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Transcription

Transcription. AHMP 5406. Objectives:. Describe the general process of DNA transcription Discuss the function of RNA polymerases Differentiate between messenger RNA, small nuclear RNA, ribosomal RNA, and transfer RNA Explain the purpose of the promoter and terminator

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  1. Transcription AHMP 5406

  2. Objectives: • Describe the general process of DNA transcription • Discuss the function of RNA polymerases • Differentiate between messenger RNA, small nuclear RNA, ribosomal RNA, and transfer RNA • Explain the purpose of the promoter and terminator • Compare and contrast the three types of RNA polymerase in eukaryotic nuclei: RNA polymerase I, RNA polymerase II, and RNA polymerase III • Discuss RNA capping & RNA splicing • Explain the selective exportation of mRNAs from the nucleus • Discuss ribosomal RNA and its function

  3. What is Transcription ? • DNA does not direct protein synthesis directly • RNA is used as intermediate • Transcription = Copying DNA into RNA

  4. RNA • Ribonucleic Acids • Contain ribose instead of deoxyribose (DNA) • Polymers formed by A, G, C, and U • U = uracil • Complementary to Thymine • But sometimes binds to G • Single stranded • Can bind to itself and form 3D structures

  5. DNA to RNA • Transcribed RNA is complementary to 1 DNA strand • Transcribe RNA = transcript • Transcripts can only reach a few thousand bases in length • Unidirectional • 5’-3’

  6. RNA Types • Coding • Messenger RNA (mRNA) • Non-coding • Ribosomal RNA (rRNA) • Transfer RNA (tRNA) • Small nuclear RNA (snRNA) • Small nucleolar RNA (snoRNA)

  7. mRNA • Codes for proteins • Can be modified by splicing, 5’ and 3’ additions

  8. Non-Coding RNAs • rRNA • Form basic structure of ribosome • tRNA • Adaptors b/w mRNA and AA • snRNA • Several nuclear processes • Assist in splicing • snoRNA • Process and chemically modify rRNA

  9. DNA sequences used as signals • Sequences signal transcription to start and stop • Promoters – start signals • s factor binds to promoter in bacteria • Terminators – stop signals • In bacteria A-T region causes fold

  10. RNA polymerases • RNA pols perform transcription • Immediate release of transcript allows many RNA to be synth. in a short time • More error prone than DNA pols • They have limited error correction abilities

  11. Types of RNA polymerases • RNA pol I • Transcribe 5.8S, 18S and 28S genes • S = Svedberg units, sedimentation coefficient related to centrifugation • The bigger the number the bigger the molecule • RNA pol II • Transcribe protein-coding genes, snoRNA genes • Some snRNA genes • RNA pol III • Transcribe tRNA genes and some snRNA genes

  12. Transcription in Euks. • Requires general transcription factors to initiate transcription • GTFs help position RNA pol II correctly • TFIIA, TFIIB, TFIID, TFIIE, TFIIF, TFIIH • Recognize specific sites on DNA

  13. RNA pol start points • TATA Box promoter • Recognized by subunit of TFIID • TBP – TATA Box Binding protein • approx 25 bp upstream of initiation point • Causes conformational change in DNA

  14. Other necessary proteins for initiation of transcription • Transcriptional activators • Attract RNA pol II to start site • Mediators • Allow communication between activator and GTFs • Chromatin remodeling proteins • Modify DNA conformation

  15. Elongation Factors • Increase affinity of RNA Pol II to DNA • Assist in moving through chromatin structure

  16. mRNA modifications • 5’ capping • Splicing = removing introns • 3’ Polyadenylation

  17. 5’ Capping • When RNA Pol II at 25 bases 5’ cap is added to transcript • Modified guanine • Three enzymes • Phosphatase = removes phosphate from 5’ end • Guanyl transferase = adds GMP • Reversed linkage 5’-5’ • Methyltransferase = adds methyl group to guanosine

  18. Splicing • Removes introns (or sometimes shuffling) • Sequences signal where to cut • Spliceosome performs cutting • Complex of snRNAs (Us) and protein

  19. Self Splicing Mechanisms • Group I • Bind free G nucleotide to specific site • Group II • Uses reactive A nucleotide • Unusual mechanisms

  20. Poly A tail • Cleavage stimulating factor F (CstF) • Cleavage and polyadenylation specificity factor (CPSF)

  21. Export of mRNA • Goes through nuclear pore complexes • Mature mRNA are modified and protein bound • Signal (passport) for transport of mRNA outside of nucleus • 5’ cap proceeds first

  22. rRNAs • Transcribed by RNA Pol I • rRNAs are made from a larger precursor • Sections are modified, cleaved and assembled into ribosomes in nucleolus

  23. Subnuclear structures • Sites where snRNA processing machinery is assembled, stored and recycled • Cajal bodies • GEMS (Gemini of coiled-bodies) • Interchromatin granule structures

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