LECTURE 29 - MIDTERM 3

Question 1

What is an overview of gene, genome, chromatin and chromosome?

Answer 1

– Gene: a chromosomal segment that encodes for a single polypeptide chain or RNA molecule or plays a regulatory function

– Genome - the total genetic information contained in a cell, an organism, or a virus

– Chromatin - DNA complexed with histones and other proteins; typically dispersed throughout the nucleus during interphase

– Chromosome – a compact strand of DNA that is encoded with genes; composed chromatin; formed during nuclear division

Question 2

What is the comparison between bacterial vs. viral genomes?

Answer 2

– Bacterial, archael, viral or eukaryotic cells all have one thing in common, they must all be able to pack large amounts of genetic material into very limited space

– packing of genetic material must be done in organized fashion, so that organism can undergo rapid replication on demand

– some viruses have RNA for their primary genetic material

– Many of these are important and useful as tools in biological research

Question 3

Describe the compaction of DNA.

Answer 3

– All cells must compact DNA to very limited space

– Must still be organized so that it can be replicated and transcribed, as necessary

– Bacteria compact their chromosomes by negative supercoiling and organizing into loops bound to proteins, bacterial nucleiod

– exists in cytosol

– Eukaryotic DNA is confined to the nucleus and wrapped around histone proteins to further compact the DNA

Question 4

Describe the relationship between genome size and organismal complexity.

Answer 4

– not a simple relationship between the two

– Bacteria only have one set of chromosomes whereas Eukaryotes have two sets (except for sex chromosomes)

• • Eukaryotes contain lots of DNA sequence that doesn’t directly encode proteins or RNA for transcription, called non-coding genes
    
    • -> or they code for RNA machinery of protein synthesis

Question 5

T or F, Bacteria has haploid DNA whereas Eukaryotes have diploid DNA

Answer 5

True

Question 6

T or F, Eukaryotic chromosomes comprise different types of DNA sequences.

Answer 6

True; Genes (37.5%) - introns, non-coding regions within a gene
- exons, protein (or functional RNA) coding regions

Inter-genes (62.5%) - regulatory elements (ex: promoters)

                              - genome-wide repeats (ex: transposons)
                              - microstaellite repeats (ex:CACACACACA...)

Question 7

T or F, most of the human genome is made of repeated DNAs and regulatory elements

Answer 7

True

Question 8

Describe DNA and the central Dogman.

Answer 8

– DNA serves as a “blueprint” for all of the proteins in the body

– Genes are segments of DNA that code for specific parts of protein

– These proteins perform a vast array of functions

   - - structural proteins (e.g, collagen). -- enzymes (e.g. aromatase)
   - - ion channels (e.g., Na+/K+ pump)  -- transcription factors (signal specific genes to turn on or off)

Question 9

What is removed during mRNA splicing?

Answer 9

– introns

Question 10

Where is eukaryotic chromatin contained?

Answer 10

– in the nucleus

– Transport between the nucleus and the cytoplasm occurs through small pores in the nuclear envelope called Nuclear Pores

– RNA and proteins can be imported and exported

– Transcription will occur in the nucleus, but translation will occur in the cytoplasm unlike in prokaryotes

Question 11

Describe how DNA is complexed into a chromosome.

Answer 11

– DNA is complexed with histones to form nucleosomes

– It is then stacked and as it becomes more and more stacked it eventually becomes chromosome

– nucleosome –> DNA + protein

Question 12

T or F, DNA organized in bacteria is circular and haploid

Answer 12

True

Question 13

What does the nucleosome consist of?

Answer 13

– histone core and the DNA wrapped around it

– linker DNA in nucleosomes is Naked DNA that separates one nucleosome from another around 20-60 base pairs

– Nucleosomes compact the DNA about 6-fold

– The DNA is wound 1.65 times around histone core

– very abundant, among the most conserved proteins in evolution

Question 14

What is Histone H1? (purpose wise)

Answer 14

– “linker histone” tightens the nucleosomal strucuture

– apart of the reason as to why DNA is able to remain wrapped around the histone

– acts like tape

Question 15

What are histones?

Answer 15

– small positively charged protons

– these are the second reason as to why DNA is able to remain wrapped around histone

– they have lysine and arginine which are positively charged and DNA is negatively charged due to phosphate so there’s this affinity that occurs

Question 16

What are features of Histone Modifications?

Answer 16

– Covalently attached groups (usually to histone tails)

–> modify amino acids on histone tails

– Reversible and Dynamic

      - -> Enzymes that add/remove modification
     - -> Signals

– Have diverse biological functions

Question 17

How is chromatin further compacted?

Answer 17

– by association with a protein scaffold

– chromatin loops (40-90 Kb)

Question 18

How does the compaction of the genome occur? (in terms of steps)

Answer 18

a. have the DNA being wrapped around histones
b. then become nucleosomes with about 30-40 base pairs between each nucleosome (aka linker DNA)
c. these nucleosomes are then stacked on top of each other
d. then with the nucleosomes stacked there are these loops of 30 nm fibre attached to central protein scaffolding –> central protein scaffolding

Question 19

What happens with histones and access to DNA-specific binding proteins?

Answer 19

– DNA-binding proteins generally prefer to bind naked DNA

– more “exposed” binding sequences are more accessible to DNA-binding proteins

– the way DNA stacks and loops makes itself unavailable for it to be exposed; binding sites are unavailable

– this is where acetylation and methylation comes in

 -----> acetylation allows for unraveling where as methylation allows it to further condense

– DNA has to shift around/unravel itself in order to make itself available

Question 20

T or F, chromosome compaction varies throughout the cell cycle

Answer 20

True

Question 21

What is the difference between Euchromatin and Heterochromatin?

Answer 21

• • Euchromatin:
    
    • –> Loosely packed regions of chromatin which stains light
    • –> Transcriptionally active chromatin ( a lot of naked DNA)
    • –> genes will be transcribed at one time
    • –> less condensed, gene rich, replicated through S phase, recombination during meiosis, at chromosome arms, contains unique sequences

– Heterochromatin

• –> Densely packed regions of chromatin which stains dark
• –> Transcriptionally inactive: too tightly compact to allow transcription to happen
• –> highly condensed, at centromeres and telomeres, contains repetitious sequences, gene-poor, replicated in late S phase, no meiotic recombination

Question 22

How are genomes studied and manipulated?

Answer 22

– Recombinant DNA technology

– restriction enzymes bind DNA as dimers, act like “molecular scissors.” (will cut specific regions on DNA)

– They recognize palindromic DNA sequences (two half-sites) –> reads one way in one direction and reads same way in other direction

– Cleave DNA symmetrically on both strands of the DNA

– restriction enzymes are produced by bacteria to protect against invading DNA (e..g. phage infection)

– Each restriction endonuclease recognizes a specific sequence of nucleotides, cuts more than once within a certain DNA molecule which results in having fragments of different length

– blunt or staggered end

Question 23

Description of the function of Restriction Endonucleases.

Answer 23

– enzyme is cutting phosphodiester bond that is apart of backbone of DNA

– restriction enzyme cleavage may generate blunt or overhanging (sticky) ends

– complementary base-pairing by sticky ends facilitates their ligation back together

– sticky ends are more cohesive compared to blunt ends

– the restriction endonuclease will bind to a specific DNA sequence and makes one cut in each of the sugar phosphate backbones of the double helix – by hydrolyzing the phosphodiester bond (specifically between the 3’ O atom and the P atom is broken)

Question 24

How do restriction-modification systems protec the host genome from Bacterial DNA Viruses?

Answer 24

– these enzymes cut foreign DNA, thus protecting the bacteria from infection (via bacteriophages)

– restriction enzymes prevent the replication of the phage by cleaving its DNA at specific sites

– The host DNA is protected by methylases which add methyl groups to adenine or cytosine bases within the recognition site thereby modifying the site and protecting the DNA from cleavage

Question 25

T or F, viruses over time can come up w’ their own methylation to be protected

Answer 25

True

Question 26

T or F, different restriction Endonucleases Recognize Different Length Sequences

Answer 26

– The length of the recognition sequence determines the approximate cleavage frequency

– these are Type II RE, but there are also I and III that cut outside their recognition sequences

Question 27

How can restriction enzymes be used to analyze genomes?

Answer 27

– can help w/ the isolation of specific genome to study

– fragments produced from restriction enzyme digestion can be analyzed to give a physical map of the RE sites, within a DNA molecule called Restriction Maps

– these help u understand and study specific regions