Class Note for MCB 411 with Professor Fares at UA
Class Note for MCB 411 with Professor Fares at UA
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Date Created: 02/06/15
EUKARYOTIC TRANSCRIPTION chromusumn 039 15 x ui nuclnmidu pairs conlmnmg about 3mm gnrlns 05 ul cllrumosomu cumuminy 15 unncs WWW 539 GAAATTCAAGCCCTG 339 W myw 3 CTTTAAGTTCGGGAC 5 mRNA 5 7 GAAAUUCAAGCCCUG 7 3 mm 9 mm L IA mm wunu W mum ruhm um um m w w mm m m or MEoI F 1995 5mm Pam shng m TRANSCRIPTION mRNA TRANSLATION pmloln mRNA TRANSLATION molein mm The Art of HEM 99s Garland Pam smng Inc A EUCARYdjTES cytoplasm nucteus exons inlrons Ira nscription unik quot TRANSCRIPTION primary RNA transcriptquot 539 CAPPING 5513 SLICING RNA 39 ADENYLATION mRNA AAAA EXPORT mRNA AAAA lTRANSLATION protein El Figure 641 part 1 of 2 Molecular Blology oithe Ce 4th Edition TABLE blThe Three RNA Pnlymemes in Eucaryutic Cell TYPE OF FOLYHEMSE GEVETEANKRIBED RNA puimunuvl INA pub a39 185 Im J 255 EINA 3mm plutulmuding gum 111m mullM gum HIM mlllL39 MINA gums HNA pulf numuv Ill IRMA gum is I RNA Rem zwmu zlllNA gum and gum5 flu lllht l mull RNA Lwll M m W w CHAIN TERNHNATION CHAIN y HWl n K INITIATION WW WWW t CH ELONGATION I From lne an or M80123 l995 Garland PuhlrsmmL In 41W 0w Eukagote Gene DNA Gene DNA RNA Polymer RNA Domini RNA NADA What does this tell you TATA ix x Z A l O TFllA Bl Cl start of transcription TBP TFIID UTP ATP 0 TFIIE TFIIF other factors r TFIIE re TFlIH Figure 16 RNA polymerase II Molecular Biology of the Cell 4th Edition transcription start point 35 30 quot 30 q BRE INR DPE TATA general element 0521quotges tran t rtigrtion BRE GC GC GAtC G C C TFllB TATA TATAATAAT TBP INR CIT CTA N TIA CT CIT TFIID DPE AG G NT 3 GT G TFIID Figure 6 17t Molecular Biology of the Cell 4th Edition What is another difference between prokaryotic and eukaryotic chromosomes chromatin remode ng complex 5 CAP TRANSCRIPTION BEGINS Figure 6719 Molecular Biology of the Cell 4th Edition 539 mm ul mum m 7 wwlllylulmnosinc 5 no a mRNA 39 m m mmNpr r mman n wn mvlmsnlvaw lmdnu mmNpND n ml mnxhy mm a base 0 GupuNpr and mum Hour 1 Hum um 3 H595 5mm Fm i w upuprNp u UH E RNA polymerase polyadenylation splicing factors factors 1 What do you think will happen if the RNA moelcnle capping factors isn t capped 2 Contrast that with Prokaryotes RNA Figure 5723 Molecuiar Biology of the Ce 4th Edition mm 1 Wu CHAIN s INITIATION WAWMR Iwzux my CHAIN ELONGATION CHAIN TERMINATION HrmimwArn n AW aiM may rt INDIAtqu From The Art or M8053 1995 Garland Publishing Inc TATA start of transcription box 39 1 Al li TBP TFllD Bl O TFIIA TFIIB D la 00 Cl 1 RNA TFm other factors N TRANSCRIPTION TFllE TFIIH RNA polymerase II Figure 6 16 Molecular Biology of the Cell 4th Edition site for incoming ribonucleoside triphosphate displaced RNA chain W WWW CHAIN a 7 INITIATION JAWWAWJ CHAIN ELONGATION 3m CHAIN TERMINATION 3 From The Art of MRS a was Garland Fuhlysmng lne DNA helix unwinding rewlnding site site short region of DNARNA helix From The Art of NEE 31995 Garland Puhlvghing Inc 10 30 s 30 nucleotides nucleotides GAl G Urich or Urich l CLEAVAGE a AAUA AA CAlOHl GU rich or Uwrich degraded in the nucleus PolyaA ADDITION AA AA A A39 OH 250 Figure 6737 Molecular Biology of the Cell 4th Edition Figure 638 cleavage and poly signals encoded in DNA RNA polymerase O polyA polymerase lPAPl polyAbinding protein CPSF POLYVA LENGTH polyA polymerase 0 polyaAabinding protein RNA polymerase eventually terminates mature 3 end of Molecular Biology of the Cell Al O 0 i AAUAAA AAAAAAAAA additional polyaArbinding AAUAAA AAAAAAAAAA 200AAAAAAAAAA 39 additional cleavage factors RNA CLEAVED RNA polymerase eventually terminates PAP additional cleavage REGULATION protein actors Q RNA CLEAVED O an mRNA molecule mam INAWManHL mm mmamw w W39 5 gr m 0 cm wmmuso EmNnmoN am wucmammmmms mums mm aimmin 4315 D nvw N a pubA 3 21 mmon rsmmmu 0 am WNW 35 5 can par mm m mmquot mm mmm Pram m m 01 M2053 9 was mm mnsmnq m Is the polyA tail encoded in the genome DNA g g I 5 3 h DNA 3 r 5 Direction aftranscriptian 7 Primary transcrlpt Synthesis of RNA i Templa e quotanscri noquot strand 0 DNA P I pnmary gt transci t mRNA 5 Release prem NA Exons Synlhesis of ponpeplide 39mmns translation Polypeptide IMPquot are Discarded W 39 MHZ COOH 5quot me inlrons gt mRNA Prueessed transcript snarl exnn mmm slop we 554mm sequence In A3 D5 A5 i amm MA DH 3 Intron Splicing D1 A1 D2 ZDAM nsAs 07 A7 l mu nucleaudes INYRON SEOUENCES REMOVED BV RNA SPLIC NG 5m awn an wvw 5 3 TRANSLATION avumumm Pram m m 01 M2053 9 was mm mnsmnq m 7Sequences removed are called introns or intervening sequences 7Coding sequences anking introns are called eXons 7Exons are not removed and are in the mRNA 7Intron removal is referred to as splicing 7Splicing is mediated by a particle called a spliceosome 7A spliceosome is made of snRNA and protein 7There are five snRNAs in a spliceosome called U1 to U6 7There are as many as 100 proteins in a spliceosome Where do the snRNAs U1 to U6 come from A v r 555259 Upstream 5399quot 539 sue F Do m S39WURAGU lt 39 39 39V IYYYVVVVYYVNYAG 3 Exon lntron Exon 3 539 m 3 39 39 39 VVVYVVVVYVVVNVAGE 339 C I 39 gtYVVVVVVYYVVVNYAG 339 5393 u Spliced R axons A 39lJG 5 to 2 bond mmm xnn sequence wean 7 A u u u u u u u U u u u c a 57 9 AG mar AG u 77777 A 77777 ararorororurornrorornr N or 777 3 A G c c c c c c c c c c c u consensus sequence for branch consensus sequence far 339 splice site 5 splice sue donor sire pom A acceptor silaquot From m m cf MEDD3 1995 Garland Publishing inn What the hell is 5 3 or 2 What are those 5 to 3 linkages 5 to 5 linkage or 2 to 5 linkage Nucleoside Nucleotide OH Base quot0 H 5c 2 o A GIT or c 0 4 H I 2 H Phosphate 0139 H Sugar This group is OH This group is OH in RNA in RNA A D Y 39 Branch Qc i ie Upstream Sphce Slte S39Ite p 39 00mm S39WURAEU v VYYYYVVYVVYVNYAG 3 Exon lntron Exon B l 539 m 339 39 VYYYVYYYYYVVNYAGm 339 C V t gtYYYYYVVYVYYVNYAG 339 539m3 u Spliced R axons A 39l1G 5 to 2 bond If RNA utilized dNTP instead of NTP could you do the splicing reaction CLEAVAGE AND JOINING OF TWO 3 SPLICE SITE EXON SEQUENCES excised intron sequence ft in the form of a lariat 393 I intron RNA will be degraded 1 3 OH in the nucleus snFlNPs will be recycled 5 exon l exon 2 I portion of 3 splice site 5 splice site BBP UZAF texon 1 intron ERES em portion of 5 A 3 a We U1 snFlNP mRNA ESP 439 U2 snRNP lranscrlpt UZAF U1 snRNP U2 SHRNP intrn 3 lUAUS U5 quottriplequotsnRNP U4U6 snRNP U5 snRNP LARIAT FORMATION U1 U4 AND 539 SPLICE SITE CLEAVAGE Iariat U6 snRNP Flgure 6729 pan 1 of 2 Molecular Biology of the Cell 4th Edition Figure 6729 part 2 of2 Molecular Biology of the Cell 4th Edition You have to bring the branch point and the 539 splice site close to each other for the first cleavage to occur and then you have to bring the 539 splice site and the 339 splice site together for the second reaction attachment to occur If you had to design a spliceosome to do that What kind of molecule would it have and What would be its properties U l I xon l CAUUCA exon 1 3amp3 5 3 rearrangement GA U6 BBP exon 2 A 5 3 i 3 rearrangement U a r exon 2 Figure 6 30 part 1 of Z Molecular Biology of the Cell 4th Edition A e B Figure 6 30 pan 2 of 2 Molecular Biology of the Cell 4th Edition preimRNA GA UGUCGAQGAGGGAGUGA CUGCAG preimRNA GAUGUCGAQGAGGGAGUGA CUGCAG Splice out this intron mRNA GAUGUCGACUGCAG 1 Huh preimRNA GA UGUCGAGUGAGGGGUGAAGCUGCAG 2 Splice out the intron What do you need as information quot 3555 quotmm spllce slte slle P v Damquot 539 mumou vvvvwvvvvvavAG I 339 Exon V Intron Exon 3 539mwa39 39VVVVVVVVVVVVNVAG 3 C r vvvvvvvvvvvvuvna 339 Emmy u R 3 Nu 5 to 2 bond A exon 1 exon 2 exon 3 5 339 exon skipping exon 1 exon 3 39 3 Bi exon 1 8X0quot 2 5 quotcryp Cu cfypuc splce splicing slte selection signals portion of exon 2 exon 1 5 Figure 8731 Molecular Biology 01 the Cell 4th Edition preimRNA GAUGUCGAGUGAGGGGUGAAGCUGCAG mRNA GAUGUCGACUGCAG DNA 5 GATGTCGAGTGAGGGGTGAAGCTGCAG 3 C TACAGCTCACTCCCCACTTCGACGTC preimRNA 5 GAUGUC GAGUGAGGGGUGAAGCUGCAG 3 mRNA 5 GAUGUCGACUGCAG 3 urtropomyosin gene 32 31 DNA A i axons introns TRANSCRIPTION SPLICING AND 3 CLEAVAGEPOLYADENYLATION yMVWWlA a39 striated muscle mRNA 5W3 smooth muscle mRNA 5W3 broblast mRNA 539WWMWVh 339 broblast mRNA 3W 3 brain mRNA Figure 5 27 Molecular Biology DI the Cell 41h Edition Al exon 1 exon 2 exon 3 39 3 exon skipping exon 1 exon 3 5393 B exon 1 exon 2 5 3 l I quotcryp on cryptic splice splicing site selection signals portion of exon 2 exon 1 5 3 Figure 6 31 Molecular Biology of the Cell 4th Edition Al NORMAL ADULT BGLOBIN PRIMARY RNA TRANSCRIPT exon exon exon 1 2 intron sequences normal mRNA is formed from three exons IBI SOME SINGLEANUCLEOTIDE CHANGES THAT DESTROY A NORMAL SPLICE SITE CAUSE EXON SKIPPING mRNA with exon 2 missing IC SOMESINGLEVNUCLEOTIDE CHANGES THAT DESTROY NORMAL SPLICE SITES ACTIVATE CRVPTIC SPLICE SITES 3 mRNA with extended exon 3 Di SOME SINGLEANUCLEOTIDE CHANGES THAT CREATE NEW SPLICE SITES CAUSE NEW EXONS TO BE INCORPORATED ge mRNA with extra exon inserted between exon 2 and exon 3 Figure 6735 Molecular Biology DI the Cell 4th Edition Does the cell need to have mechanisms to fixhandle errors in transcription or in splicing 10 Al EUCA RYOTE s cytoplasm nucleus introns exons DNA m tra nscription unit quotprimary ITRANSCHlPTION RNA transcript 5 CAPPING Nuclear Export of mR NA g ligfsucwe NA 9 ADENYLATION mRNA AAAA EXPORT mRNA AAAA lTRANSLATION protein Figure e21 part1 012 Molecular Biology of the Cell Ath Edition initiation factors for intron intron nuclear protein synthesis i l lO lO5 exon 200 107105 pore am nucleotides nucleotides nucleotides RNA complex eIF4G ll ll l EXPO SR 6 factor CBC polyeA binding proteins complexes Figure 6733 Molecular Biology ofthe Cell 4th Edition o L L AAAAAAAAAZDU l O V 039 poyAbinding lob hnRNP pmtems proteins nuclear exportready mRNA restricted proteins 44AAA NUCLEUS cwosm Figure EHtO part1 ol 2 Molecular Biology 0 the Cell 4th Edition gt TRANSLATION I 444150 Figure 6740 part 2 ol 2 Molecular Biology of the Cell 4th Edition A nunlear pore quotexporvreadyquot complex RNA as it emerges from RNA polymerase NUCLEUS CYTOF LASM chromatin TRANSCRIPTION gt l l 200 nm Figure 6 39 Molecular Biology of the Cell 4th Edition 11 What about other RNAs snRNA catal is loop of nucleolar organlzer DNA rRNA gene 45 rRNA precursor proteins involved in processing of rR NA RNAs ribosom gl rge Ibo Pr rg eelw r nualemJquot 39 roteIn cytoplasm gamcle RE m m J quotL r 53 rRNA I NUCLEOLUS rRNA PROCEe we 1 I D6380 Tali rase e Omerase quot e proteins Bulb Iglrrgnea ure telomerase Ef sUbunll CYTOP LAS M TRANSPORT AND FlNAL ASSEMBLV OF RIBOSOMES 408 605 subunit subunit Figure 6747 Molecular Biology of the Cell 4th Edition 5 458 precursor rRNA 3 PPP I OH l l3000 nucleotides l l CHEMICAL MODIFICATION A A A A H tul degraded regions of CLEAVAGE nucleotide sequence 18S rRNA 588 rRNA 28 rRNA q SS rRNA made elsewhere incorporated into incorporated into small ribosomal large ribosomal subunit subunit Figure 6 42 Molecular Biology ofthe Cell14th Edition A C Hk NH I I icy CH HO 7 H36 C Hoe HZC o l ribose ribose Oi Oi GI 3 lt Hi pseudouridine 2 Omethylated nucleotide HNA modifying enzyme precursor rRN RNA modifying enzyme Figure 6 43 Molecular Biology of the Cell 4th Edition tRNAs are made as larger precursors that are processes They also contain introns blue that are excised by a protein endonuclease and the exons are ligated by another protein ligase Different than mRNA splicing Figure 6 54 Molecular Biology of the Cell 4th Edition 12 two methyl groups added to G NNrdimethy G two hydrogens added to U ldihydro U sulfur replaces oxygen in U l4rthiourldinel deamination ofG linusine Figure 955 Molecular Biology of the Cell 41h Edition 13
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