Ahmad Kashan Kashan Kashan yes BBTF19BM024.pptx

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1. RN A SYNTHESI S IN EUKARYOTES Name:Ahmad Kashan Roll No. BBTF19BM024 submitted to : Dr M. Kabir subject: Plant ecology

2. Transcription in Eukaryotes Much more complex phenomenon that in prokaryotes Eukaryotic transcription and translation are separated in space and time This separation has enabled eukaryotes to evolve post-transcriptional regulation strategies Transcription is directed by A single type of RNA polymerase in prokaryotes While four types of RNA polymerase are known in eukaryotes

3. Eukaryotic RNA Polymerase In Eukaryotes there are at least four types of polymerase, which differ in Template specificity Location within nucleus Susceptibility to inhibitors

4. Type Location Cellular transcripts Effects of α-amanitin I Nucleolus 18S, 5.8S and 28S rRNA Insensitive II Nucleoplasm mRNA precursors and snRNA Strongly inhibited III Nucleoplasm tRNA and 5S rRNA Inhibited by high concentration Table: Eukaryotic RNA polymerase

5. All these polymerase are large proteins, containing from 8-14 subunits and having a total molecular mass greater than 500 kDa These RNA polymerase are homologous to each other and to prokaryotic RNA polymerase, but RNA polymerase Ⅱ contains a unique carboxyl-terminal domain Activities of RNA Pol Ⅱ are regulated by serine/ threonine phosphorylation present in carboxyl-terminal domain

6. spRNAP- Ⅳ spRNAP- Ⅳ i.e. single-polypeptide RNA polymerase- Ⅳ was recently reported It is a homolog of mitochondrial RNA polymerase Localized in the nucleus Transcribes some genes in human and rodents It is insensitive to α- amanitin S It dost not respond to transcriptional enhancers.

7. α-Amanitin sensitivity α-Amanitin is produced by poisonous mushroom Amanita phalloides (commonly called death cup or destroying angel) It is a cyclic octapeptide that contains several modified amino acids It binds very tightly to RNA polymerase Ⅱ and blocks the elongation phase of transcription It also inhabit RNA polymerase Ⅲ but at much higher concentration RNA polymerase Ⅰ is insensitive to this toxin Amanita phalloides , a poisonous mushroom that produces -amanitin

8. Transcription Elements Cis-acting elements Promoters Enhancers Trans-acting elements

9. Cis-acting Elements Eukaryotic RNA polymerase also need specific promoter for transcriptional initiation Eukaryotic promoters also consist of conserved regions Promoters for different RNA polymerase differ in sequence and position Promoters are Generally located Upstream of gene, however, they maybe Embedded either in the gene or maybe Located downstream

10. Promoters of RNA Pol-I It transcribes ribosomal RN A Genes are arranged in several hundred tandem repeats Each repeat containing one copy each of 28S, 18S and 5.8S rRNA genes TATA-like sequence called ribosomal initiator element (rInr) is found at transcriptional start site. An Upstream promoter Element (UPE) is also available between -150 to -200 Both these elements bind to proteins that help to recruit RNA Polymerase-I

11. The RNA polymerase I promoter consists of a ribosomal initiator (rInr) and an upstream promoter element (UPE).

12. Promoters of RNA Pol-II Promoters of RNA Pol-II are located on 5’ side of the start of transcription For RNA Pol-II TATA box is the most important cis-acting element which is situated between position -30 and -100 TATA may be compulsory but not sufficient Promoter can contain any combination of a number of possible elements May also involve enhancer element, unique to eukaryotes

13. Other elements situated between -40 to +150 also important e.g. GC & CAAT boxes present on template strand. ‘’GC’’ box is required for constative gene expression ‘’CAAT’’ box is important for inducible gene expression -10 and -35 in prokaryotes act as direct binding sites for RNA pol. While TATA, CAAT, GC and other elements are binding sites for proteins other than RNA polymerase, called Transcription factors.

14. Promoters of RNA Pol-III L o cated with i n t r ans c rib e d sequ e n c e, do w nst r eam o f t h e start site These are of two types Type Ⅰ found in 5S rRNA genes, containing two short conserved sequence called A block and C block T y pe Ⅱ f o u n d i n tRN A ge n e s, consists o f tw o 11 b p se q ue n ce called A block and B block

15. RNA polymerase III promoters consist of conserved sequences that lie within the transcribed genes.

16. Enhancers Eukaryotes and their viruses contain DNA sequences called enhancers These are not promoters by themselves but enormously increase effectiveness of the promoters Position of the enhancers is not fixed and vary greatly Enhancer sequences can stimulate transcription at start sites located thousands of nucleotides away Enhancers may be located upstream, downstream or even in the midst of a transcribed gene

17. They influence the gene regulation in specific tissues and at specific developmental stage e.g. The immunoglobulin enhancer function in B lymphocytes but not elsewhere Presence of similar promoter and influence of similar enhancer create batteries of genes operative in a given set of internal/external environmental conditions.

18. Trans-acting elements/factors Cis-acting elements in eukaryotes are binding sites for proteins called transcription factor. These proteins are regarded as trans-acting elements/factors because their genes may be located on a different DNA molecules (other than their target gene) These proteins generally called transcription factors help RNA polymerase to bind to its promoter