LEC22: Chromatin and Gene Structure Flashcards
what are the molecules that make up chromatin?
DNA + histone octamer core (H2A, H2B, H3, H4) + H1
what is the direction of nucleic acid synthesis?
in which direction is the template strand read?
strand is read 3’ to 5’
synthesis is 5’ to 3’
name the types of sequences that are present in a pre-mRNA transcript
exons, introns
how can DNA in double strand be bound by proteins?
1) binding between amino acid side chains of alpha helices of a DNA binding protein, which sit on DNA, bind to DNA bases; this is **protein a.a. sequence has high specificity, **plugs into DNA sequence w/ its available H-bonding
(specific amino acid - base sequence pairing)
2) a homotetram (4 identical proteins that are binding a particular DNA sequence) have a specific binding site on DNA, it’s kinky, they bind in a specific spot therefore
(overall DNA sequence is recognized by DNA binding proteins)
what is RNA’s secondary structure?
how does it form?
what is it called?
what base pairs form?
RNA usually = single stranded
can have 2ndary linear, single strand of RNA fold up on itself via complementary base-pairing within the strand
this forms a stem-loop structure of intramolecular double stranding
A-U, G-C, G-U also possible
what is this
RNA secondary structure - stem-loop
intramolecular double stranding
what kind of genome does HIV have? why is this significant re: structure of that genome?
RNA genome
lots of secondary structure required for proper functioning of this RNA genome
what is DNA denaturation?
heating up a double stranded DNA molecule separates the 2 strands b/c they are only held together by H-bonds
what happens if you denature 2 strands of double stranded DNA molecules in a test tube and then lower the temperature?
why does this occur?
raise temperature: strands come apart
lower temperature: complementary strands will find each other again, get renaturation back to what you had originally
this is bc of complementary base pairing
what is hybridization?
complementary base pairing between 2 nucleic acid strands which are not naturally together
how does a probe work?
probe = complementary piece of nucleic acid (DNA usually, can be RNA though)
make it complementary to the sequence that you are trying to locate in your sample
if put probe in with mixed up sample, denature, and then renature, it will bind if its complement is present; and can have it signal if binding occurs
this is nucleic acid hybridization
what is the nucleosome
packing mechanism that allows 2 meters of DNA to get into a 10 micron nucleus
nucleosome = double stranded DNA (150 bp) + histone proteins core
what is teh structure of the histone proteins of the nucleosome?
2 molecules each of H2A, H2B, H3, H4, and 1 H1 that staples DNA to that core protein octamer
what comprises a nucleosome
DNA-histone protein unit
what is chromatin
DNA in its bound form to histones in the nucleus
what is euchromatin vs heterochromatin? where is each found?
euchromatin: transcriptionally active region of chromosome
heterochromatin: tightly wound, dense structures of chromosome that’re inaccessible to RNA polymerase
what is centromere
connects chromatins
what is telomere
at end of chromosome
what are the processes of nucleic acid synthesis?
1) DNA replication
2) Transcription
in DNA replication, which strand(s) is/are used as template(s)?
what does the replication?
what direction is the new product synthesized? which direction is the unit being replicated read?
what do you end up with?
both strands are used as templates
DNA polymerase (DNA dependent DNA polymerase) reads nucleotide sequence of template strand in the 3’ to 5’ direction, and synthesizes new DNA in the 5’ to 3’ direction
result: 2 identical double stranded DNA molecules
what is teh basis of DNA replication?
complementarity
what does transcription? are both strands templates? how is the template read, how is RNA synthesized? what is the product resulting?
done by RNA polymerase, a DNA dependent RNA polymerase
only 1 template strand, 1 is non-template
RNA polymerase reads template strand from 3’ to 5’, makes RNA in 5’ to 3’ direction
product: 1 single stranded RNA molecule
newly synthesized RNA transcript has same sequence as what?
same as the non-template strand of the DNA that was transcribed, but has U’s not T’s
what problem does the tight packing of DNA present?
what determines if DNA will be expressed?
generates problem of accessibility to the DNA by proteins that regulate transcription or are engaged in replication
histone modifications - acetylations, methylations, phosphorylations - determine if DNA info will be expressed
what are the substrates of DNA polymerase?
the end of the previously replicaed DNA
and
incoming deoxyribonucleoside triphosphates (dATP, dCTP, dGTP, dTTP)
what are ribonucleoside triphosphates?
ATP, CTP, GTP, UTP
are what RNA polymerase incorporates into growing RNA chain as it transcribes DNA info into RNA
what’s another name for the DNA non-template strand? why?
“coding strand’ because it has the same sequence as the RNA that is transcribed from the DNA template strand
what is the +1 site?
the base pair where txn initiation occurs
aka “TSS” “transcription start site”
what is the transcription unit?
what are its boundaries?
what happens to it and does it result in?
the piece of DNA that is transcribed by RNA polymerase
from +1 site to terminator sequence
acted on by RNA polyermase
makes pre-mRNA
what are the steps of pre-mRNA processing? what is the result?
1) splicing: exons are knit together, introns are spliced out
2) capping: 5’ CAP is put on pre-mRNA
3) poly-adenylation: poly-AAAAA tail put on 3’ end of pre-mRNA
what’s splicing
during pre-mRNA processing, pre-mRNA is spliced
introns are spliced out (“intervening”); exons stay in (“expressed”) to the mRNA
no functional consequence of this for the DNA, only for the pre-mRNA and mRNA
what does poly adenylation onto what?
a DNA-independent RNA polymerase adds a poly-A tail on to the 3’ end of mRNA
where does mature mRNA go?
cytoplasm
what is the open reading frame?
what are its features?
the part of the mRNA that ribosome will actually translate, code for a protein
begins w/ AUG, codon for methionine
ends w/ UAA, UAG, UGA, stop codons
what are the ends of the mRNA?
5’ untranslated region, 3’ untranslated region poly-A tail
whole thing is:
5’UTR, coding sequence (ORF), 3’UTR, poly-A tail
what is a codon
a string of 3 nucleotides
encodes for an amino acid
what does it mean that the code is degenerate?
multiple codons code for the same amino acid
what do RNA Pol I, II, and III transcribe for?
RNA Pol I: rRNA genes
RNA Pol II: protein-coding genes!
RNA Pol III: tRNA genes
what is a silent mutation?
occurs b/c code is degenerate so 1 base change may still code for same amino acid
**does not effect function **
what is a missense mutation?
single nucleotide change results in codon that codes for a different amino acid
may or may not effect function depending on how different amino acid that’s put in is
what is a nonsense mutation?
point mutation that results in premature STOP codon in coding sequence
results in truncated (shorter), nonfunctional protein
almost always effects function
what is a frameshift mutation?
insertion or deletion of a nucleotide/nucleotides in a DNA sequence that isn’t divisible by 3, so shifts entire ORF
resutls in totally different translation from original
the earlier in the sequence the deletion/insertion occurs, the more altered the protein
causes reading of codons after the mutation to code for different amino acids
what is the first nucleoside in pre-mRNA? why?
a nucleoside 5’ triphosphate
b/c RNA synthesis can begin de novo
how does RNA pol II know where to start txn?
needs help of general transcription factors (TFII complexes of TFIIB, D, E, F, H) and mediator which guide RNA Pol II to sit down on DNA, start txn
guide the RNA Pol II to the TATA region
what is a cis vs. trans element?
cis: something on teh DNA itself, intrinsic pt of DNA
trans: soemthing that binds DNA from outside, has an effect on it
what is the promoter?
a cis element of the DNA
where txn begins
TATA sequence of promoter is notable
what is a consensus sequence?
an example?
a sequence that is most often found at a particular position
TATA is an example of this
where is the TATA sequence located?
-25 or -30 location, upstream of +1 site where txn begins
what is TBP? what does it do?
TBP: TATA box binding protein
recognizes the TATA sequence and so binds, has TFIID attached and allows binding of TFIID to DNA at initiation site
causes a bend when binds
what is characteristic of the carboxyterminal domain of RNA Pol II’s largest subunit?
multiple repeats of a short sequence highly enriched in serine
gets phosphorylated by TFIIH, a protein kinase
this phosphorylation singals a switch to elongation mode
how does RNA Pol II get recycled? why?
gets recyled for repeated rounds of txn
protein factors rearrange elongating txn complex such that RNA Pol II is released from DNA template, is returned to its hypophosphrylated state by a CTD phosphatase
