Lecture #2

Question 1

Nucleotides are the Building Blocks of DNA

Answer 1

DNA: deoxyribonucleic acid 
Each nucleotide contains: 
    - Phosphate group 
    - sugar 
    - nitrogenous base 
sugars 
    -DNA 
    - RNA  (one has a hydroxyl group, one doesn't)

Question 2

The Nitrogenous Bases

Answer 2

2 types of bases 
   - pyrimidines 
    - purines  
Pyrimidine 
   - Cytosine, Thymine, Uracil 
Purine 
   - Adenine and Guanine

Question 3

Creating a DNA strand

Answer 3

Nucleotides covalently bind to each other through the phosphate group.
- sugar interacts w/ phosphate group at the 3’ and 5’ position.
- strands of DNA exhibit “chemical polarity”
5’ to 3’
Hydrogen bonds create the second strand

Question 4

Double Stranded DNA

Answer 4

Nucleotides pair with thei rpartner through hydrogen bonds.
-A-T
-C-G (has to do with hydrogen bond pairing)
Creates a complementary strand
- 3’ to 5’
-Cytosine and guanine is more stable b/c of the more hydrogen bonds
Hydrogen bonding creates a planar region

Question 5

DNA double Helix

Answer 5

DNA molecular twists upon itself to form a helix.
- Right handed helix
- 10 bp/ turn
This conformation is the most energetically stable
- Bases are on the inside
- Sugar/Phosphate backbone
Various proteins interact using the grooves
~2m of DNA per cell
- Nucleus is 5-8 um

Question 6

DNA is packed into chromosomes

Answer 6

DNA is coiled around proteins and packed into higher ordered structures.
- Humans: 3.2x10^9 nucleotides w/ 24 pairs of chromosomes.
- 22 pairs of autosomal chromosomes; 2 sex chromosomes.
Each cell contains two copies of each chromosome
- maternal and paternal
- homologous chromosomes
chromosome

Question 7

Chromosome Characteristics

Answer 7

Gene vs non- gene encoding regions 
Gene recording regions contain the instructions for making a particular protein 
Non- gene encoding regions 
   -"Junk DNA" 
   - regulatory regions 
   - RNA

Question 8

Major Regions

Answer 8

Heterochromatin: Highly compacted (gene poor)
Euchromatin: More relaxed (gene rich. more proteins can get in)
Centromere: Constricted region on the chromosome used to attach sister chromatids.
Telomere: sequences of repeating nucleotides located on both ends of the chromosome. (Protect DNA)

Question 9

Chromosomes can exist in multiple states

Answer 9

Chromosomes change as the cell passes through the cell cycle
Interphase: G1 to G2
-Chromosomes are more relaxed and extended.
Mitosis: Chromosomes are more condensed.
(S)–> chromosomes get replicated

Question 10

Interphase chromosomes are “relaxed”: Why?

Answer 10

Chromosomes are replicated during the “S” stage of the cell cycle
- Synthesis
Chromosomes replication occurs at replication origins.
-Multiple points
- Why?
Our chromosomes are much longer.
Must replicate quickly

Question 11

Mitotic vs. Interphase chromosomes

Answer 11

Chromosomes entering mitosis are highly condensed
- All chromosomes closer together, and easily divided at this point.

Sister chromatids held together by centromeres.

Easily separated into different cells.

Question 12

Technique to visualize chromosomes

Answer 12

Karyotyping: Display of the 46 human chromosomes

Staining

• Giemsa: works (reacts) better w/ A and T’s b/c they are not as strong.
• Dark and light banding
• The banding for each chromosome is unique.

Painting

• Make probes for specific chromosomes regions.
• Label probes w/ flouresent dyes
• View the chromosomes through a flourescence microscope.

Question 13

Technique to visualize chromosomes part 2

Answer 13

Painting can be done when cell is in interphase or Mitosis

Interphase chromosomes occupy discrete regions within the nucleus
- Attached to the nuclear envelope.

Question 14

How are chromosomes compacted?

Answer 14

Nucleosomes are basic unit of chromatin compaction.
- Less compacted form of chromosome
- DnA + proteins
147bp of DNA wrapped around an 8 proteins called histones.
- 2 copies of H2A, H2B, H3 and H4
- H1 histones link nucleosomes together

Question 15

Histones: building blocks of the nucleosome

Answer 15

H2A, H2B, H3 and H4 
   - 2 copies of H3 and H4 form a tetramer 
   - one copy of H2A and H2B form a dimer 
Highly basic (positively charged) 
      pH lysings and argenins 
      pH 7--> (+) charged neutral 
Interact with DNA (negatively charged) 
     sugars--> nutral 
     Nitragenous base--> neg.

Question 16

Why does the packaging need to be dynamic?

Answer 16

Changes in the nucleosome structure allows access to DNA.
Chromatin Remodeling Complexes
- ATP dependent; needs multiple cycles of ATP hydrolysis.
- Acts to reposition and partially de-condense the chromatin
- Some are deactivated during mitosis; Why is this a good idea?
So the job can get done.

Question 17

Why does the packaging need to be dynamic part 2

Answer 17

Protein Modifications (Post-translational Modifications) 
   - Small chemical groups or proteins attached to a protein after translation has occured.  

Modifications 
   - Acetylation 
   - Methylation 
   - Phosphorylation 
    - Ubiquitination
          Small protein 
          76 amino acids 
          found in most Eukaryotic cell

Question 18

Protein modifications affect nucleosome stability

Answer 18

Protein modifications can alter histone function, nucleosome stability and histone-protein interactions.

Histones are modified on their N (closest to amino group) and C termini (closest to carboxyl group)

Different patterns of modifications yield different outcomes.

Modifications affect higher order packing more significantly than individual nucleosome stability

- small disturbances vs. large alterations 
- chemical groups vs. small proteins

Question 19

Protein modifications affect nucleosome stability Part 2

Answer 19

Modifications can either de-stabilize (relax) or stabilize (condense) nucleosomes

• Methylation of histones generally leads to gene silencing.
• Ubiquitination of H2B generally leads to gene activation
• Ubquitination of H2A generally leads to gene silencing.

Question 20

Chromosomes contain relaxed and condensed regions

Answer 20

-Regions of euchromatin are usually gene rich and expressed.
- Regions of heterochromatin are usually silenced.
- 10% of interphase chromosome
- Take up Giemsa stain more readily (easily stained)
- Some regions are permanently silenced.
X- inactivation
Why does it matter?
Gene expression

Question 21

Changes in chromatin structure can be inherited

Answer 21

Epigenetic Inheritance
- “Epi” in greek means “on”
- superimposed on genetic inheritance based on DNA.
During mitosis each daughter cell receives 1/2 its DNA from the parent cell
- Inherits the nucleosome condensed chromosome
The daughter cell has a copy of the modification made in the parent cell
- DNA and histone
Upon replication the daughter cell “remembers” the modifications.