Polymerases in eukaryotes

The primary transcripts generated by RNA polymerase II—one of three distinct nuclear DNA-depen-dent RNA polymerases in eukaryotes—are promptly capped by 7-methylguanosine triphosphate caps (Figure 35-10) that persist and eventually appear on the 5 end of mature cytoplasmic mRNA. These caps are necessary for the subsequent processing of the primary transcript to mRNA, for the translation of the mRNA, and for protection of the mRNA against exonucleolytic attack. 

There s not just one DNA polymerase there s a whole army. DNA replication actually occurs in large complexes containing many proteins and sometimes many polymerases. In eukaryotic cells we have to replicate both mitochondrial and nuclear DNA, and there are specific DNA polymerases for each. In addition to DNA replication, you have to make new DNA when you repair. Consequently, the function may be specialized for repair or replication. There can also be specialization for making the leading or lagging strand. Some of the activities of DNA polymerases from eukaryotes and prokaryotes are shown in the table on the next page. 

With respect to the general mechanism of RNA chain growth, are there any differences among the various types of RNA polymerase in eukaryotic cells ... 

General transcription factors belong to a group of proteins which assemble at the TATA box or a similar region in the promoter.They are required for the initiation of transcription by the DNA-dependent RNA polymerase in eukaryotic cells. 

Rifampicin (Fig. 10.70) is a semisynthetic rifamycin made from rifamycin B—an antibiotic isolated from Streptomyces mediterranei. It inhibits Gram-positive bacteria and works by binding non-covalently to RNA polymerase and inhibiting RNA synthesis. The DNA-dependent RNA polymerases in eukaryotic cells are unaffected, since the drug binds to a peptide chain not present in the mammalian RNA polymerase. It is therefore highly selective. 

Tujo distinct polymerases are needed to copy a eukaryotic replicon. An initiator polymerase called polymerase a begins replication but is soon replaced by a more processive enzyme. This process is called polymerase switching because one polymerase has replaced another. This second enzyme, called DNA polymerase 5, is the principal replicative polymerase in eukaryotes (Table 28.2). 

Polymerase a was the first discovered, and it has the most subunits. It also has the ability to make primers, but it lacks a 3 5 proofreading activity and has low processivity. After making the RNA primer, Pol a adds about 20 nucleotides and is then replaced by Pol 5 and e. Polymerase 5 is the principal DNA polymerase in eukaryotes. It interacts with a special protein called PCNA ior proliferating cell nuclear antigen). PCNA is the eukaryotic equivalent of the part of Pol III that functions as a sliding clamp (P). It is a trimer of three identical proteins that surround the DNA (Figure 10.19). The role of DNA polymerase e is less clear. It may replace polymerase 5 in lagging strand synthesis. DNA polymerase p appears to be a repair enzyme. DNA polymerase y carries out DNA replication in mitochondria. Several... 

There are three RNA polymerases in eukaryotes, of which Pol II produces mRNA. 

RNA polymerase II the RNA polymerase in eukaryotes that makes mRNA also called RNA polymerase B (11.4) rRNA (ribosomal RNA) the kind of RNA found in ribosomes (9.5)... 

There are three RNA polymerases in eukaryotes, compared with one in prokaryotes. There are many more transcription factors in eukaryotes, including complexes of them necessary for polymerase recruitment. RNA is extensively processed after transcription in eukaryotes, and, in most cases, the mRNA must leave the nucleus to be translated, whereas translation and transcription can occur at the same time in prokaryotes. 

Roeder, R.G., Rutter, W.J. Multiple forms of DNA-dependent RNA polymerase in eukaryotic organisms. Nature (Lond.) 224,234-237 (1969)... 

RNA polymerases interact with unique cw-active regions of genes, termed promoters, in order to form preinitiation complexes (PICs) capable of initiation. In eukaryotes the process of PIC formation is facilitated by multiple general transcription factors (GTFs), TFIIA, B, D, E, F, and H. 

A second example of the differences between important biomolecules in archaebacteria and eubacteria is their DNA-dependent RNA polymerase. The enzyme found in archaea resembles that in eukaryotes more than it does those in bacteria ... 

Regulation of transcription is a central mechanism by which cells respond to developmental and environmental cues. RNA polymerase Il-mediated transcription in eukaryotes is to a large extent regulated at the level of chromatin, which forms a physical barrier for the binding of proteins to the promoter region of a target gene. The basic unit of chromatin is the nucleosome, which consists of an octamer of histone proteins around which the DNA is wrapped (see Fig. la). 

Telomeres are r etitive sequences at the ends of linear DNA molecules in eukaryotic chromosomes. With each round of replication in most normal cells, the telomeres are shortened because DNA polymerase cannot complete synthesis of the 5 end of each strand. This contributes to the aging of cells, because eventually the telomeres become so short that the chromosomes cannot function properly and the cells die. 

Transcription factors (such as TFIID for RNA polymerase II) help to initiate transcription. The requirements for termination of transcription in eukaryotes are not well understood. All transcription can be inhibited by actinomycin D. In addition, RNA polymerase II is inhibited by a-amanitin (a toxin from certain mushrooms). These points are summarized in Table 1-3-1,... 

RNA polymerase II ends transcription when it reaches a termination signal. These signals are not well understood in eukaryotes. 

In eukaryotes, general transcription factors must bind to the promoter to allow RNA polymerase II to bind and form the initiation complex at the start site for transcription. General manscription factors are common to most genes. The general transcription factor TFIID (the TATA fector) must bind to the TATA box before RNA polymerase II can bind. Other examples delude SP-1 and NF-.l that modulate basal transcription of many genes. 

In eukaryotic cells, there are three classes of RNA polymerases (I, II and III) which synthesise different classes of RNA, as follows ... 

DNA-directed RNA polymerase [EC 2.1.1.6] catalyzes the DNA-template-directed extension of the 3 -end of an RNA strand by one nucleotide at a time thus, n nucleoside triphosphate generate RNA and n pyrophosphate. The enzyme can initiate a chain de novo. Three forms of the enzyme have been distinguished in eukaryotes on the basis of sensitivity of a-amanitin and the type of RNA synthesized. See also Replicase... 

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