Centromeres and telomeres

Question 1

Centromeres

Answer 1

Constricted region of heterochromatin in each eukaryotic chromosome
Includes spindle attachment site
Ensures correct segregation of chromosomes and prevents chromosome breakage

Question 2

Types of centromeres

Answer 2

Point centromeres (S. cerevisae). 1 microtubule attaches. Specific DNA sequence
Regional centromeres (S. pombe, humans). Repetitive DNA sequences
Dispersed/holocentric (C. elegans)

Epigenetic structures

Question 3

S. cerevisiae centromeres

Answer 3

CEN elements (define centromeres)
CEN elements consist of short, conserved sequences (centromere DNA elements)
CDE-I/III flank AT rich region CDE-II

Question 4

CDE

Answer 4

DNA at CDE-II wound around alternative nucleosome containing Cse4
CBF3 tetrameric protein complex binds CDE-III
Cbf1 homodimer binds CDE-I
Proteins bound at CDE-I/II/III interact with Ctf19 complex (links centromeric complex to kinetochore)
Proteins act as assembly platform for kinetochore

Question 5

Regional centromeres

Answer 5

Core sequence flanked by repetitive DNA
Recruit specific histones (CENP-A) into nucleosomes of central region
Whole structure specified as heterochromatin

Question 6

Formation of centromeric heterochromatin in S. pombe

Answer 6

Centromeric repeats transcribed by RNA pol II
dsRNA formed by convergent transcription and RNA-dependent RNA polymerase (RDRP)
siRNAs produced by dicer and loaded into RNA-induced transcriptional silencing complex (RITS)
RITS binds nascent transcripts and recruits HMT and RDRP
H3K9 methylation recruits HP1
Heterochromatin spreading

Question 7

Human centromere

Answer 7

Long arrays of simple tandem repeats
Higher order repeat of alpha-I satellite DNA (copied thousands of times to form chromatin)
Pericentromere is divergent repetitive sequences and retrotransposons (alpha-II satellite DNA)

Question 8

Telomere structure

Answer 8

TTAGGG repeat sequence in vertebrates
G rich 5'-3' towards telomere
C rich 5'-3' towards centromere
Variable length
G-rich 3' ss tail results from post-replicative processing of C-rich strand to remove primers and iDNA

Question 9

Chromosome capping

Answer 9

3’ single stranded G-rich strand forms T-loop
Displaces complementary sequence in an upstream region of the telomere
Catalysed by TRF2

Question 10

Telomerase

Answer 10

Only active in stem and cancer cells

Ribonucleoprotein complex composed of telomerase reverse transcriptase (TERT) and RNA template (TERC)

Question 11

Shelterin

Answer 11

Coats telomeres and protects from DNA damage repair pathways
Regulates telomere length
Inhibits telomerase

Question 12

Shelterin complex

Answer 12

Telomeric ds repeat binding factors 1/2 (TRF1/2)
TRF2-interacting factor repressor and activator protein 1 (Rap1)
Telomeric protection factor (POT1)
Bridging molecules TIN2 and TPP1

Question 13

Telomere heterochromatin

Answer 13

Resembles pericentric chromatin
H3K9, H4K20 methylation
HP1 binding (reader)
Telomeric DNA cannot be methylated (no CpG)
Sub-telomeric DNA methylated
May regulate telomere length

Question 14

TERRA RNAs

Answer 14

Telomere repeat containing RNA (lncRNA)
5’-UUAGGG-3’ repeats at 3’ end
Base pairs with C or G-rich strand of telomeres
Promotes recruitment of enzymes (heterochromatin formation and DNA replication)
Control telomere length (inhibit telomerase)

Question 15

Importance of telomeres

Answer 15

Senescence (limiting lifespan of normal cells). Limits mutation accumulation in dividing cells
Promote genome stability