Functions & Dysfunctions of Genetic Regulation Flashcards
Central Dogma of Genetics
DNA–> RNA–>protein (via transcription, translation)
Structure of DNA
double helix, sugar/phosphate backbone w bases (AGCT); antiparallel strands
Euchromatin
protein involved w DNA Packaging, ubiquitous within DNA, almost always activated
Heterochromatin
condensed DNA, found mainly in centromeres and at the edges, very little gene expression
methylation on packaging
adding a methyl group
base pairing
A and T have 2 double bonds, G and C have 3 (stronger); 142 h-bonds b/w DNA and histone octamer
DNA packaging
DNA wraps around a histone w condensed chromosomes that prevent mutations and protect against damage
Structure of histone
Proteins made up of a lot of AA but specifically many Lysine and Arginine (basic, positively charged) that attract the negatively (from phosphate groups) charged DNA
Mnemonic: Lysine Arginine Histone
Human genome project
Began in late 90’s early 2000’s and took a very long time to finish and it was only 90% done; cost a billion dollars whereas now it takes about 24 hours and $1000
Personalized medicine
Uses each patient’s individual DNA to hybridize information
Binding proteins to DNA
1) Histones
2) non-histone binding
Nucleosomes
Basic unit of chromosome packing (includes histone and non-histone portion); 8 histone proteins (histone octamer)
Chromatin
Protein + DNA
Position effect
Gene activity is based on location on the chromosome
If a previously actively expressed gene is moved near the heterochromatin (centromere or telomere)—> becomes silenced
Alternative RNA splicing
Exons/introns can be cut differently on the RNA to produce different gene combos (avg: 2 different ways/ genes)
Histone Deacetylation (HDAC)
HDAC activity represses gene activity by removing acetyl group and resestablishing normal DNA-histone interaction
Histone Acetyl Transferase (HAT)
Add an acetyl group to histone (N-terminal lysine) which reduces DNA interaction because the positive charge is removed;promotes gene expression as its not as tightly wound
Ex: estrogen (steroid hormone) promotes gene expression
DNA Methylation
Adding methyl group to DNA (cytosine and adenine) by specific methyl transferase enzymes
Change activity but not DNA sequence (ex: will repress gene expression when its at a gene promoter)
Key processes affected: genomic imprinting, xchromosome inactivation, repression of transposable elements, aging, carcinogenesis
Hypermethylation
**important in Cancer Development
Gene promotes will lead to silencing/target
Hypomethylation
Too little methylation—> inactivate chromosome and loss of imprinting abilities
DNA Replication
DNA polymerase (DNA-dependent) synthesizes new DNA from 5’-3’; NEEDS free 3’ OH
New chain is assembled based on the template of the old chain (needs to be separated and then double are made)
Requires dATP, dGTP, dCTP, dTTP
Replication fork
Origin & direction of expansion
Semi-discontinuous
HAS to go from 5’—3’ but not both sides can be the correct way sooo one side is leading and one is lagging
Lagging strand goes from 3’-5’ of ORIGINAl but is made 5’-3’ in Okazaki fragments
Helicase
Unzips & unwinds helix by using ATP to bind and conformationally change
Topoisomerase
Uncoils the tangled coils by breaking phosphodiester bonds
Target for anticancer drugs because they block the cell cycle and generate breakage which harms the entire genome—> apoptosis and cell death
Single stranded binding proteins
Sit on the single strand of the DNA helix to keep it open and expose the bases and prevent formation of hair puns, stabilizes the DNA
DNA Ligase
Glues back together the pieces of Okazaki fragments
DNA polymerase
Adds DNA to replicating DNA
Nucleoside analogues inhibitors
These lack the 3’ OH group and act as drugs that can inhibit DNA replication (anticancer); need to be converted to dNTPs before they can actually inhibit
Ex: ara-c, acyclovir, azidothymidine
Spontaneous damage
Depurination, Deamination
Depurination
Essentially: removal of purine (adenine or guanine)
Due to cutting off n-glycosidic bond
Deamination
Amino group of either base is hydrolyzed and adenine turns to hypoxathine, guanine turns to xanthine and cytosine turns to uracil
Ionizing Radiation
ex: xrays
Change base damages, directly break strands, DNA-protein cross links
Non-ionizing radiation
Ex: UV light
Add a covalent bond between adjacent pyrimidine bases—> toxic, mutagenic and carcinogenic photoproducts
Metabolic activation
Ex: (Benzo(a)pyrene and aflatoxin B1)
Activated by hepatic cytochrome P450 and eventually forms a bulky adduct w exocyclic amino which prevent replication and gene expression
Agents that direct modify DNA
Ex: crosslinking agents, alkylation agents and intercalating agents
Crosslinking agents
cross links between DNA bases either in the same strand or complementary strands
Ex: nitrogen mustard, crisp Latin, carmustine, mitt ya in C (anticancer)
Alkylating agents
methylated base changes and changing phosphodiester into phosphtriesters
Ex: DMS and MMS
Products include: 7-methylguanine, 3-methyladenine
Intercalating agents
Come in between stacked bases of DNA double helix—> unwinding and separating base pairs
Ex: ethidium bromide, thalidomide and aanthracycline antibiotics
Thymine modifications
In CpG sites/islands (cytosine-phosphate-guanine) and it replaces a T w a G—> stably silences the genes so it’s useful for cancer cells
Direct repair mechanism
Repairs cyclobutane pyrimidine dimers by using DNA photolyase which binds to the dimer, absorbs the light and then breaks the bond (photoreactivation)
Direct Repair Mechanism
Of O6-methylgaunine
Uses methylguanine methyltransferase to move methyl from guanine to the cysteine and removes damage
Base excision repair (BER)
Repairs single base mismatches and small alterations
Uses: DNA glycosylases (they can flip out th base)
Recognize wrong base, DNA Glycosylase comes in and hydrolyzes the N-glycosidic link to remove the altered base, recognized by AP endonuclease —> cuts the phosphodiester bond, AP Lyase removes it; DNA Polymerase B and DNA ligase replace it and glue it back together
Nucleotide excision repair (NER)
Repairs chemicals that alter or distort normal change
Ex: fixes UV light induced pyrimidine dimers, BPDE from chemical agent mutation, cisplatin additions
Distortion is detected, unwind double strand,nick in either side of the damage, nucleotides (29 for humans, 13 in E.coli) are removed
DNA polymerase e (DNA polymerase I in Ecoli fill in the gap using the undamaged side as the template and then glued together using ligase
Clinical: xeroderma pigmentosum
Mismatch excision repair
When a wrong nucleotide is inserted—> wrong base pairing= wrong number of hydrogen bonds=deformity
Binds to DNA at deformation and and distinguish between parent and daughter to identify daughter cell; endonuclease comes in to cut the daughter strand and helicase/exonuclase removes a segment of DNA that has the mismatch, polymerase fills the gap, ligase seals the nick
Deficiencies: increase susceptibility to hereditary non-polyposis colorectal cancers
Recombination repair
Due to ionizing radiation or anti cancer agents—> breaking of double strand or inner strand cross linking
Homologous/Nonhomologous
Nonhomologous end joining recombination repair
Nonhomologous end joining: ends are joined together by DNA ligase which fixes the break in double strand but leaves a few bases out; preferred method w translocation or deletions
Homologous repair
Homologous recombination: when the DNA breaks after replication but before division —> helices are close together
Broken ends have 3’ overhang, single stranded region invades good copy and pairs it w the complementary strand, DNA polymerase begins filling in the gaps and then they’re glued back to the 5’ ends
Transcription-coupled repair
RNA polymerase stalls during transcription
(In eurkaryotes: ERCC-6 and ERCC-8 recognize it and bring in other enzymes to help fix)
Defective ERCC-6 and ERCC-8–> Cockayne syndrome =growth defects
Translesion synthesis
Uses DNA polymerase n and some weird shape to fix thymine dimers or apurini AP sites
Xeroderma pigmentosum
Skin is extremely sensitive to light and are more sensitive to direct sunlight and are prone to developing melanomas and sarcomas because UV light usually makes dimers which can be fixed in normal people w NER but these have defects in XP proteins in the NER
Hereditary nonpolyposis colorectal cancer
Mutations in MER complex—> no good copy and then the MER complex cant fix the tumors
Cockayne Syndrome
Rare autosomal recessive congenital disorder
Biochem: mutations in the ERCC6 and ERCC8 in TCR of DNA
Developmental and neurological delay, photosensitivty, premature aging, hearing loss, eye abnormalities
BRCA mutations and breast cancer
BRCA1 and BRCA2 (breast cancer susceptibility genes)—> tumor suppressor genes
Mutations= bigger risk of women developing cancer
Mutations= increased risk of breast cancer for men’s
BRCA1=increase risk of cervical, uterine, pancreatic and colon cancer in women, and pancreatic testitcular and prostate in men
BRCA2=risk of melanoma, pancreatic stomach, gall bladder, bile duct cancer=women
Methylation of CpG islands
Stable silence of genes
Occurs at 5 of the pyrimidine ring of Cytosine (within CpG sites)—> 5-methylcytosine
UPS Protein Degradation Pathway
Sustains homeostasis
Oxidative stress, cellular differentiation, varying nutrient supply, elevating/reducing temperature, stress
Ubiquitin
Small amino acid that is in all cells and sticks to the lysine residues on the target proteins; attach to each other to form chains which are targeted for destruction
SUMOylation Cycle
Sumo exists as a precursor w two glycines that will chop off the XX part
Resevouir of modification—> active from that gets integrated into various substrates
SUMO/Ubiquitin
NF4 will stick ubiquitin onto proteins and has some SUMO interacting motifs to bind sumos
