Structure of Genomes I & II (lectures 2-3)

Question 1

Number of genomes sequenced

(February 2024;
https://www.ncbi.nlm.nih.gov/genome)

Answer 1

• Eukaryotes: 30,530
• Prokaryotes: 567,228

Question 2

Why is understanding genome structure important? = 3

Answer 2

1 * Medicine – predisposition to certain diseases, response to drugs

2 * Explanations for evolutionary change

3 * Production of “better” food (plants and animals), fodder, and fuel

Question 3

Gene Ontology (GO) Annotations (from the website: https://geneontology.org/):

“Associations of gene products to GO terms are statements that describe”: 3

Answer 3

1. “Molecular Function:
   - the molecular activities of individual gene products”
2. “Cellular Component:
   - where the gene products are active”
3. “Biological Process:
   - the pathways and larger processes to which that gene product’s activities contributes”.

Question 4

Prokaryotic genomes: What are they?

differences?

Answer 4

1. Archea
2. Bacteria

— differences:
different MOLECULAR and GENETIC characteristics

Question 5

Prokaryotic genomes:

Bacteria and Archea

Similarities?

Answer 5

1 * no extensive internal compartments

2 * chromosome / nucleoid

3 * plasmid(s)
————4 * integrative or independent

Question 6

Prokaryotic genomes: How is DNA packaged? = 6

Answer 6

1. DNA packaged with DNA-binding proteins
   2 * typically circular genomes
   3 * negative supercoiling
2. Protein core limits loss of supercoiling if a break occurs
   5 * domains
   6 * loops ~10 – 100 kb

Question 7

Escherichia coli nucleoid:

Answer 7

Know most about ‘Escherichia coli’ nucleoid but DNA-binding proteins found in other species, including archea.

Question 8

Prokaryotic genomes cont’d = 5

shape and coiling

Answer 8

1. Circular, double-stranded DNA
2. Remove a few turns of the double helix
3. Molecule forms a negative supercoil

diagram Protein core, Supercoiled DNA loops, Broken loop- no supercoiling

Look at Diagrams on it

Question 9

Prokaryotic genome size:

Answer 9

Prokaryotic genome size is variable

• most < 5 Mb
• range: 112 kb – 14.8 Mb
• average gene density: ~950 genes / 1Mb

Question 10

Prokaryotic genome size: Largest genome and smallest genome:

Answer 10

1 * largest genomes found in free-living soil bacteria – ability to respond to changing environment

2* smallest genomes in endosymbiotic bacteria – more consistent environment

Question 11

Prokaryotic genome size - commonly arranged …?

Answer 11

• commonly arranged in OPERONS (not universal)
• group of genes
• involved in the same biochemical pathway and
• expressed as a single unit

Question 12

Prokaryotic genome size…is proportional to?

Answer 12

Prokaryotic GENOME size is proportional to GENE NUMBER

Question 13

Prokaryotic genome organisation and content: 4

Answer 13

1. Majority of BACTERIAL and ARCHEAL genomes are CIRCULAR
   …..2 * SOME are LINEAR,
   e.g. Borrelia burgdorferi (causative agent of Lyme disease)
2. Many PROKARYOTIC genomes are MULTIPARTE
   ….4 * two or more molecules

Question 14

Prokaryotic genome organisation and content :

PLASMIDS = 7

Answer 14

1. typically CIRCULAR
2. REPLICATION INDEPENDENT of nucleoid
3. up to 1000s of COPIES / cell
4. PARTITIONED TO NEW CELLS INDEPENDENT OF NUCLEOID
5. contain GENES NOT ESSENTIAL FOR SURVIVAL IN PERMISSIVE HABITATS/CONDITIONS
6. TRANSFERRED to / TAKEN UP BY VARIOUS SPECIES
7. argument: plasmids not
   be included in definition
   of prokaryotic genomes
   BUT….

Question 15

Essential genes are found on ‘D. radiodurans’ R1 plasmids….

EXPLAIN = 4

Answer 15

‘B. burgdorferi’ (causative agent of Lyme disease)

• 1 linear chromosome
• 19 linear and circular plasmids
• indispensable genes, e.g. encoding some membrane proteins

Question 16

Chromosome vs chromid vs plasmid

Answer 16

1. CHROMOSOME (s) – located in nucleoid, carries essential genes
2. CHROMID – uses plasmid partitioning system, carries essential genes
3. PLASMID– uses plasmid partitioning system, carries nonessential genes

Question 17

’ V. cholerae’ vs ‘D. radiodurans’

Answer 17

‘V. cholerae’ : one chromosome, one chromid

‘D. radiodurans’ : two chromosomes and two chromids

Question 18

‘E.coli’ genome… space? separation?

other parts

Answer 18

1. 11% of genome = non-coding DNA
2. • little space between genes
3. • some genes separated by only a single
     nucleotide (thrA and thrB) or none (thrB and
     thrC)
4. • thrA-C = operon; encodes proteins for
     threonine biosynthesis
5. Some archeal genes have introns
6. Some prokaryotes contain nested genes = genes encoded within other genes (aka overlapping genes)
7. Bacterial genes = slightly longer than archeal genes

Question 18

What is in ‘E.coli’ genome SEQUENCES? = 8

Answer 18

1. repeat sequences
   2. • few high-copy-number interspersed repeat families (compared with eukaryotes;)
   3. • insertion sequences (IS)
   4. • mobile elements (transposons) repeated in the genome
2. • nontransposable repeat elements
     6. • repetitive extragenic palindromic (REP) sequences
3. • gene regulation?
     8. * clustered regularly interspaced short palindromic repeats (CRISPRs)

Question 19

‘Prokaryotic genome organisation and content’

Lateral (aka horizontal) gene transfer; where, who? = 5

Answer 19

1. • gene flow between prokaryotic species
     
     2. • frequent
2. • most prokaryotic genomes contain hundreds of kb of DNA from different
     prokaryotic species

      4.* transfers occur between bacteria and archea

3. • DNA originates from the environment, exchange of plasmids and viral
     vectors

Question 20

‘Prokaryotic genome organisation and content’

Lateral (aka horizontal) gene transfer; how? = 7

Answer 20

1. multiple genes in a singe transfer
2. • mechanisms of transfer
     3. * transformation
     4. * conjugation
     5. * transduction
3. • confuses species relationships
     7. * laterally transferred gene will have relatively similar sequences in two species – due to little time for sequence divergence

Question 21

‘Prokaryotic genome organisation and content’

Lateral (aka horizontal) gene transfer;EXAMPLES? = 4

Answer 21

1. • antibiotic resistance
2. • ability tolerate hot environments
3. • anaerobic to aerobic GROWTH HABITS
4. • METABOLIC PATHWAYS

Question 22

Prokaryotic genome organisation and content cont’d DIAGRAMS

Answer 22

SLIDE 18

Question 23

Prokaryotic gene function catalogue (GO Terms):

‘E. coli’ genome = 5
FUNCTION
GENE FAMILYS

Answer 23

1. function of many genes still not known
   2. • no significant similarity to any known genes in other bacteria
2. • gene families
   4. • genes having arisen from gene duplication events
      5. • e.g. rRNA genes in bacteria and archea

Question 24

Eukaryotic genomes: Heterochromatin vs Euchromatin = 16

Answer 24

1. LINEAR CHROMOSOMES with NEGATIVE SUPERCOILING in MEMBRANE-BOUND NUCLEUS 'Heterochromatin' 2. * densely staining regions in interphase nucleus 3. * chromatin densely packed 4. * constitutive 5. * permanently condensed chromatin 6. * DNA is gene poor - but does contain some genes 7. * centromeric, telomeric 8. * many repeat regions 9. * other chromosome regions 10. * most of the human Y chromosome 11. * facultative 12. * not permanently condensed 13. * exists in some cells at some times 14. * DNA encodes genes that are inactive at particular times or in particular cells 'Euchromatin' 15. * cannot see in interphase nucleus 16.* DNA is less condensed and gene rich

Question 25

Eukaryotic genome organisation; 'Nucleosomes' = 7

Answer 25

1. * protein core = histone octamer 2. * 2 subunits of histones H2A, H2B, H3, and H4 3. * DNA ~147 bp 4. * “slide” 5. * expose chromatin regions for transcription 6. * also involves chromatin-remodelling proteins 7. * removed and replaced during DNA replication

Question 26

Eukaryotic genome organization cont’d: 'Higher orders of chromatin structure' = 4 UNDERSTANDING EUCHROMATIN LOOPS

Answer 26

1. Euchromatin loops ARE 2. * dynamic 3. * extension / merging – allow access to transcriptional machinery 4. * condensation – repress transcription

Question 27

Eukaryotic genome organization cont’d: 'Higher orders of chromatin structure' = DIAGRAM

Answer 27

SLIDE 22

Question 28

Eukaryotic genome organization: 'Organisation of chromosomes in interphase nucleus' = 13

Answer 28

1. Organisation of chromosomes in interphase nucleus 2. CT – chromosome territory 3. * specific for each chromosome 4. LAD – lamin-associated domains 5. * heterochromatin interacting with nuclear lamins at nuclear periphery 6. TAD – topologically associating domain 7. * Compartment A 8.* transcriptionally active 9.* enhancer-promoter interactions 10. * Compartment B 11. * transcriptionally repressed (facultative heterochromatin) 13. * chromatin switches between compartments, depending gene expression demands

Question 29

Eukaryotic genome organization: 'INSULATORS' = 7

Answer 29

1. define TAD boundaries 2. * 1-2 kb long 3. * in many (all?) eukaryotes 4. * DNAse I insensitive 5. * interact with specific DNA-binding proteins 6. * establish functional domains, e.g. loop 7. * prevent cross-talk of regulatory domains between functional domains

Question 30

Eukaryotic genome organization: 'INSULATORS' DIAGRAM

Answer 30

SLIDE 24

Question 31

Eukaryotic genome size: 3

Answer 31

1. Great VARIABILITY in eukaryotic genome size 2. * 10 Mb – 100,000 Mb 3. * EUKARYOTIC GENOME SIZE is NOT PROPORTIONAL TO GENE NUMBER

Question 32

Eukaryotic genome size: correlation? = 4

Answer 32

1. Overall correlation of genome size with morphological complexity of organisms 2. * HOWEVER, no precise correlation between genome size and complexity 3. *especially evident when looking within eukaryotic groups 4. * C-value paradox/enigma (C-value = haploid genome size)

Question 33

Eukaryotic genome size: 'Factors contributing to the C-value paradox/enigma' = 3

Answer 33

Factors contributing to the C-value paradox/enigma 1. * non-protein coding DNA 2. * gene density 3. * “split genes” – # introns / gene

Question 34

Non-protein coding DNA scales with morphological complexity DIAGRAM

Answer 34

SLIDE 27 PROKARYOTES AT THE BOTTOM EUKARYOTES = MOST

Question 35

Repeat sequences in eukaryotic genomes: 'INTERSPERSED REPEATS' = 10

Answer 35

1. GENOMES of most MULTICELLULAR EUKARYOTES have substantial amounts of moderately and HIGHLY REPETITIVE SEQUENCES 2. Interspersed repeats 3. * repeat units distributed (seemingly) randomly around the genome 4. * in intergenic regions and introns 5. * DNA transposons 6. * retrotransposons 7. * LTR – long terminal repeat retrotransposons 8. * Non-LTR retrotransposons 9. * SINE – short interspersed nuclear element 10. * LINE – long interspersed nuclear element

Question 36

Repeat sequences in eukaryotic genomes: TANDEM REPEATS = 16

Answer 36

Tandem repeats 1. * repeat units located next to each other 2. * satellite DNA (satellite bands after fractionation and density gradient centrifugation of genomic DNA) 3. * repeat unit < 5 bp to > 200 bp 4. * clusters 100s of kb in length 5. * e.g. centromeric DNA 6. * minisatellites 7.* not part of satellite bands on gradients 8. *repeat unit up to 25 bp 9. * clusters up to 20 kb 10. * e.g. telomeric DNA 11. * microsatellites 12. * not part of satellite bands on gradients 13. * repeat unit < 13 bp 14. * clusters < 150 bp 15. * used to establish kinship 16.* an individual’s genetic profile DIAGRAM SLIDE 29

Question 37

Eukaryotic genomes contain pseudogenes (2 TYPES) AND gene relics = 9

Answer 37

1. Conventional pseudogene 2. * inactivated due to mutation 3. Processed pseudogene 4. * derived from a mRNA that is converted to cDNA and reinserts into genome 5. * no introns or regulatory regions that ancestral gene had 6. * inactivated 7. Gene relics 8. * truncated gene – from 5’ or 3’ end 9. * gene fragments

Question 38

Eukaryotic genomes contain pseudogenes and gene relics = DIAGRAM

Answer 38

SLIDE 30

Question 39

Eukaryotic gene density

Answer 39

Genes are more closely packed along the chromosomes of less complex organisms

Question 40

Less complex organisms contain fewer split genes

Answer 40

Genome of yeast compared to genomes of more complex eukaryotes * few genes with introns – yeast genome has 239; human genome > 300,000

Question 41

G-value paradox

Answer 41

- Gene number does not scale with morphological complexity - Alternative splicing leads to multiple mRNAs and proteins from a single gene * explains part of the C- and G-value paradoxes

Question 42

G-value paradox = DIAGRAM

Answer 42

SLIDE 33

Question 43

Eukaryotic genomes contain gene deserts: 9 WHAT, SIGNIFICANCE? IN HUMAN GENOME?

Answer 43

1. Large regions of chromosomes (10^5-10^6 bp) devoid of known genes or other functional genetic elements 2. * human genome 3. * 25% consists of gene deserts 4. * chromosomes 4, 5 and 13 (30-40% of the chromosomes) 5. * significance of gene deserts 6. * not known 7. * some contain regulatory sequences that act over large distances to control gene expression 8. * others show no clear function 9. * superfluous regions of genomes??

Question 44

Eukaryotic genomes contain gene families: SIMPLE VS COMPLEX: SIMPLE...= 9

Answer 44

1. Simple (aka classical) gene families 2. * all members have identical or nearly identical sequences 3. * arose from gene duplication events 4. * rRNA genes 5. *humans: 6. * 2000 genes for 5S rRNA 7. * single cluster on chromosome 1 8. * 280 copies of 28S, 5.8S, 18S repeat unit 9. * 50-70 repeats clustered on multiple chromosomes

Question 45

Eukaryotic genomes contain gene families: SIMPLE VS COMPLEX: COMPLEX...= 7

Answer 45

1. Complex gene families 2. * members have similar sequences 3. * different enough to code for gene products with different properties 4. * arose from gene duplication events 5. * mammalian globin genes 6. * expressed at different developmental stages 7. * biochemical properties correlate to physiological needs during development

Question 46

Eukaryotic genomes contain nested genes? How many Categories? =7

Answer 46

1. Overlapping genes found in the genomes of yeast, protists and metazoans 2. Two major categories 3. * genes nested within intron of another gene (= external host gene) 4. * relatively common in eukaryotes 5. * non-intronic genes nested opposite coding sequence of external host gene 6. * no clear evidence of these in metazoan genomes 7. * present in yeast and protistan genomes (and prokaryotic genomes)

Question 47

Eukaryotic genomes contain nested genes diagram

Answer 47

slide 36

Question 48

Eukaryotic gene function catalogues (GO Terms): = 8

Answer 48

1. Human genome 2. * greatest number of genes in all categories except metabolism 3. * many more genes involved in defence and immunity 4. 'Caenorhabditis elegans' (nematode worm) genome 6. * high number of genes in cell-cell communication category 7. *1000 genes vs 1250 in humans 8. *BUT only 959 cells vs 1013 cells in humans

Question 49

The Hidden Genome: Non-coding (nc)RNAs

Answer 49

Non-coding (nc)RNAs * tRNAs, rRNAs, circRNA (circular RNA), eRNA (enhancer RNA), lincRNA (long intergenic non-coding RNA), microRNA (miRNA), NAT (natural antisense transcript), piRNA (PIWI RNA), scaRNA (small Cajal body-specific RNA), siRNA (small interfering RNA), snRNA (small nuclear RNA), snoRNA (small nucleolar RNA)

Question 50

The Hidden Genome = RNAs encoding microproteins and peptides = 8

Answer 50

1. smORF = small open reading frame 2. * shorter than 100 amino acids 3. * dORF = downstream open reading frame 4. * located in the 3’-UTR of known proteincoding genes 5. * uORF = upstream-encoded smORF 6. * located in the 5’-UTR of known proteincoding genes 7. * nuORF = novel unannotated open reading frame 8. * SEP = small peptide

Question 51

The Hidden Genome DIAGRAM

Answer 51

SLIDE 38

Question 52

The Forbidden Genome: 4

Answer 52

1. Short DNA sequences not compatible with life 2. * minimal absent words (MAWs) 3. * not found in a particular genome (nullomers) 4. * not found in any genome (primes)

Question 53

The Forbidden Genome: USES = 4

Answer 53

1. * tags to distinguish samples (e.g. control or reference samples vs forensic samples) 2. * suicide genes that could be encoded by genetically modified organism and activated to destroy them if they prove dangerous 3 * anticancer peptides (NulloPs) 4. * biomarkers for cancers