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BIOL 200 > KH8 > Flashcards

KH8 Flashcards

1
Q

what does the genome of each species of eukaryote consist of

A

a characteristic number of independent linear DNA molecules

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2
Q

each linear DNA molecules is a

A

chromosome

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3
Q

what do chromosomes never exist as

A

naked DNA

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4
Q

what do chromosome always exist as

A

DNA/protein complex termed chromatin

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5
Q

describe a key feature of chromatin organization

A

condensation or compaction
DNA molecule in average human chromosome is 5 cm long if stretched out = ~5000 times longer than width of typical cell nucleus
So DNA molecule is highly folded/packed/coiled even in the interphase nucleus

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6
Q

when are chromosomes more tightly folded/packed/coiled

A

mitotic metaphase (more packed than interphase)

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7
Q

why are chromosomes more tightly packed during mitotic metaphase

A

to facilitate equal distribution between the 2 daughter cells

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8
Q

describe metaphase

A

highly condensed for transmission to daughter cells
no DNA replication or transcription
only concerned with getting chromosomes to right place

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9
Q

describe interphase

A

real functional chromosome
undergoing replication and transcription
where action is
chromatin fiber of chromosome unwinds to a degree (from metaphase to interphase) - never to DNA/simplest chromosome structure
dynamic and controlled

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10
Q

what is chromatin

A

eukaryotic DNA and associated proteins

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11
Q

how many base pairs of DNA in each chromosome for humans

A

~50-250 million base pairs

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12
Q

what are topological domains

A

folded domains with definite boundaries
interact among themselves

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13
Q

what is nucleosome

A

DNA wrapped around histone octamer

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14
Q

where are giant interphase chromosomes from

A

polytene chromosomes of the fly (drosophila) salivary galdn

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15
Q

describe polytene chromosomes - why they happen

A

cellular gigantism driven by DNA ~10 cycles of DNA replication without cell division
duplicates but chromosome does not separate

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16
Q

describe polytene chromosomes - physically

A

all ~1024 daughter chromatids are in perfect alignment making a giant chromosome with regional differences in chromatin condensation
many parallel chromatids

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17
Q

describe regional differences in chromatin condensation

A

dark bands = condensed chromatin (topological domains)
light bands = boundary elements

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18
Q

describe interphase chromatin organization

A

dynamic
polytene chromosome puffs show how chromatin decondensation with transcriptional activation

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19
Q

describe puffs of polytene chromosome

A

puff up = active transcription regions, open and close
associated with active form of RNA polymerase 2 = active transcription

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20
Q

describe puffs of polytene chromosome - colours

A

red = pol 2 phosphoCTD = active
green = non phosphorylated = inactive

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21
Q

name parts of metaphase chromosome

A

sister chromatids
centromere
telomere
chromatid

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22
Q

describe metaphase sister chromatids

A

identical products of the previous semiconservative replication of a single chromosomal DNA molecule

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23
Q

what do metaphase chromosomes show

A

karyotype (morphology)
chromosomal complement of the species

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24
Q

what characteristics of chromosomes vary

A

number
shape
size
species specific and sometimes sex specific

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25
Q

describe ex of karyotype

A

human female karyotype revealed by fluorescence in situ hybridization (FISH) with a panel of probes representing sequences differentially distributed among the chromosomes
“chromosome painting” - identification

26
Q

what are chromosome rearrangements

A

breaks/translocation

27
Q

describe chromosome rearrangements

A

chromosomes can break and rejoin giving translocations

28
Q

when can translocations happen

A

mutations can happen during somatic cell division cycle in life and can cause disease like cancer
also can occur in germ line

29
Q

describe ex of chromosome rearrangements (cancer)

A

chronic myelogenous leukemia
results of chromosome breakage and fusion in blood cells
chromosome fusion generates chimeric gene encoding an oncogenic fusion protein
creates protein that deregulated cell growth in blood cells with mutation = cancer
called philadelphia chromosome

30
Q

describe chromosome rearrangements in germ line

A

gives gametes - eggs or sperm - with variant chromosomes
offspring often have reduced fertility
usually a dead end
this is why karyotype is so consistent across a species
can happen that it is successful and passed on = rare and karyotype can evolve over time

31
Q

describe evolution of human karyotype

A

from primate ancestral karyotype
unchanged Chr 11 (=13)
breakage Chr 14, 15 (=5)
reciprocal translocation Chr 12, 22 (=14, 21) end-to-end fusion Chr 2 (=9 + 11)

32
Q

what are the elements required for replication and stable inheritance of linear chromosomes - 3

A

origin of replication
centromere
2 telomeres - ends

33
Q

how do we learn about protein structure and function

A

by building artificial chromosomes

34
Q

how did we discover elements required for chromosome function

A

Experimental discovery in a simple eukaryote: yeast

35
Q

describe what yeast cells need

A

yeast leu- cells have LEU gene inactivated by a mutation and need exogenous leucine for growth

36
Q

what does LEU gene code for

A

makes leucine

37
Q

describe initial experiment done with yeast

A

wild type LEU gene cloned into circular bacterial plasmid
introduce to yeast cells and ask if it rescues leucine independent growth
aka does LEU plasmid replicate as cell grows

38
Q

describe initial experiment done with yeast - results

A

LEU plasmid replicates well in bacteria BUT INCAPABLE OF REPLICATION IN YEAST
since bacterial origins of replication do not work in eukaryotes

39
Q

how do we fix plasmid not working in yeast

A

insert into plasmid a random piece of yeast DNA that happened to contain a yeast origin of replication
now plasmid can support growth of leu - yeast in absence of leucine

40
Q

describe origin of replication (yeast experiment)

A

ARS = autonomously replicating sequenced = yeast origin of DNA replication
REQUIRED FOR REPLICATION
but cells not portioned well between daughter cells - mitotic segregation is faulty

41
Q

how to improve mitotic segregation

A

add centromere
CEN = DNA sequence from yeast chromosome centromere
drives good mitotic segregation

42
Q

what is kinetochore

A

attached to centromere
spindle microtubules attach here
then 2 chromatids separate and walk in opposite directions

43
Q

the centromere…

A

link to spindle microtubules

44
Q

describe yeast CEN

A

sequences common to various yeast centromere
present on nucleosome that includes a centromere specific histone variant CENP-A (centromeric protein A)

45
Q

what does CENP-A do

A

recruits CBF3 complex which in turn recruits Ndc80 complex which attaches to microtubules
initiates cascade of assembly

46
Q

describe image of CENP-A

A

different colour since its a different histone
tags it so it can recruit kinetochore apparatus - lateral attachment then to end on conversion
Kinetochores attached here and migrates and microtubule dissolves behind it as it moves
drags whole chromosome with it - this is how chromosome moves apart

47
Q

what shape are yeast chromosomes

A

linear DNA molecules
not circular
Previously we used a plasmid in yeast experiment which is circular

48
Q

how to convert circular plasmids to linear DNA molecules

A

cutting a single site with restriction endonucleases

49
Q

Does a plasmid with ARS and CEN that works well as an experimental circular chromosome in yeast also work well as a linear chromosome?

A

nOOOOOOOOOO
lose everything - linear does not work
so add telomeres

50
Q

what are telomeres

A

special sequences at ends of eukaryotic chromosomes
cloning a bit of telomere DNA at end and it works - needed for linear molecules to survive

51
Q

describe conclusions of yeast experiment - telomeres

A

cut plasmid without telomeres = unstable
linear plasmids containing ARS and CEN act normal if genomic fragment telomeres added to both ends

52
Q

name the functions of telomeres - 3

A

protect from exonuclease
prevent end-to-end fusion
solve a replication problem faced by linear DNA

53
Q

what is telomere problem

A

because lagging strand synthesis cannot be completed - chromosomes shorten at ends in each replication
unsustainable since at some point you will lose an essential gene
15-20 bases of RNA and no way to make it DNA

54
Q

what is solution to telomere problem

A

telomerase
DNA polymerase that can extend telomeres
restores chromosome length to overcome lagging strand end shortening

55
Q

what do telomeres contain

A

simple repeat DNA sequences
ciliate protist tetrahymena (TTGGGG)
human (TTAGGG)

56
Q

describe telomerase

A

reverse transcriptase that carries its own template RNA complementary to the telomeric DNA repeat
by extending template strand telomerase gives primase more template DNA to prime on

57
Q

what is a reverse transcriptase

A

DNA polymerase that uses RNA as template

58
Q

describe how telomerase works informally

A

binds to telomere
complementary to telomere
so reverse transcriptase can extend it
slips so can extend next primer - adds telomere repeats

59
Q

where is telomerase active

A

germ cells and stem cells not in somatic cells since divide only a few times (so existing telomeric repeats are long enough)

60
Q

when is telomerase often reactivated

A

in cancer cells - a target for cancer therapy

61
Q

describe mice lacking telomerase gene

A

ok for 3 generations then fertility decline
has enough repeats so it can go through 3 generations before ends are shortened too much and a key gene is not replicated (it is lost)

62
Q

describe ciliated protist tetrahymena

A

unusual gene expression mechanism
does not express genes from its “main” genome. maintained in a non-transcribed form in the micronucleus
transcription occurs in the macronucleus from millions of gene-sized DNA pieces.
presence of so many DNA ends made it possible for researchers to identify the telomere sequence and to isolate telomerase

BIOL 200 (29 decks)

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