MBB lec 21 eukaryotic DNA

when is dna in most condensed/visible form

metaphase chromosome (X-shaped)

functions of dna being packaged compact

protect

contribute to regulation of replication/transcription

name the levels of dna packaging from most condensed to most simple

-metaphase chromosome (1400nm) (fully condensed during division)

-condensed chromatin (700nm) (folded/compact)

-looped domains (300nm) (looped chromatin for organization)

-chromatin fiber (30nm) (coiled nucleosomes, form thick fiber)

-nucleosomes (11nm) (dna wrapped around histones)(beads on string)

-dna (2nm)

label each dna packaging level

right to left: dna, nucleosomes (circles is histone in middle), chromatin fiber, looped domains, condensed chromatin, metaphase chromatin)

describe the first level of chromatin packaging

Nucleosome

-dna wrapped around histone proteins (bead on string structure)

-aka 11nm fiber

-even tho dna packed, still accessible (important for transcription)

what are histones? are they charged? why? how assembled

-major PROTEIN component of CHROMATIN

-positive charge (rich in Lys and Arg) binds to negative DNA

-assembly requires histone chaperones (highly conserved)

5 types of histones and which one is in nucleosomes

H2A, H2B, H4, H3 (core histones)

H1 (clamps dna wrapped around nucleosome)

how does histone core protect dna in nucleosome

protect from degradation by nuclease

what is the 2nd level of chromatin organization

-30 nm fiber (aka solenoids or chromatin fiber) made by coiled 11nm fibers

-chromosomes in this structure most of time (interphase)

-H1 histone on interior of 30nm fiber

what interactions help pack the 11nm fiber into more compact 30nm fiber

positive N-terminal tail BIND negative sugar-phosphate backbone of neighbouring nucleosomes

what are the more open, less compact regions with less H1. REGIONS that are more accessible to transcription machinery

actively transcribed regions (eu-chromatin)

in solenoids (30nm fiber), what repositions/removes histones to increase dna accessibility

atp-dependent chromatin remodeling complexes

dynamic chromatin structure allows dna to switch between ___ and ___ states. which is active and which isnt

condensed (inactive) and open (active) states

what 2 processes increase accessibility for chromatins dynamic structure to switch between closed/open states (expose dna and allow for transcription replication repair)

1. Chromatin remodeling complexes (use ATP to reposition/remove nucleosomes)

2. Histone-modifying enzymes (ex. histone acetyltransferases HAT and deacetylases) alter histone charge and dna interaction

further compacted 30nm fiber forms the next level, what is it and what is it attached to. this organization allows dna to be compacted up to about ____ fold

looped domains

-attached to a nuclear scaffold (nuclear scaffold proteins like H1, topoisomerase II stabilize high-order structure)

-compacted up to 10k-fold

describe size and appearance of chromatin in interphase vs M phase mitosis

interphase: less condensed (30nm fiber) and exist as induvidual chromosomes

M phase: highly condensed into visible chromosomes

what is the site where dna relication begins during s phase

replication origin

where do splindle fibers attach to pull chromatids apart during cell division

centromere

whats 2 functions of telomeres

1. protect from degradation

2. solve end replication problem

explain the end replication problem and its solution

-dna poly cant start synthesis without rna primer

-at end of lagging strand, final rna primer removed

-PROBLEM: no upstream 3’ OH available so dna cant be filled, leaving primer gap

-RESULT: chromosomes get shorter each replication

-Solution: telomerase extends 3’ end of chromosome

steps: 1. bind to 3’ end of strand overhand and its rna base pairs with telomere dna

2. extends 3’ end (acts as reverse transcriptase, uring rna template to add dna repeats)

3. once extended, primase adds rna primer for dna pol to finish dna synthesis

how are ssdna ends of euk chromosomes protected from degradation

-telomeres form protective T-loop at chromo ends

-3’ overhand folds back and invades dna (D-loop formation) that hides chromo end, reventing it from being recognized as dna damage

-telomere-binding proteins stabilize structure and protect against nucleases