Exam 3 Review Flashcards
describe a feed forward circuit
type of network where signals pass in one direction—from input to output—without looping back. It often involves an intermediate step and helps control timing, amplify signals, or filter noise.
describe positive feedback loop
regulatory mechanism where the output of a process enhances or amplifies the initial signal. In cells, this means that once a certain molecule or gene is activated, it promotes its own continued activation, often leading to a stable, self-sustaining state
how are positive feedback loops related to cell memory
Positive feedback reinforces a response, and by maintaining activity after the trigger is gone, it helps cells remember and maintain long-term states.
what is a polysome
cluster of ribosomes bound to a single mRNA molecule, all translating it simultaneously. This allows the cell to make multiple copies of a protein from one mRNA strand at the same time, increasing efficiency in protein synthesis.
what is purpose of polysome
increase efficiency of translation
differences between hsp70 90 and chaperonins
chaperonins are used when HSP aren’t sufficient to properly fold the protein, they give the protein its own compartment to properly fold without interruption, HSP works during translation, both give a correctly folded protein
in Bulk RNA seq, what does red mean
activated genes
in bulk RNA seq, when does time start
when serum is added
what can BULK RNA seq not to
tell you individual gene expression in individual cells
what can single cell RNA seq do
tell you about heterogeneity of a cell, measures mRNA 1 cell at a time for 1000s - 10000s cells
cells close together in single cell RNA seq are what
similar, similar gene expression or cell type
what can be hard to discern with single cell RNA seq
cell state vs type
what does in situ hybridization do
tells where in organism mRNA is expressed
what binds to target mRNA in in situ hybridization , what special about them
probes, they are fluorescently labeled
what is a limit of in situ
can’t cover massive amounts of different mRNA
describe prokaryote transcription
more efficient, multiple proteins coded per mRNA, no modification
describe eukaryote transcription
more regulated, inefficient, extra coding allow for regulation
what bind to cis reg seq
transciption factors
system of survalence for viruses
CRISPR
where do coactivators and co repressors bind
transcription factors
RNA transcribed from CRISPR locus is bound to what
Cas Protein
what is in complex with Cas protein and what does it do
small crRNA, seeks out and destroys viral sequence s
list non conventional base pairs between RNA and itself
G-u, a-g, c-u
what is present at active site of RNA polymerase
Mg2+
how does RNA poly move
stepwise along DNA
energy source for RNA poly
ribonucleoside triphosphate
function in variety of nuclear processes including splicing of pre mRNA
snRNAs
help to process and chemically modify rRNAs
snoRNAs
not all apear to have function, some serve as scaffolds and regulate diverse cell processes
lncRNAs
regulate gene exp by blocking translation of specific mRNAs and causing their degradation
miRNAs
turn off gene exp by directing degradation of selective mRNAs and helping to establish repressive chromatin structures
siRNAs
RNA poly 1 transcribes what genes
5.8S, 18S, 28S rRNA
RNA poly 2 mainly transcribed what
all protein coding genes
RNA poly 3 transcribes what
tRNA genes, 5S rRNA, snRNA
in euk RNA, introns removed via what and from what
splicing, pre mRNA
what happens to3’ end of mRNA euk
3’ end cleaved and poly A tail added
describe the premRNA splicing reaction
- An adenine nucleotide in the intron
attacks the 5′ splice site, cutting the RNA
backbone. - The cut 5′ end of the intron links to the
adenine, forming a loop (lariat structure). - The free 3′-OH end of the exon reacts with
the next exon, joining them together. - The intron is released as a lariat and later
degraded into single nucleotides for
recycling
key seq for intron removal
in intron : GU at 5’ splice site, AG at 3’ splice site
what directs direct cleavage and poly A tail addition in euk mRNA
sequences in the mRNA
what is the poly a tail sequence signal
AAUAAA
what is cleavage signal for euk mRNA 3’
CA
GU rich element beyond cleavage site is bound by what
CstF
DNA probes can bind to both RNA and DNA targets. How could you design a probe that
would bind to a gene’s DNA sequence but not its mRNA sequence? How could you design
a probe to bind to a gene’s mRNA sequence but not its DNA sequence?
DNA not RNA - design a probe that binds to introns
RNA not DNA - design probe that spans exon-exon junction s
translation reads mRNA in what direction
5’ to 3’
match amino acids to codons in mRNA
tRNAs
what base pairs with mRNA codon
anticodon
amino acid attached where and to what
3’ end of tRNA
tRNAs contain unusual bases like what
pseudouridine, dihydrouridine
what enables some tRNAs to recognize multiple codons for the same AA
wobble posiiton
inosine formed by what
deamination of adenosine
aminoacyl tRNA syntheses do what
attach amino acids to corresponding tRNAs
steps of attaching AA to tRNA
Step 1: Activation – The amino acid’s carboxyl group binds to AMP, forming an adenylated amino acid.
This reaction is driven by ATP hydrolysis.
Step 2: Transfer to tRNA – The amino acid is transferred from AMP to the 3′ end of one the tRNAs that
can code for it. This forms a high-energy ester linkage, creating aminoacyl-tRNA
Incoming amino acid covalently linked to growing chain through
peptide bond
drives peptide bond formation
high energy peptide tRNA bond
what does HSP70 recognize
exposed hydrophobic regions
common promotor sequence
TATA box
where is TATA box located
about 30 nt upstream of transcription start site
TATA box recruits what for what
gen transcription factors needed for initiation of transcription by RNA poly 2
DNA sequence where general transcription factors and RNA polymerase assemble.
promotor
site for transcriptional factor/regulator
cis regulatory sequence
preferred nucleotides for specific transcription factor binding
sequence logo rep
many transcription factors form what
homo or heterodimers
euk transcription regulators form what
multiprotein complexes
noncoding RNAs in multiprotein komplexes function as what
scaffolds to stabilize protein assemblies
The repressor can bind near the activator on DNA and…
interfere with activator function, blocking coactivator recruitment
repressor can do what
stabilize an intermediate in the transcription factor assembly process
True/False: The general transcription factors are a group of DNA binding
proteins that interact as homo- or hetero- dimers with cis-regulatory DNA
sequences outside the TATA box. why
GTF help recruit RNA poly to promotor
feed forward loop can measure what
duration of a signal
a regulatory
mechanism for gene silencing and posttranscriptional control in eukaryotes.
RNAi
what guides RNAi
single strand interfering RNAs like siRNAs or miRNAs
Single-strand interfering RNAs (like siRNAs or miRNAs)
guide RNAi by…
bp w complimentary target RNAs
possible outcomes of RNAi
Cleavage and degradation of target mRNA
* Translational repression or target mRNA
* Heterochromatin formation on DNA from
which RNA is being transcribed
miRNAs transcribed as what
primary miRNAs
primary miRNAs for what
hairpin structures
primary miRNA processed into what
precursor miRNA
dicer enzyme cleaves what and forms what
cleaves precursor miRNA, generated mature miRNA
key protein in RISC
argonaute
RISC binds to what initially
both strains of the miRNA
argonauts does what
cleaves and discards one strand of miRNA, uses remaining strand to guide RISC to complementary mRNA targets
what cleaves premiRNA to get mature miRNA
dicer enzyme
what determines the outcome of what RISC is doing
extend of bp btw miRNA and mRNA
miRNA regulation:extensive base-pairing leads to
mRNA cleavage and degradation
miRNA regulation : partial bp results in
inhibition of translation, eventual mRNA degradation
what cuts dsRNA to turn it into siRNA
dicer
introduction os dsRNA is for what
gene knockdown, gets turned to siRNA then taken up by argonauts and RISC
Cas9 binds what
guide RNA
guide RNA contains what regions
one required for Cas9 binding, sequence matching specific genome target
Cas9 creates what in the targeted genomic site
double strand break
double strand break by Cas9 usually repaired by what , can introduce what
non homologous ends joining, mutations
what can be used in the double strand break to precisely edit, and what does this enable and require
homologous recombination, requires a altered repair template, which is supplied by the experimenter, enables to alter genes
mutant Cas9 - properties and used for what
cannot cleave DNA, used for gene regulation
mutant Cas9 fused to activator does what
turn on dormant gene
How would you expect the usage of rare codons in a coding
sequence to affect protein translation? Why?
Rare codons are likely to have lower translational efficiency as tRNAs
may be less abundant for that codon
What experiment would you perform to test this?How would you expect the usage of rare codons in a coding
sequence to affect protein translation?
Make a optimal codon versus a rare
codon version of the protein in an expression vector. Have cells
express each version and measure resulting protein output
Under what circumstances would using a rare codon be
advantageous?
This could be used to reduce the output for this particular protein that
is meant to be at low levels. Also could make it responsive to the
upregulation of a particular tRNA. Enables modular expression of a
set of proteins based on tRNA regulation
Viruses, which rely on the translational machinery of the host cell, can
evolve to infect other hosts.
a)How might codon usage bias contribute to the success of a virus in
a particular host?
Virus would use codons for efficient replication in that host by
matching its codons with those most frequently used by host cells. 1
point
How do you hypothesize codon usage could change when viruses
crossover to a new host?
Virus would alter the codon usage to better match the new host after
the acquiring the ability to infect that new host
What experiment would you perform to test this hypothesis?: Virus would alter the codon usage to better match the new host after
the acquiring the ability to infect that new host
Look at examples of host-species
transitions in viruses and analyze sequence of the virus before and
after the transition. Compare with codon frequency tables to see if
codon usage shifted to more closely resemble the codon usage bias
of the new host
What factor(s) do you hypothesize can contribute to this and why: Codon usage is not equal within a species showing some are used
more frequently than others. Across species, codon usage can also
vary for the same amino acid
Differences in GC content, differences in the abundance of particular
tRNAs in a cell.
You synthesize the sequence, but the probe still does not work. Explain why the
probe sequence does not effectively bind to any RNAs.
The probe forms a
hairpin structure with itself. This prevents it from effectively binding the target region.
If you want to use genomic DNA to PCR amplify a region for generating the
probe, which regions of the gene should you target and why? Which regions
should you not target?
You can target exonic regions but want to avoid
intronic regions. Do not cross exon-intron junctions with the amplicon
Differential exon splicing? does what
Generates a variety of polypeptide sequences from the same sequence
what do bacteria not have that eukaryotes have
intron processing mechanisms
polypeptide chain is synthesized from….
N terminus to C terminus
determine whether cells uniformly regulate gene transcription in
response to a signal or have different responses to the same input
single cell RNA seq
used to identify sets of coordinately regulated genes across samples
bulk RNAseq
Difference between prokaryotic and eukaryotic mRNA?
Prokaryotic mRNA remains unmodified at
the 5’ and 3’ ends
Know how mRNA and the ribosome interact with each other- reading from…
5’ to 3’
a techniques can test the sufficiency of cis-regulatory DNA sequence in
promoting gene expression
gene reporting
