1938-524: The discrepancy between the small number of protein coding regions of genes revealed by the human genome project and the large diversity of proteins seen in an organism: different proteins encoded by the same gene could increase the diversity of the proteome . Isoforms at the RNA level are readily characterized by cDNA transcript studies. Many human genes possess confirmed alternative splicing isoforms. It has been estimated that ~100,000 expressed sequence tags ( ESTs ) can be identified in humans. Isoforms at

1989-464: The function of each isoform must generally be determined separately, most identified and predicted isoforms still have unknown functions. A glycoform is an isoform of a protein that differs only with respect to the number or type of attached glycan . Glycoproteins often consist of a number of different glycoforms, with alterations in the attached saccharide or oligosaccharide . These modifications may result from differences in biosynthesis during

2040-486: The functions of the bacterial general transcription factor sigma are performed by multiple general transcription factors that work together. The RNA polymerase-promoter closed complex is usually referred to as the " transcription preinitiation complex ." After binding to the DNA, the RNA polymerase switches from a closed complex to an open complex. This change involves the separation of the DNA strands to form an unwound section of DNA of approximately 13 bp, referred to as

2091-429: The initiating nucleotide hold RNAP rigidly in place, facilitating chemical attack on the incoming nucleotide. Such specific interactions explain why RNAP prefers to start transcripts with ATP (followed by GTP, UTP, and then CTP). In contrast to DNA polymerase , RNAP includes helicase activity, therefore no separate enzyme is needed to unwind DNA. RNA polymerase binding in bacteria involves the sigma factor recognizing

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2142-475: The initiation complex. During the promoter escape transition, RNA polymerase is considered a "stressed intermediate." Thermodynamically the stress accumulates from the DNA-unwinding and DNA-compaction activities. Once the DNA-RNA heteroduplex is long enough (~10 bp), RNA polymerase releases its upstream contacts and effectively achieves the promoter escape transition into the elongation phase. The heteroduplex at

2193-413: The isoforms of 1,492 genes and determined that most isoforms behave as "functional alloforms." The authors came to the conclusion that isoforms behave like distinct proteins after observing that the functional of most isoforms did not overlap. Because the study was conducted on cells in vitro , it is not known if the isoforms in the expressed human proteome share these characteristics. Additionally, because

2244-456: The preferred form is α2β2γ1. But in the human liver, the most abundant form is α1β2γ1. The primary mechanisms that produce protein isoforms are alternative splicing and variable promoter usage, though modifications due to genetic changes, such as mutations and polymorphisms are sometimes also considered distinct isoforms. Alternative splicing is the main post-transcriptional modification process that produces mRNA transcript isoforms, and

2295-404: The process of glycosylation , or due to the action of glycosidases or glycosyltransferases . Glycoforms may be detected through detailed chemical analysis of separated glycoforms, but more conveniently detected through differential reaction with lectins , as in lectin affinity chromatography and lectin affinity electrophoresis . Typical examples of glycoproteins consisting of glycoforms are

2346-407: The protein level can manifest in the deletion of whole domains or shorter loops, usually located on the surface of the protein. One single gene has the ability to produce multiple proteins that differ both in structure and composition; this process is regulated by the alternative splicing of mRNA, though it is not clear to what extent such a process affects the diversity of the human proteome, as

2397-447: The recognition of its promoters, containing the -10 and -35 motifs. Despite the many commonalities between plant organellar and bacterial RNA polymerases and their structure, PEP additionally requires the association of a number of nuclear encoded proteins, termed PAPs (PEP-associated proteins), which form essential components that are closely associated with the PEP complex in plants. Initially,

2448-500: The template DNA strand according to Watson-Crick base-pairing interactions. As noted above, RNA polymerase makes contacts with the promoter region. However these stabilizing contacts inhibit the enzyme's ability to access DNA further downstream and thus the synthesis of the full-length product. In order to continue RNA synthesis, RNA polymerase must escape the promoter. It must maintain promoter contacts while unwinding more downstream DNA for synthesis, "scrunching" more downstream DNA into

2499-466: The two types of enzymes would be structurally related. However, x-ray crystallographic studies of both types of enzymes reveal that, other than containing a critical Mg ion at the catalytic site, they are virtually unrelated to each other; indeed template-dependent nucleotide polymerizing enzymes seem to have arisen independently twice during the early evolution of cells. One lineage led to the modern DNA polymerases and reverse transcriptases, as well as to

2550-422: The unproductive cycling of RNA polymerase before the promoter escape transition, results in short RNA fragments of around 9 bp in a process known as abortive transcription. The extent of abortive initiation depends on the presence of transcription factors and the strength of the promoter contacts. The 17-bp transcriptional complex has an 8-bp DNA-RNA hybrid, that is, 8 base-pairs involve the RNA transcript bound to

2601-476: The α-phosphate of the NTP to be added. This allows the nucleophilic attack of the 3′-OH from the RNA transcript, adding another NTP to the chain. The second Mg will hold on to the pyrophosphate of the NTP. The overall reaction equation is: Unlike the proofreading mechanisms of DNA polymerase those of RNAP have only recently been investigated. Proofreading begins with separation of the mis-incorporated nucleotide from

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