1-2 Nuclear Structure and Function

Question 1

Describe structure of nuclear envelope

Answer 1

a double membrane barrier between nucleus and cytoplasm. Punctuated by nuclear pores where all traffic into and out of nucleus occurs

• outer - contiguous with ER, has ribosomes, proteins translated on these insert into lumen within membrane
• inner - attached to and supported by nuclear lamina

Question 2

Describe structure and function of the nuclear lamina

Answer 2

Meshwork of intermediate filaments formed from three proteins Lamin A, B, and C.

Attaches to inner membrane of Nuclear envelope and chromatin to stabalize envelope and organize chromosomes

Question 3

Describe the structure of nuclear pores

Answer 3

Nuclear Pore Complexes (NPC) are proteinaceous channels through nuclear envelope. Where all molecules enter/exit nucleus.

Arranged in 8-fold symmetry forming channel which small proteins (44kDa or less) can passively diffuse. Proteins 60kDa or larger must contain a nuclear localization signal (NLS)

Question 4

What happens to the lamin proteins during mitosis?

Answer 4

Lamin phosphorylation at mitosis induces disassembly of the nuclear lamina which causes nuclear breakdown

process reversed after mitosis, causing re-assembly of the nucleus

Question 5

Describe the structure of the nuclear pores

Answer 5

Nuclear Pore Complexes (NPCs) are proteinaceous channels which traverse the nuclear envelope. Pathway for all molecules moving into and out of nucleus.

Question 6

Describe the basic mechanism of transport through the pores

Answer 6

1. Carrier proteins recognize transport signals (NLocalS/NExportS) on substrate.
2. Carrier protein (importin/exportin) binds substrate and escorts through pore
3. Cargo/Carrier protein dissociate, carrier recycled back through pore.

Question 7

The functions of the nuclear pores allows for..

Answer 7

different protein and RNA compositions between the nucleus and cytoplasm

Question 8

What is the “cargo” exportin binds to in the nucleus?

Answer 8

mRNA, tRNA, or rRNA

Question 9

Transport in either direction is dependent on?

Answer 9

Small GTPase called RAN and two associated factors

• RanGTPase activating protein (GAP) on the cytoplasmic side (both have letter P)
• Ran GTPase exchange factor (GEF) on the nuclear side (both have letter E)

Question 10

What powers the transport in and out of the nucleus?

Answer 10

The assymetric distribution of GTP bound ran(activated) vs GDP-bound ran sets up a gradient which drives reactions.

Question 11

Describe protein import specifically

Answer 11

1. Import requires Nuclear Localization Signal on a substrate which is bound by importin carrier protein.
2. Importin directs docking of complex to fibrils extending from cytoplasmic face of NPC
3. translocation of complex
4. GTP bound ran directs dissocation of the carrier and cargo by binding to importin and lowering importin’s affinity for the cargo.
5. RanGTP bound importin is then recycled out of the nucleus into the cyctoplasm, where RanGTP is hydrolyzed to RanGDP by GAP, causing it’s dissocation from the importin.

Question 12

Describe protein export specifically

Answer 12

1. Export requires Nuclear Export Signal on the substrate which is bound by exportin carrier protein and RanGTP.
2. Exportin directs movement through the NPC to the cytoplasm
3. Once in the cytoplasm, GAP hydrolyzes the RanGTP to RanGDP, which dissociates the complex due to change in affinity. Releasing the cargo, Ran-GDP, and exportin which is recycled back into the nucleus.

Question 13

Describe the nucleoulus

Answer 13

non-membrane bound sub-compartment of the nucleus assembled on the rDNA repeats of 5 different autosomes with 2 homologes each for a total of 10 chromosomes.

sidenote: rDNA is ribosomal DNA which is a sequence that codes for rRNA

Question 14

Describe the pathway of ribosomal biogenesis

Answer 14

1. begins in nucleolus, transcription of genes for 45s rRNA large precursor
2. single 45s precursor RNA formed and processed into pre-ribosome by endo/exo nucleases into 28s, 18s, and 5.8s.
3. 18s incorporated into small subunit, 28s, 5.8s, and 5s (made elsewhere by RNA poly 3) incorporate into large ribo subunit
4. ribosome exported from nucleus

Question 15

Describe the structure and relative numbers of Chromosomes

Answer 15

-extremely long, linear polymers of double stranded DNA coated with structural/regulatory proteins called chromatin

• 22 different autosomes, 2 sex chromosomes
• haploid cell - 22 autosomes, 1 sex =23
• diploid cell - 44 autosomes, 2 sex = 46

Question 16

what is required to stabilize and duplicate chromosomes?

Answer 16

Three prominent sequences are required to stabalize and duplicate chromosomes

1. Replication origin- initiation of DNA replication
2. Centromers - segregation of duplicated chromosomes at mitosis and meiosis
3. Telomeres - stablility and replication of chromosome ends. Protects chromosomes against shortening during DNA replication

Question 17

There are two levels of packaging in ____phase cells.

Answer 17

interphase

euchromatin - less condensed, DNA being actively transcribed

Heterochromatin - more condensed, not currently being expressed

Question 18

When are chromosomes most condensed?

Answer 18

During Mitosis

Question 19

DNA packaging is mediated by _____. Types?

Answer 19

Histones. 5 Kinds (H1, H2A, H2B, H3, H4)

Question 20

Loss of function mutation that involves RanGEF is likely to have what effect?

Answer 20

loss of mRNA export from nucleus

Question 21

Describe how chromatin is formed

Answer 21

1) Positive residues of histones attatch to negative residues of DNA.
2) 8 histones (2 of each type) combine to form a histone octamer (nucleosome)
3) Multiple nucleosomes coil together and stack
4) resultant packed fiber of nucleosomes is 30nm thick and called chromatin

Question 22

Define “gene”.

Answer 22

Genes code for proteins such as RNAs (rRNA, tRNA, mRNA, snRNA, snoRNA, microRNA, IncRNA).??????????/

Question 23

Describe nuclear architecture

Answer 23

The nuclear matrix is a dense, underlying fibrous network composed mainly of proteins that provides structure/organization to the nucleus.
-Chromosomes attatch to the matrix through sequences called matrix associated regions (MARS)

-the matrix helps bring together regions of chromosomes being actively transcribed to specific foci (transcription/splicing factories)

Question 24

When does nuclear architecture change? how?

Answer 24

During Mitosis. Most eurkaryotes (including humans) undergo open mitosis which nucleus disassembles.

1. Transcription stops
2. Nucleolus dissolves
3. Chromosomes condense
4. Modification of the lamins (phosphorylation) causes the nuclear envelope to break down into vesicles

After mitosis - Lamins dephosphorylated.
nuclear matrix/envelope reassembles around de-condensed chromosomes
chromosomes fuse to recreate complete nucleus
rDNA transcription resumes creating 10 pre-nucleolar foci that fuse to reform nucleus

Question 25

Epigenetics of histones

Answer 25

each of the four main histones (H2A, H2B, H3, H4) have highly conserved n-tail domains that protrude from the nucleosome. - subject to many post-translational modifications such as acylation, methylation, phosphorylation. - regulated by a diverse set of histone acetylases and de-acetylases - these modifications can persist through several cell divisions and in some cases are heritable.

Question 26

Lupus

Answer 26

caused by production of antibodies to self antigens in the nucleus. -able to study new sub-compartments in the nucleus

Question 27

acute promyelocytic leukemia

Answer 27

caused by mutation in nuclear PML protein normally required to form a subcompartment of the nucleus, called PML bodies.

Question 28

Spinal muscular atrophy

Answer 28

second most common neuromuscular disease of childhood, duchenne MS being most common. inherited or mutation in the protein SMN (survival of motor neurons) beleived to be involved in RNA splicing.

Question 29

describe the compartments of the nucleolus

Answer 29

Regions 1) Fibrillar center - region contains DNA not being actively transcribed 2) Dense fibrillar compartment - contains rRNA molecules actively being synthesized 3) Granular compartment - contains maturing ribosomal precursor molecules

Question 30

what is the composition of the nucleosome?

Answer 30

2 copies each of histones H2A, H2B, H3, H4

Question 31

Where are ribosomes made? | What is the size of the large rRNA precursor and how is this broken up?

Answer 31

Ribosomes are made in the nucleolus region of the nucleus. Ribosome biogenesis involves forming a single 45s precurosr rRNA which is processed in the pre-ribosome by endo/exo-nucleases into 28s, 18s, and 5.8s Ribosome biogenesis is complete following export from the nucleus

Question 32

During mitosis, would you expect to see heterochromatin or euchromatin?

Answer 32

During mitosis, all of the chromosomes achieve their most condensed state...which is why we would expect to see heterchromatin (stains darker)

Question 33

Which statement concerning human chromosomes is wrong? A. Gametes contain 23 chromosomes. B. Homologues carry different alleles of the same genes while sister chromatids have the same alleles. C. Independent assortment to make gametes can have 2^46 possible outcomes. D. Recombination between sister chromatids would not contribute to genetic diversity. E. Recombination between homologues does contribute to genetic diversity.

Answer 33

C. 2^23

Question 34

``` The frequency of genetic recombination between two loci on a given chromosome will determine: Selected Answer: Incorrect A. linkage of genetic traits. Answers: A. linkage of genetic traits. ``` B. linkage of RFLPs. C. linkage of genetic traits to RFLPs. D. Only A and B are correct E. A, B, and C are correct.

Answer 34

E