Genomic Regulation Flashcards
Central Dogma of Genetics
1) DNA-> RNA-> Protein
2) Replication->Transcription-> Translation
Transcription and it’s steps
- DNA–>RNA is the goal
1) Initiation
2) Elongation
3) Termination
RNA Polymerase
1) Link nucleotides to for an RNA strand using DNA as a template
2) Adds RNa nucleotides 5’ to 3’
Initiation
1) RNA Polymerase binds to DNA promoter
2) RNA Poly separates DNA to make signle-strand template
Elongation
DNA template is read and RNA polymerase add a corresponding RNA base to a new RNA molecule
Termination
1) Terminators signal RNA transcript is done
2) Transcript is released from RNA polymerase
Pre-mRNA
Transcript of a protein-coding gene in eukaryotic cells
Splicing
Removing introns and extrons from pre-mRNA before assembling mature DNA sequence
-Increases the stability of mRNA
Nucleoside structure
Ribose sugar, phosphate group and nitrogen base
DNA Structure
1) Double stranded
2) Anti-parellel (5->3 and 3->5)
3) Sugar-phosphate backbone
4) Double helix w/ major and minor grooves
Importance of DNA condensation
To prevent physical damage to DNA as chromosomes are separated and based to daughter cells
What is Reverse Transcription?
- The synthesis of DNA from RNA
- Typically mediated by RNA viruses, which used Reverse Transcriptase to perform this event, which allows their genetic info to integrate with the host cell’s
Histones
1) Proteins DNA wraps around into super condensed form called chromatin
2) Linked by hydrophobic interaction and salt linkage
3) Lysine and Arginine very present residue (target of post-trans modification)
4) POSITIVELY CHARGED (DNA is negatively charged)
Nucleosomes
- DNA packed around a cluster of histones make a nucleosomes
- Basic unit of chromosome packing
Chromatin
-A DNA-PRotein complex made up of Protein + DNA in a large quantity and wrapped very tightly
Types of chromatin
1) Euchromatin - lightly packed (because usually under active transcription), highly enriched in genes
2) Heterochromatin - very condensed chromatin; present at centromeres and telomeres; few active genes present
Position effect
Gene activity depends on position within chromosome (i.e. genes near heterochromatin)
CGH Arrays
Comparative Genome Hydridization — test that allows us to determine copy number variations in our DNA to determine who has what difference with respect to disease states
-Gains and losses of Chromosomes in genome
Long Terminal Repeats (LTR)
- Identical sequences of DNA
- Used by viruses to insert their genetic material into host genomes
- Via reserve transcription
RNAi
- RNA Interference
- Process that allows RNA molecules (miRNA) to stop gene expression or translation in host genomes by targeting mRNA
miRNA
- microRNA
- Important in post-transcription gene expression — silencing
- Will bind to a protein, create complex that binds to sequence and will block translation of certain mRNA segments
Alternative splicing
When exons are either retained in the mRNA or targeted for removal in different combinations creating diversity in mRNA from PreRNA and thus an opportunity for mutations
DNA Helicase
-Unwinds DNA during replication, creating replication fork
Topoisomerase
- Moves ahead of helicase to remover overwound strands of DNA from portion created by helicase
- Target of anti-cancer drugs because inhibition leads to strand breaks, cell cycle arrest and cell death.
Primase
-Adds RNA primer segment to initiate start of replication
DNA polymerase
-Starting at RNA primer, moves 5’-3’ adding base pairs continuously on leading strand
Okazaki Fragments
-Short segments added 5’->3’ on lagging strand, which are primed by RNA primer
Exonuclease
-Removes RNA primers and replaces RNA with complementary DNA strand
DNA Ligase
-Seals DNA strands together by forming phosphodiester bonds between nucleotides
High Fidelity Replication
- Low errors in DNA transcription
- DNA proofread by DNA Polymerase and repair by others
- Germ and somatic errors low to 1) maintain species and2) avoid cancer proliferation
Histone Deacetylase (HDAC)
-Acetyl groups on histones are removed, causing DNA to wind extremely tight, resulting in repressed gene expression
Histone Acetyl Transferase (HAT)
-Promotes gene expressing by adding acetyl groups, unwinding DNA and allowing to be transcriptionally active
Post-Translational Modification (PTM)
- Enzymatic modification of proteins following synthesis
- Key for epigenetics
PTM on Histones
- H3 & H4 histone proteins have long tails that have various PTMs and are the target of PTMs:
- Methylation
- acetylation
- phosphorylation
- Ubiquitination
- SUMOylation
KEY: SUM of these modifications determine if a gene is turned on or off
DNA Damage Types
1) Ionizing Radiation
2) Non-Ionizing Radiation
3) Spontaneous Damage
4) Thymine Modifications
5) Intercalation
Ionizing Radiation DNA Damage
1) DNA strand breaks
2) base modification
3) Cross-linked DNA-proteins (thymine-tyrosine)
Non-Ionizing Radiation DNA Damage
Formation of pyrimidine dimers (i.e. pyrimidine cyclobutane dimer —– interfere with base pairing during replication leading to mutations)
Spontaneous DNA Damage
1) Depurination – 5000 purines lost per day
2) Deamination - C to U change (100 per day)
-Base deletion or substitution (One strand is good while the other is bad due to change — semi-conservative rep the reason)
Leads to negative effects with DNA replication, OBVI!!!!!
CpG Islands and Methylation (Spontaneous DNA Damage)
- CpG islands are associated with inactive genes (concentrated around promoters) and methylation results in gene silencing (typically DNA repair genes, leading possibly to cancer)
- Methylation of C leads to pairing with T, not G
Cross-Linking & Intercalating
When chemical agents (or something else) resulting in molecules to cross-link, leading to a potentially harmful molecule (i.e. BPDE, and carcinogen created from burning grilled meats (charcoal))
-In intercalation, agent inserts itself within DNA
Chemical Agent Damage to DNA
1) Cross-linking agents (mustard gas)
2) Alkylating Agents (Dimethyl sulfate)
3) Intercalating agents (Thalidimide)
Types of DNA Repair
1) Direct Repair
2) Base excision repair (BER)
3) Nucleotide excision repair (NER)
4) Mismatch excision repair (MER)
5) Recombination repair
6) Transcription Coupled Repair (TCR)
7) Translesion synthesis (bypass synthesis)
Genetic Defects in DNA Repair Pathways
1) MSH2, 3, 6, MLH1, PMS2 (Colon cancer)
2) Xeroderma pigmentosum (skin cancer)
3) Ataxia telangiectasia (AT) (Leukemia)
4) BRCA2 (Breast, Ovarian, prostate)
5) Franconi anemia (leukemia, abnormalities)
MSH2, 3, 6, MLH1, PMS2
- genetic defect in DNA repair
- Colon cancer
- Caused by mismatch repair
Xeroderma pigmetosum
- genetic defect in DNA repair
- Skin cancer
- nucleotide-excision repair
Ataxia telangiectasia (AT)
- genetic defect in DNA repair
- Leukemia
- ATM protein, activated by double-strand breaks
Franconi anemia group
- genetic defect in DNA repair
- Leukemia, congenital abnormality
- DNA cross-link repair
Direct repair
-i.e. photolyase removing thymine dimer from DNA, allowing light to pass
Base excision repair
-bad base is snipped out by DNA glycolyase and replaced with correct base
Nucleotide excision repair
-Segment of DNA strand removed and replaced by excision nuclease and DNA polymerase (i.e. removal of a pyrimidine dimer)
Clinical
Mismatch excision repair (MER)
- Error (Nick) in new DNA strand detected by DNA polymerase and followed and fixed by MutS and MutL
- Strand is removed
- Repair is made via DNA synthesis, Ligase seals
i.e. colorectal cancer (Autosomal dominant)
Double-strand base repair
1) Non-homologous -> broken ends from breaks brought together, leading to one or more nucleotides being lost - NBD
2) Homologous -> Template strand present (sister chromatids) so damage can be perfectly repaired
Transcription Coupled Repair (TCR)
- DNA repair directed to specific sequence that is being actively transcribed (i.e. urgent repair needed) – image occurs during expression
- RNa polymerase goes to problem area and direct machinery there
i.e Cockayne’s syndrome a defect in this repair system
Rifampicin
- Antibiotic that inhibits RNA Synthesis in bacteria
- -Side effects: upreg of hepatic cytochrome P450 enzymes – increase other drug metabolism
Epigenetics
Change in gene expression, not in the DNA sequence (PTMs) that turns genes on or off
Usually at histone tails
SUMOylation
- Caused by stress
- When Small Ubiquitin-like Modifier (SUMO) adds/removes an amino group to a protein, altering function / may feed into degradation
Ubiquitination
Ubiquin added to protein surface (PTM), targeting it for degredation (need ATP)
DNA Methylation
- methyl group added to DNA molecule (FOUND in some dietary sources)
- REPRESSES REGULATION AT PROMOTER
- Essential for Normal development
- Leads to:
- Carginogenesis
- Genetic imprinting
- Aging
Methylation in cancer (CPG Islands)
- Abnormal hypermethylation (but hypo is bad too!)
- Transcriptional silencing (of tumor suppressors?)
- INHERITED by daughter cell following division
