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Lecture 13 Notes

by: AnnaClippinger

Lecture 13 Notes BIOL-L211 2521

GPA 3.3
Molecular Biology
Megan Dunn

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Molecular Biology
Megan Dunn
Class Notes
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This 16 page Class Notes was uploaded by AnnaClippinger on Monday October 19, 2015. The Class Notes belongs to BIOL-L211 2521 at Indiana University taught by Megan Dunn in Summer 2015. Since its upload, it has received 25 views. For similar materials see Molecular Biology in Biology at Indiana University.


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Date Created: 10/19/15
erdai Lecture 13 ct 13 2015 39 Last Time 39 Transcription in Prokaryotes Transcriptional Regulation in Prokaryotes 39 Today Transcriptional Regulation in Prokaryotes aqu MS Mn MEIquot1 3 39 Examples of P rokaryo lc Gene Expression 1 pg Mg quot W W Next Time 39 Problem Set 2 Due Transcriptional regulation We will discuss two general examples 2 Decrease transcription of a gene Examples DNA binding proteins in the lac operon andl in quorurm sensing Sigma factors review air Proteins that bind speci c promoter sequences and inst rn ct RNA p lymerese where to begin transcription One sigma factor is called 1 39 omdirects transcription of Mun genes quot1 CE 41mm wl ESSEnn uI or Rout Inttummy Alternonve srgmafoctors if E coil nu rtmm 0quot wl ormr mum Nbuni N f N Difiertnr 0 factor I ELUQNI39E durum lJr 0 Mat E r shaman housekeepmg a 9 life rmi gem Wits can be E w mud EN 3an l m Enema A way of coordinately regulating hundreds of genes of similar function throughout the genome rgtarvation l TszmmW alierna w 6 futrm t a Bind m g 2 3339 agggrtnyc s d ff F f ALS39 quotN VE lquot8 l9h 039 In di Via uul Nuclco tide ll nfll Sti ll REEm mm In or has Alternative sigma fctors Organisms can control the order in which genes are transcribed Example Lytic bacteriophage infects bacterium which Infected bacterium replicates phage DNA transcribes phage genes translates hage proteins EWPmmn t ath 0 n walled lo urtmq a Igttrnatiw J lq ma in rm R N h P A t 3 man u l 1 HUUM RECP39I El fatter M ma w I teeter v aquot factor premeter prame ert Fig 1315 quotLatequot infection genes promote 39n K Ea rly infection genes including gene for alternative tr fatter lm i W W Multan 39 t5 Home t i slut a lawn POW frunmmnn a Alterna ve ngma factors Middle infectien genes including gene for a Itern ati ve e fa cter j 40 E 3quotquot P l W1 N Man in I JN UVi HEnt mm E39nurrmmenl Quasi May activate transcription ef genes for ribesemes cell wall biesynthesis RNA polymerase core cell division etc Alternative sigma factors WWW cm htqmrm RUn law on quotnumerals a 5 quot l 6 9 6 z w quot W U b quot l I Care 273 m t illty 7 May activate tra script ian of genes forquot flagellum biosvnthesis Alternative sigma factors Example IS 1mm l Milan r activate transcriiptimn uf genes for Detoxifying enzvmes transporters etc Molecular biology in the news The Economist May 2011 5 3 r0 H OH Emm m m Occu rs in re lles jelly sh etc A w cmrlu quot n a phatoprotein Discovered in a quotelllv sh Aequoren Vic orln sf ruuure Ma mm Map rumtqphnreln39ctnkr Braan Fluarescent Protein EeIaaSheel 7 Fnljygpe tide VVStrucur a Makes 23 V HEM EBITEIquot aan Ghmmoplmre Figure 1 9 uaaA GF P and differently COIDFEd variants I m like can be Ll m mm m gennm l ecula biology if s an organism Protein locallza on in a bacterial cell Regrowth 3 UR lfQE 1 m9 l l 39l g I71quot 170 P W tam i salamander limbs muzv f39TDr rcglmwii m wluwpunsmn 0 Jim Two diflerent proteins labelecl with CFP and YFP i I 39 quot Ruppy the fr 7 glowlng puppy VIN ll Ruppy s pupples The figChe operon governs oge ar biosynthesis B subi39 is oche operorr Comprised 01 32 gemes 27 M long Promoter P oce I Z A V g I 7 EXP i M S d vib rm atrium Bi RNA j court 139 H Mercer rnq Sigma mm r iPrerre 39 che ope m n i ii 39 i i i g 1 i I i g i r ch28 in ll diet 2 i f 3 I e N 1 A IVE I if I NJ iii rim rimquot ml 391 yiriquot iii rial 51 WA er the MEN rim mu rim lm rim th irt Cheri chef rr39y mm I Mung USPS 0pm Filament Hook cap rFig re HID l39amenil mp limkslrrrgzlum HE E 39 3K 7 H3911 lrainemstmtmre 39 FigKjrindlen preterm Slater Proton cheme Moi I Motorpretevaum 1quot Fig L junction protein 3353 Note colorcoded genes Iii Hr quotm mquotmr opean correspond to proteins Hgfrodslmmrm 39 FI39Ii wmdi structure 1er rod simrimo Secretion Secreiian apparatus Fil Secreition appammsh HIP Seoretlmappara msFlinn V i i l n JJ39Ierbaga lrmg 1 7 7 I i r 7 7 r SEEIQHG39quot appat ws r FIIR V 7 i 39 llieumkngw I I 5 l I r r 7 H Encreiiuniped ciwa im 39 39l v 4 39 Swarm smel ciwe I 39lilli we 39539 quotml r chaperonec iilroirarlli 39 39 Mam COMM l p 39 m quotbl Q flaw E i n is unci er a g Mukhe and Keernsr 2014 Ann Rev Genet The alternative sigma factor oD controls transcription of the ogellin gene gem ayChe operon i i g l g i i i x l i V 7 Chang chew chef Her e iii i rift iiiM rid rm yirF rm yi mil film er the NM uquot Mi rim Him WM hF rm che fret 5er 5er i i D if quot be pro poled rrwurd win 0D e 7 e ncoci i rig a n 39 1quot 39i39 or used To direct rm no or me s the protein tirat is useci to build 19 T he i mqeum mm H T in The quotI F gis 41 i C2 he J 12 l r l 739 Vr h W v V A I 7 W n I i 7 I r rEf39l I i r H T r 39 39 w l 5 Hm Huh liii FM 5rle Hm ail yin ll linti Hitquot the rim lill39 ll Mail illlll HM 1 th Mir than metquot angli swrii l l i l 7gt 72 J r u a t The finthe eperon is too long for the ENTIRE transcript to be made EVERY ti rne Semetimes 51590 is not transcribed before transcription terminates lithis ihap em 0 protein would NOT be made 77 a 39 r v an t tflaqtllin gt m wn39um ELF bt rrnscrrmd i if a D lS quot quot rnmcrilatd w Flaqcum wl II ampT be a wru wv affect How was this experimentally determined a 0330 L ec vitv ofth ii itrwas measured by fluorescence microscopy M S O p a l MI Built UN MO l CLll r39 techniau 13 r i ir i i 391 i7 a l 7 LENJ li lll i739l quot139 f ettt39 li0t MH I NW Mimi in quotwill RE SQGquot0W ample Green nerescent prete ianFPl Emmi WE Him t r tithe N TRANCEquot BDEWM Your Fm39muler Di 7 ni me if w w lNKInWinqwn laminae arr mama P age in Para i P a m age iancl gfp are each transcribed i from their own promoters ageiiin gfp neither are in operons Rama ting W39HBLE M m Mar Jusr adding P egeiiin 39l l 39 la 31 l l 7 Ill l39tir for 1W Eloqtl39lm Elena l quot grp the 9 p the Promoter is u SW u M W w W Transcriptional fusions Can use the activity of th it an indication of whether or not your promoter of interest is activated for transcription i The gfp gene is transcribed from the q Ytt 2 ogeiiin promoter what cowl 39i W 39i runad39trwn on grp Flour CHIS bl ii quot Prlogeum39 5 P 395 transcribed 1 G1R39FE N 39 R f V39 C D L URL ESS39 ELL 5 quotl39TthcriPti rim 39 Cell is green Cell is NOT green quot1P HgEiiinhgfp is transcribed 1N9 anis F39ilWDl Df E ge l n gfp JeThc 39539 Q 39 Pmmnlir l w f fo r tr nScriPti on l39m WiPii quoti 15 Becausebis at the of a rlt 1211quot f it is transcribed infrequently a a is required to gene transcriptio39 N Ul TM 1 MM gill in END I is located Closer to a Therefo re f more Could be made and Tram cripn on Bi minim MM from he pm momv wan manned Artificially rnove ii different positions throughout the i New positions indicated by black arrows Blpid Gig D glint 11 DE MM Ef i j 39l39NHQLH W Begrmm a loamy0n a a is i a a rhe eemm w E l 7 I I dieE chew cried TH 7 7 a I r 3 right rigr mi r r39 iii Id m M yixF Hm M yizi Hit me Hitquot Me mil in Wm WM th mm cheat ahaC 1ng ma A A f l 39 WW W W or quotii iitllm 16 g E a o hgogemm E it 3 chew ch91 r rquot1 x 39 DY oE jri39 it rat31 1 I 3 7 r 7 r r r I I L v V gt 1 7 I I A gt A mo rm fr nJ Jrer MK g l the Mill rim Hm Imr Him HM h F h I39m391 Pref sin swr A A A native si39gID i V 1 V 7 7 original position ionka 1 trammfptmndi Fusions Pmm reigD iF sigD figE sigD in MM h mnwm It gnu 11w a mi tweu 1mg milk J 39 Remember A green celi indicates A for transcriptio open 7 7 r mm n V rneosrgo oheC sigD V netirefr39gD ringmm H LN M U R WEMN T When qrgb is t m Fr39u39nro r the Optimu P oge inagfp m Fot39HLQI S is SL1 pp Dimcot Cozy and Kearns 2011 Moi Microbioii 1 Transcriptional regulation 39 refers to mechanisms by which gene expression is ed in response to the environment We wiil discuss two general exampies 39 Decrease transcription of a gene loo operon and quorum sensing 18 Acn39vo tors and repressors RNA perimete I I 3 r 39 RNAP binds promoters weakly inmates 13mm irvm Hamiin 0i immr39iii basal ievel ot39 39anscri p lion ii Hiiiij 39 39 aetiuatoe opereler binding site Phiquotme binds to a n E sequence Block RNHP from my th mmuler l hm bits Hamwrpnon oi Mene 5 0 C179 as n pnqsrcn l rr rem pmemt w Blake trm39mi on binds a sequence away from the promoter Recruit RNAP form pramurer Hnn was of Lemon am 9mg 339 1 Hue anew wt prawn wt can nrui n DNM RN A WM mm min togaMar 19 no transcription The iac operon in E coli ii amewit 393 din promm U i CAP site in eramr 7 L l prornTOEr in response to environmental L Jnditionsr expression of the 39 i Increased 31 unanimity OR Re Wm W1 Rtpreno r 2D The lac operon in E coli is dedvlcated Wreaking dovr it ll i cl mu l min alumsL 39i fall Mam 1luwgc l9 We Pit Pena loadfo m r m Emu aw meme W0 39 0 E coli controls expressiotglrgf hg Pilng glmogb operon based on fh Ur bsmm 0le lllLl39cG a Lactose Galactme Elmcase ll JUQl alum Wm M Repwased RNA polymerase a The promoter for the lac nperon lever wealk Very low levels df transcription occur l e I I u M lifa gt r 7 7 7 basal level in q 39 nftranscriplinn r A 7 ele Demeter 7 pmmuler F r 39 lfh llt l Ir 3939 21 The lac operan in E coli m 39 yielddag 39lrwui 5 c 7 3 l 7 El l 2 MW Wife t m l N HP To me pm mu 5 Tmmcrlpl39l an all lac game ad valid 0NL w hm Ldtl oee re lm r n vaellvated level V of hanscrlplion r quotquot 39 39 ep essor binds to an aperator PT EWle mm M l Pl l an lllc QEHEG lumem 1 r f v mjlei s 39 wquot W M1 Lula r u 5 r quot 2 11395quot 2139 quot llquot ll 39 a a 1 Y quotnll39nr gquotr4 i39 39iu i v quotu 39 339 l 39 399 EF E i Ile F quotiii Lil Lilla av L yjy iv lg 111 ue jl ifjx jf i lrl VI no transcription CAP sill Operator promoter 22 CAP activator and repressor bind speci c regions CAP binds to the CAPbinding site 7 Recruit RN n to rm promoter Repressor binds the Operator Phusicmw pawmm RN H P From birding the pram mix Gr I p P bziii ingsitg 7 DNA severed by RNA pe iymerase 7 39 H Ax 4LLL 7717 4 A 39 n9 i 3 mRNA 5 quot M ecam feeeneeTitaniaCanere neriesche39dceneemermemin zeemrennet1ieeneemeeeeweenieeaemeeemmmnrmemnmgrammesv at Teasemaneeecncecmeeeaeeeaeemeeeeeeeeemeeemeemenameeereeammeemggncemmcmweeeeeneeeemaaeeeeeeeeemeeeeea 35 10 1 Figi lea DNA attered by represent Tm Optm i r mrtri i The PM 0 W 23 Quorum sensing quotThe minimai number of of cers and members of a committee or organization usuaiiv a majorityI who must be present for valid transaction of biusinesequot in i used by bacteria to communicate with each miter byng detecting and respending to Chemical signals quotEMU quotN n 039 h a MIC mdi mm THEW P pum on 61mm RtePDnd W i hangar m amt expme 1 w mm Ummh hum mm m mg was lit a mum wraith fitme need to new 39 Gene expression Gene expressien Gene expression OFF OFF ON Luxl synthesizes euteinducer in Vibrio scheri Cells sense an extracellular signaling molecule called 6 r 39 Ami n riduwr i9 Madt 3m a Pm rem calm o O a AUloindllll Autoinduvczer diffuses out of the cell and into tine environment 25 a transcriptiprlall activator V BUT if ream m9 0i high lievcl CDHCCHTM HOI39I 0F AWI D l Mum in ma BNA 39 Ntfd mm th aid 1 0 Gillian mine mi viion Accumulatinglevels of 1 UN 39l mime r O O 0 39 9 Autoinduar Bir gluxR o 0 f O o O H m 7 f CE Tm Mari Ph on 39Uhl l39 25 Celi toce communication critical cell clerisit ir quot a g gene expressren cell densrty autoihducer concentration time Mlhl WM thrashed r Auto uln ther accumulates in Droporlrnh wl all eltnsihl At the critical cell density auteiad ucer concentration reaches a threshold to activate gene expression 2 Quorum sensing sighals are specr c g u39 t 39r ra c u A s 12 AV hL LIH arquotcsquot a EA g H A a Bd HSL Sin115a Email rr39eaj39 15ml H Purimmm azmghm39 Will AZWMMM squrli39ddlai ml 0H t H gt M o Emma04am 39 lil 39 a m t tmwm ll Cl Hr N 5 39qw n ms SqII Ill 0 Wamo m m39lm ebream Evil Benderpm our n l den 7th 0 El H W 39 Mae EHSL H 39 It ui rmgme I En nEermm 39i Egan H El caretmun subsp xanJCati til I a o H gwleru aucrdggia ww E331 Each species at bacteria makes a W in calstmmr r quotm l m P rcu39tr MES H la structurally unique autoihducer V H EECVi P D l39l m V tamcg m fawn hmcm D H 39 r E D vmumriemmna i e ll 0 WI m u n icati on Mitigate D l 0 NmdHSL 39 Ham39sEater pr rm n39uwerl K H 0 DH CI H 0 amp39a39hlquotdmw39m4 HSL l H R kabim qaminammwm U 23 Querum sensing example Bioiumi nescence m the presence of autoinducer Lm R ammi fhetmmc pl mn 0F Buio lummes tm gmee i RNWIS L H l T C l r o o l a u 29 Quorum sensing exampie Bfoluminescence The bacteriu m regulates the expression of a I 3zf efe bv quorum sensing 7 LUCI mm mm Elm m Eml h ll39t gm 3i mu39mmescem Autoinducer A U GHTI I V scheri grown in liquid and on a petri prl ate 30 Quorum sensing example Bioluminescence 392 f0 rms a relationship with a small squid called 1 m a quotu l39 L r The bacteria live within the squid s light organ Blol39ulminesmnu via Quorumrte Ming Samaria pmwdEW Wm tr l uml n tion quotquot furme mind No a hadow39l 1 mm Uitd m pm Harlan MEthtlm39Q39m Wing Moonliqm l u illuminam lg lith Rumwl 31 From villain musk 1 an Shadow apredatm L A MT isquot me tin tam thmru Review l Mlmbiulnay


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