Chromatin Basics

download Chromatin Basics

of 12

Transcript of Chromatin Basics

  • 8/20/2019 Chromatin Basics

    1/31

    This slide series has been used for teaching to students of the Montpellier University in 2004.

    It explains basic features of the folding of DN in nucleoso!es and chro!atin fibers" and

    discusses the regulatory roles of post#translational histone !odificatons

    The !aterial displayed is inspired fro! several sources. The literature used is cited $ithin

    the slides. Moreover" several slides %indicated by the &'( logo) have been copied or !odified

    fro! &acob (aterborg" University of Missouri" *ansas +ity To see his chro!atin !aterial in

    full" see the $ebsite, http,--sbs.u!c.edu-$aterborg-

  • 8/20/2019 Chromatin Basics

    2/31

    3-99&'(

    DNA compactioncompaction  in a human nucleus

    1bp (0.3nm)

    10,000 nm

    30nm

    11 nm

    Compaction of !" by hi#tone# Compaction by chromo#ome #caffo$% & n'c$ear matri

  • 8/20/2019 Chromatin Basics

    3/31

    3-99&'(

    Nuclear # chro!oso!e co!paction

    Compaction by chromo#ome #caffo$% & n'c$ear matri

    ra%ia$ $oop

    chro#omo#ome mo%e$

    1µm

       1   0     µ  m

    chromatidchromatid

      m   i   t  o   t   i  c  c   h  r  o  m  o

      s  o  m  e

    + 2M NaCl

    histones

    + 2M NaCl

    histones

    Mitosis!" $oop#

  • 8/20/2019 Chromatin Basics

    4/31

    3-99&'(

    'I/TN1/are

     highly consered!

    small! basic proteins

    H1

    H*

    H"

    H3

    H+

    he$i

    ariab$e

    con#ere%

    "istone acetylation

    i# a reer#ib$e mo%ification

    of $y#ine# in the !-termini

    of the core hi#tone#.  #esult/ re%'ce% bin%ing to !"/ %e#tabi$iation of chromatin

    in2er hi#tone

    Core hi#tone#

    !

  • 8/20/2019 Chromatin Basics

    5/313-99&'(

    +ore 'istones

    he ba#ic #tr'ct're of "  core hi#tone# i# the #ame/  1 $ong hy%rophobic a$pha-he$i,

      bor%ere% by/  #hort hy%rophobic a$pha he$ice#

      that form pair#/  H" - H* an% H3 - H+  4hich interact.

    #e$erences% Moudrianakis et al. PNAS 88, 10138 (1991);

      PNAS 90, 10489 (1993); PNAS 92, 11170 (199)

    The histone#fold

  • 8/20/2019 Chromatin Basics

    6/313-99&'(

    'istone octa!er asse!bly

    "3-"&

    tetramer  "2A-"2'

    dimer 

    "istone

    octamer 

  • 8/20/2019 Chromatin Basics

    7/31

    3-99&'(

    The Nucleoso!e as the funda!entalchro!atin unit

    5ig're, co'rte#y from im

    6ichmon%, 7H 8ric:4iter$an%

  • 8/20/2019 Chromatin Basics

    8/31

    3-99&'(

    / 1+;-1+9 bp !" in a 1.;< t'rn# of a

    f$at, $eft-han%e% #'perhe$i/ one p#e'%o t4ofo$% ai# centere% atthe =%ya%> (reference 0 he$ica$ t'rn#)

    / one ba#e-pair preci#e$y at the %ya%/ #harp ben%# at ? 1.< an% ? +-< t'rn#/ Hi#tone-fo% %omain# organie 11 bp

    of !". he !" i# bo'n% at 10 bp

    intera$# thro'gh many contact#,

    inc$'%ing penetration of arginine# at a$$

    1+ minor grooe# facing the proteincore/ he grooe# from neighboring !"

    t'rn# $ine 'p@ forming channe$#/ H3 an% H* !-termini eit one of

    the#e channe$# eery 0bp./ he H+ tai$ e#tab$i#he# contact# 4ith

    the net core partic$e.

     Nucleosome features

    H* H+H3H"

  • 8/20/2019 Chromatin Basics

    9/31

    3-99&'(

    'istone octa!er organies 34 bp of DN

    /  7ach core hi#tone %imer 

      ha# ; !" bin%ing #'rface#

      that organie 3 !" t'rn#@/  he hi#tone octamer 

      organie# 1+< bp of !"

      in 1 3&+ he$ica$ t'rn of !"+A nm of !" pac2age% in a %i#c of ; 11nm

    B ; nm

       B

       1   1

      n  m

       

  • 8/20/2019 Chromatin Basics

    10/31

    3-99&'(

    (here are the N#ter!ini of the core histones 5

    !u"er, Mader,#i$%&ond, Sar"ent ' #i$%&ond 

    Nature 3(9, 2120 (199)) 6uestion 3: What is the function of histone N-termini ?

    6uestion 2: Are all N-termini functionally equivalent ?

  • 8/20/2019 Chromatin Basics

    11/31

    3-99&'(

    "1*" g$ob'$ar protein %omain

    7iner histone '3

    ,iner histone "1 organi.es e/iting DNA up to 1( - 200 bp

    H1 #tabi$ie# interaction bet4een n'c$eo#ome# in compacte% chromatin.

    On the OUTSIDE of the DNA gyres ? Asymmetric between the DNA gyres ?*%ou et al. Nature 39! +0 (199() +ole et al. 4cience 2)&! 14 (199)

  • 8/20/2019 Chromatin Basics

    12/31

    3-99&'(

    Chromatosome

    mM

    1 mM

    7iner 'istone and histone ter!ini

    control liner DN entry-exit of chro!atoso!e in chro!atin fiber.

    Core histone octamer + 1 Liner !istone + " f#$$ t#rns of DNA %1&' b()

    H3

    H1

    !

    C

    H3

    *%ou, -er$%&an, #a&akris%nan, ra/ers,

    Mulder&ans Nature 39, +0 (199()

     An, !eua, /an olde, *latano/a

    5NA4 9, 339; (199()

  • 8/20/2019 Chromatin Basics

    13/31

    3-99&'(

    +hro!atin

    fibers

    + charged N termini

    (bin% !" on neigboring

    n'c$eo#ome#)

    highly acetylat

    core histon

    (e#pecia$$y H3 an% H

    30 nm

    chromatin $iber 

    11 nm

    bead

    /  "67" leel o$ histone "1 /  #educed leel o$ histone "1

    /  N8 gene transcription /  7ene transcription possible

  • 8/20/2019 Chromatin Basics

    14/31

    'istone !odifications

    Turner (22!" Cell 111# 2$%

    D

  • 8/20/2019 Chromatin Basics

    15/31

    3-99&'(

    cetylation of conserved lysines

    H+ !-termin'#

    H3 !-termin'#

     "c-:-E-6-E--E-E--E--E--E-E-"--6-H-6-K-F--6--? ? ? ? ? ?? ? ? ?

     "c or MeAcAcAcAc

    5   8 12   16 20

     "-6---G--"-6--:--E-E--"--6--G--"---"-"-6--:-"--

    Me "c or Me

    Ac Ac Ac Ac

    ?? ? ? ? ? ? ??

    4   9   14   18   23   27 

    The N#ter!ini of histones '4 and '8" and their acetylation patterns" are absolutely conserved.

    'ysine

    ε

    # N-Acetyl-'ysine

    'T %'istone

    cetyl#Transferase)'istone

    Deacetylasere/ersile rea$tions

    8

    N

    C

    C C

    C

    C

    CN+α   β

    γ δ

    ε5

    88

    8

    5

    8

    -

    DN

    bacbone

    binding

    no NA

     )in*in+8

    N

    C

    C C

    C

    C

    C

    8

    C

    C

     Acety$*CoA

    CoA5-

    --

    --

    ---

    --

    --

    --

    --

    --

    -

    -

    -

    -

  • 8/20/2019 Chromatin Basics

    16/31

    3-99&'(

    e$ectro%e

    e$ectro%e

    gel type% 4D4

    "&

    10 aa

    "3

    13< aa

    "3

    "&

    +

     : 

    9el 1lectrophoresis of 'istones

    A; A;<

    4eparation by% si.e si.e +charge

    si.e + charge +hydrophobicity

    "3

    "&

    "32

    "&

    "31

    01

    3

    +

    0

    3

    +

    1

    "&

    10

    ++ +

    13<

    ++ +

    "3

    0

    3

    +

    1

    + +

     :   : 

    7el scans a$fa$fa hi#tone# (+ateror", 90s)

    "&

    10

    ++ +

         3

    13<

    ++ +

    "31

         3

       3   3   3

       3

    13<

    ++ +

    "32

       3     3

    3     

    013

    +1

    3

    +

    0

  • 8/20/2019 Chromatin Basics

    17/31

    3-99&'(

     Almost  ll cetylation is Dyna!ic

    +ateror"   = 'iol Chem 2)3, D;0 (199()

    7amp$e alga

    Ch$amyomonas

    reinhartii 

    >

    0

    10

    1

    20

    2

    0

    10

    1

    20

    2

    0

    1

    3

    +

    <

    0

    1

    3

    +

    <

    H3 H4

    proteinprotein

    ,?@?, o$ Ac,ysturnoer 

    ,?@?, o$ Ac,ysturnoer 

       S   t  e  a   d  y  -  s   t  a   t  e   [   *   A  c   ]   l  a   b  e   l   i  n

      g

    ee$ of hi#tone acety$ation 32 > o$ "3  1 > o$ "&

     "erage $ee$ of acety$ation 2& Ac,ys*"3  1 Ac,ys*"& 

     "erage "cy# t'rnoer I 19 ±0.+ min  (H3) 3 ±1.1 min (H+)

     "cy# t'rnoer I (min) .A 1.A 1.A 1.; 1.<

    :raction of acetylated histone 100 92 10& 10 (2 39 ( (1 100 ) >

      sub;ect to c7ys turnover,  300 ±12 < of '8  ±, < of '4

    1 " , - . 1 " , - . /AcLys0histone

    %=>

  • 8/20/2019 Chromatin Basics

    18/31

    3-99&'(

    cetylation of +hro!atin Do!ains

    ees, laton, %orne, rane#oinson  ?M'8 = 13, 1A3 (199&)

    1xa!ple, the chicen *g$obin gene omain

    l "igh leels o$ chromatin acetylation, acro## comp$ete chromatin %omain#

    (!" $oop#), in%'ce# chromatin change# %etecte% a# =general DNase 6 sensitiity>l Within the#e chromatin %omain#, at f'nctiona$ gene# or tran#cription factor#,

    the chromatin #tr'ct're i# interr'pte% by #ma$$ =DNase 6 hypersensitie sites>

    %omain

    bo'n%ary

    %omain

    bo'n%ary

    βρ βH β " βε

    0 10 20 30 b

      !a#e

      hyper-#en#itie #ite

      β-g$obin gene#

      Ac-,ys antibody

      nucleosome ppt

    5NA loo6 do&ain

    7eneral DNase 6

      sensitiity

    Contro$

    inactie gene

    chicken

    ovalbumin

    0 1 2DNase 6

    (U&m$)

       D   N   A  r  e  m  a   i  n   i  n  g

    DNase 6

  • 8/20/2019 Chromatin Basics

    19/31

    3-99&'(

    cetylation at ?ro!oters

    Transcritional

    ./0./NThyroid Hormone Receptor ea&6le o 5NAindin" rans$ri6tion a$tor 

    co-repressor 

  • 8/20/2019 Chromatin Basics

    20/31

    3-99&'(

    Deacetylation of +hro!atinTranscri(tiona$ Si$encing 2*chromosome Inacti3ation

    Nan et al. MolCell'iol 1, +1+ (199)@ Cell ((, 1 (199))@ ones et al. Nat7enet 19, 1AD (199()

    l meC CpE !" mo%ification i# ob#ere% in

    repre##e% gene# an% inactiate% L chromo#ome#

    l meC CpE-methy$ation i# maintaine% after !"

    rep$ication by Maintenance Methy$a#e action on

    hemi-methy$ate% !"

    l meC bin%# tran#criptiona$ repre##or MeC52 

    (Methy$C-bin%ing 5rotein-2)

    l MeC52 bin%# :in3 4ith #5D3 hi#tone %eacety$a#e

    + +"ypo-acetylated repre##e% chromatin fiber 

    transcriptional repressor 

    Histone eacetylase

    4in3

    #5D3

    MeC52

     ..pCpEp..3 me

    ≡       ≡     3 ..pEpCp.. me

    co-repressor 

    Cp7 methylation

    'i t M th l ti

  • 8/20/2019 Chromatin Basics

    21/31

    C C

    'istone Methylation

    'ysine

    ε

    # N-monomethyl-'ysine

    'MT %'istone

    Methyl#Transferase)'istone

    de!ethylase5

    8

    N

    C

    C C

    C

    C

    CN+

    α   β

    γ 

    δ

    ε

    8

    N

    C

    C C

    C

    C

    C

    ε

    C

    'MT %'istone

    Methyl#Transferase)

    /#adenosyl!ethyionine

    8

    N

    C

    C C

    C

    C

    C

    ε

    C

    ε

    C

    'MT %'istoneMethyl#Transferase)

    8

    N

    C C

    C

    C

    ε

    C

    ε

    C

    ε# N-*imethyl-'ysine

    # N-trimethyl-'ysine

    DN bacbone

    binding !ay not bestrongly affected" but

    specific proteins !ay

    recognie these

    !odifications

    N+

    N+

    N+

    /#adenosyl!ethyionine

    /#adenosyl!ethyionine

    "istones can be methylated at lysines or arginines ?/ample% "3 9 methylation

    'istone !ethylation , 'istone !ethyltransferases

  • 8/20/2019 Chromatin Basics

    22/31

    'istone !ethylation , 'istone !ethyltransferases

    7nhancer of 8e#te KD:i$encing of e'chromatic gene#

    Mo%ifie% from

    achner an% Jen'

    (00). C'rrent N

    Ce$$ *io$. 1+, A;

    Multiple roles of '8 *@ !ethylation

  • 8/20/2019 Chromatin Basics

    23/31

    Multiple roles of '8 *@ !ethylation

    In contrast to histone acetylation" histone !ethylation is /TA71. This !aes of this

    !ar  a candidate for inheritance of chro!atin states.

    N. $rassa 7nhancer of

    e#te

    :. pombe to man

    ri-@ %i-@ mono-HMa#e#O

    (ri-met KD)

    an% tri- met K9

    Mo%ifie% fromachner an% Jen'4e

    (00). C'rrent Npin. C*io$. 1+, A;-9A

    'eterochro!atin

  • 8/20/2019 Chromatin Basics

    24/31

    'eterochro!atin

    irst discoered in Drosophila in genetic studies o$ chromosome rearrangements

    "eterochromatin induces gene silencing

    :ingh, . *. (199+). Mo$ec'$ar mechani#m# of ce$$'$ar %etermination their re$ation to chromatin #tr'ct're an% parenta$ imprinting. J Ce$$ :ci  107 , ;

  • 8/20/2019 Chromatin Basics

    25/31

    9enes involved in heterochro!atin

    :ingh, . *. (199+). Mo$ec'$ar mechani#m# of ce$$'$ar %etermination their re$ation to chromatin #tr'ct're an% parenta$ imprinting. J Ce$$ :ci  107 , ;

  • 8/20/2019 Chromatin Basics

    26/31

    H3

    6& 61,

    Ac Ac

    H36& 61,

    HAC

    H36& 61,

    7ethylation of 6& (Clr,8 uvar3&!

    7e

    H36& 61,

    7e9i .ecruitment of 9i or H0-1

    'eterochro!atin for!ation involves histone '8 lysine @ !ethylation by

    /u%var)8#@ and recruit!ent of '?#3

    .ecruitment of more Clr, (uvar3&! molecul )y 9i (H0-1!

    pen chro!atin

    +ondensed chro!atin

    1pigenetic regulation of centro!ere function and BN

  • 8/20/2019 Chromatin Basics

    27/31

    1pigenetic regulation of centro!ere function and BN

    Central *omain uter reeatsuter reeats

    " 0om)e Centromere

    9i

    6& 7e

     Nucleosomes

    9i

    6& 7e

     Nucleosomes*s.NAs

    This structure requires roteins that are resonsi)le for the henomenon ofBN interference

    -; .NA interference is +ene silencin+ me*iate* )y short .NA molecules ro*uce* from

    *ou)le stran*e* .NA (*s.NA!"

    -; hort .NAs are ro*uce* )y cleava+e of *s.NA )y the nuclease calle* icer" icer

     ro*uces short *s.NA calle* si.NA *ule

  • 8/20/2019 Chromatin Basics

    28/31

    pposing functions of '8 *@ versus *4 !ethylation

    achner an% Jen'4ein. (00) C'rrent Npin. Ce$$ *io$. 1+, A

    functional lin bet$een histone '8 !ethylation and

  • 8/20/2019 Chromatin Basics

    29/31

    functional lin bet$een histone '8 !ethylation and

    DN cytosine !ethylation in Arabio(sis tha$iana

    '?35

    Mo%ifie% from achner an% Jen'4ein. (00) C'rrent Npin. Ce$$ *io$. 1+,

    Multiple biological pheno!ena involving histone !ethylation

  • 8/20/2019 Chromatin Basics

    30/31

    /(=!

    Multiple biological pheno!ena involving histone !ethylation

    H3-KD methy$ation an%

    Mo%ifie% from achner an% Jen'4ein. (00) C'rrent Npin. Ce$$ *io$. 1+,

    an% H3-KD methy$ation

    4 d ti

  • 8/20/2019 Chromatin Basics

    31/31

    4ummary and perspecties

    Chromatin i# pac2age% in a hierarchy of #tr'ct're#. 7ach of the#e

    $ee$# of pac2aging ha# reg'$atory ro$e# in the genome

    he $ee$ of pac2aging 4e 2no4 be#t, a$#o than2# to formi%ab$e

    too$# techno$ogy %ee$opment in the recent year#, i# the

    n'c$eo#ome.

    n partic'$ar, po#t-tran#$ationa$ hi#tone mo%ification# p$ay 2ey ro$e#

    in reg'$ation of genome f'nction, an% the combinatoria$ po4er of

    the#e mo%ification# i# on$y beginning to be 'nrae$e%.

    5't're cha$$enge# 4i$$ be to %ecrypt the %etai$e% meaning of the#e

    mo%ification#, an% to 'n%er#tan% the ro$e# of the higher or%er $ee$#

    of chromatin an% chromo#ome fo$%ing