Biochemistry - Cellular Structure & Biomechanics Block (I) Flashcards
Euchromatin is known as ‘beads on a string’ due to its arrangement as nucleosomes connected by linker DNA. These fibers are commonly referred to by their width.
What width does euchromatin have?
11 nanometers
The weak forces connecting DNA to histones are primarily produced by electrical interactions between what structures?
DNA phosphate groups (-) and arginine and lysine side chains (+)
What are the proteins making up the histone octamer?
H2a (x2)
H2b (x2)
H3 (x2)
H4 (x2)
What histone protein is not part of the histone octamer?
H1
One fiber of euchromatin is ___ nm in width.
One fiber of heterochromatin is ___ nm in width.
11;
30
What portion of the nucleosome is the site of most modifications that allow for gene activation and silencing?
The histone tails
What major nucleosome modifier is responsible for decondensation (unwinding) of chromatin?
What effect will this have on the genes involved?
Acetylation;
increased gene expression
How does acetylation of the histone tails (specifically lysine residues) cause decondensation of the chromatin?
It neutralizes some histone positive charge
What enzyme is responsible for histone acetylation?
What enzyme is responsible for histone deacetylation?
Histone acetyltransferases (HATs);
histone deacetylases (HDACs)
Histone acetyltransferases (HATs) have what effect on chromatin?
Histone deacetylases (HDACs) have what effect on chromatin?
Decondensation (11 nm fibers);
condensation (30 nm fibers)
Histone acetyltransferases (HATs) have what effect on gene expression?
Histone deacetylases (HDACs) have what effect on gene expression?
Increased gene expression;
gene silencing
What effect does methylation typically have on chromatin?
Gene silencing
How does histone methylation cause gene silencing?
By preventing acetylation
What is the typical effect of histone phosphorylation?
To promote chromatin decondensation (unwinding)
Which histone modifications will typically result in chromatin decondensation (unwinding) and increased gene expression?
Acetylation (by HATs);
phosphorylation
Which histone modifications will typically result in chromatin condensation (coiling) and gene silencing?
Deactylation (by HDACs);
methylation
For what are SWI/SNF responsible in gene expression?
How?
Regulation of chromatin decondensation;
through HAT function
For what is H1 responsible in gene expression?
How?
Facilitating DNA condensation;
through HDACs
Which activators regulate HAT function and chromatin decondensation?
Which activators regulate HDAC function and chromatin condensation?
SWI/SNF;
H1
What is the function of the MECP2 protein?
To bind methylated DNA (not methylated histones)
and recruit HDACs
DNA methylation (as opposed to histone methylation) will result in activation of what protein and what effect?
The MECP2 protein;
HDAC recruitment;
chromatin condensation
What is the shortest phase of the cell cycle?
What phase is significantly shortened in rapidly dividing (e.g. tumor) cells?
M;
G1
How many chromosomal autosome pairs are there?
How many chromosomal sex pairs are there?
22;
1
What stain is often used in karyotyping so different chromosomes can be distinguished from one another?
What segments are marked using this staining process?
G-banding;
A-T rich sequences
Define euploidy.
Correct number of chromosome pairs
What is the term if an embryo is missing both chromosomes of one chromosomal pair?
What is the term if an embryo is missing one chromosome of a chromosomal pair?
What is the term if an embryo has both chromosomes of one chromosomal pair?
What is the term if an embryo has an extra chromosome in addition to one chromosomal pair?
Nullisomy (-2);
monosomy (-1);
euploidy (0; just right);
trisomy (+1);
What enzyme cuts specific palindromic DNA sequences?
Restriction endonucleases
DNA of interest can be recombined into a plasmid after cutting palindromic sequences on both using what type of enzyme?
What enzyme then seals their exposed palindromic sequences?
What then happens in the bacteria containing this recombined DNA?
An endonuclease;
DNA ligase;
replication (of the cell and DNA)
What in particular can in-situ hybridization be used to analyze?
Gene expression (through antibodies that show mRNA and/or protein expression)
What specialized base pairs are useful for Sanger DNA sequencing? What is important about the base pair insertions?
Dideoxynucleotides (ddATP, ddGTP, ddCTP, ddTTP);
one must attach after each base pair of the target sequence, creating sequences of all lengths from the full sequence to only one base pair
What is PCR good for?
Amplifying DNA sequences using a system of primers, heating, and Taq polymerases
How do we know that genetic polymorphisms are not simply new mutations?
They are present in ≥1% of the population
Per every 1 billion base pairs copied, how many mutations occur?
~1
What general term can be used to describe DNA replication in regards to the way older DNA strands are distributed amongst newer strands as replication occurs?
It is semi-conservative
True/False.
DNA is both antiparallel and complementary.
True.
What does it mean that DNA replication is semi-conservative?
Each new DNA duplex model is composed of one parent strand and one daughter strand
DNA origins of replication are characterized by a richness of what type of base pairing?
A-T pairings
All DNA replication occurs in what direction?
5’ to 3’
What structure is used to provide a 3’-hydroxyl group so that DNA replication can initiate after the strands are denatured from one another?
An RNA primer
What enzyme ‘unwinds’ DNA helices from one another?
What enzyme releases the tension this creates by snipping and rejoining the strands?
What proteins prevent reannealing of the strands?
Helicase;
topoisomerase;
single-stranded binding proteins
In-vitro DNA replication mutations are 1 out of every 10,000 base pairs. In-vivo human replication mutations are 1 out of every 1,000,000,000 base pairs.
What explains this discrepancy?
Extensive DNA repair systems
What bacterial DNA polymerase is the main one synthesizing new strands?
DNA polymerase III (bacterial)
What bacterial DNA polymerase replaces RNA primers with DNA?
DNA polymerase I (bacterial)
What bacterial DNA polymerases are most involved in DNA repair?
DNA polymerase I and II (bacterial)
How does DNA polymerase I (bacterial) remove RNA primers to be replaced with DNA?
3’ to 5’ exonuclease activity
What enzyme connects Okazaki fragments?
DNA ligase
How many replication origin points do eukaryotes have in each contiguous piece of DNA?
(1 or many?)
Many
Where is the hydroxyl group that the triphosphate bond attacks in DNA polymerization?
(I.e. hydroxyl groups of what prime are attacked by phosphates of what prime?)
3’-hydroxyl
5’-phosphate
For what is eukaryotic DNA polymerase Alpha useful?
Synthesizing short stretches of DNA, works with RNA primase to make RNA primers
For what is eukaryotic DNA polymerase Delta useful?
Takes over after RNA primers; lagging strand synthesis (but also can be leading)
For what is eukaryotic DNA polymerase Epsilon useful?
Leading strand synthesis (but also can be lagging)
For what is eukaryotic DNA polymerase Beta useful?
DNA repair
For what is eukaryotic DNA polymerase Gamma useful?
Mitochondrial DNA replication
What enzyme is able to ‘read’ RNA in order to synthesize new DNA?
What types of infectious agents are likely to have this enzyme?
Reverse transcriptase;
retroviruses (e.g. HIV)
DNA synthesis occurs in what direction?
5’ to 3’
What bacterial enzyme removes and replaces the RNA primers of DNA replication?
How?
DNA polymerase I;
5’ to 3’ exonuclease activity
DNA ligase forms what type of bond?
3’ to 5’ phosphodiester bonds
Which bacterial enzyme is able to perform both 5’ to 3’ DNA synthesis and 3’ to 5’ exonuclease activity?
DNA polymerase I
DNA proofreading by bacterial DNA polymerase I occurs in what direction?
3’ to 5’
How is bacterial DNA polymerase I able to perform both 5’ to 3’ synthesis and 3’ to 5’ exonuclease activity?
There are two separate active sites on the enzyme
What enzyme fills in the space left behind by RNA primers at the end of genetic sequences?
Why is this important?
Telomerase (TTAGGG);
to protect against chromosomal shortening
Why is telomerase needed in DNA replication?
The final Okazaki fragment of the lagging strand has a gap at the end where no RNA primer 3’-hydroxyl is be placed as the space is shortened
Which enzyme(s) is(are) responsible for processive DNA synthesis in bacteria?
Which enzyme(s) is(are) responsible for processive DNA synthesis in eukaryotes?
DNA polymerase III;
DNA polymerase delta, DNA polymerase epsilon
DNA is synthesized in what direction?
The template used for synthesis is read in what direction?
5’ to 3’;
3’ to 5’
What are a few exogenous sources of DNA damage?
Ionizing radiation;
UV rays;
environmental chemicals
What are a few endogenous sources of DNA damage?
Reactive oxygen species (ROS);
uncorrected errors of replication
Chemical and ROS damage to DNA often results in what sort of structural changes?
UV damage to DNA often results in what sort of structural changes?
Depurination, deamination;
thymidine dimers
Deamination of cytosine leads to:
Uracil
What are three forms of DNA repair?
(the three ‘excision repairs’)
Base excision repair
Nucleotide excision repair
Mismatch excision repair
Deamination of 5-methylcytosine leads to:
Thymine
How are basepairing errors and other minor DNA errors corrected?
What is a common example of a new basepairing error?
Base excision repair;
5-methylcytosine is deaminated to thymine
(C to T)
How are thymidine dimers and other large DNA lesions corrected?
Nucleotide excision repair
What is the most common form of basepair error?
What type of DNA repair mechanism would address this?
Cytosine to thymine mutations
(and subsequent T-G base pairs);
base excision repair
How much of the DNA strand is removed in base excision repair?
Just the errant base
How much of the DNA strand is removed in nucleotide excision repair?
Often several nucleotides in sequence
How much of the DNA strand is removed in mismatch excision repair?
Often several nucleotides in sequence
What are the two endonucleases of nucleotide excision repair?
XP-F and XP-G
What is the rate-limiting enzyme of nucleotide excision repair?
What gene codes for it?
XP-F;
ERCC1
Cisplatin is used to damage DNA and induce apoptosis.
What defense mechanism is fixing this damage?
Cells overexpressing what gene will be resistant to cisplatin?
Nucleotide excision repair;
ERCC1 (XP-F gene)
What type of DNA repair mechanism catches uncorrected errors of replication?
Mismatch excision repair
What is the difference between nucleotide excision repair (NER) and mismatch excision repair (MER)?
NER corrects large defects (e.g. thymidine dimers);
MER corrects errors of replication (e.g. C-A pairing)
Which of the three forms of DNA excision repair (base, nucleotide, mismatch) involve(s) helicase, endonucleases, polymerase, and ligase in removing sequences of DNA (i.e. more than one nucleotide) and refilling them?
Nucleotide, mismatch
(base only removes a single, solitary base)
What are the two types of gene (based on whether the gene is virtually always active and needed or not)?
Constitutive;
regulated
What is a constitutive gene?
(as opposed to regulated)
A gene that is always active in virtually all cells
(needed for basic cell survival)
What is a regulated gene?
(as opposed to constitutive)
A gene that is only needed sometimes;
they are turned on and off according to cellular needs/signals
Constitutively expressed genes are always active and needed. They operate at a _______ level that is then induced or repressed as needed.
Basal
What type of prokaryotic protein represses gene expression by interacting with the gene promoter?
An oppressor protein
What type of prokaryotic protein increases gene expression by interacting with the gene promoter?
A gene activator
To what area of the gene do opressor proteins bind?
The repressor region
To what area of the gene do activator proteins bind?
Just adjacent to the promotor
What is typically the mechanism by which oppressor and activator proteins are activated or inhibited in prokaryotes?
Single effector signaling molecules
Regulation of the lactose operon depends on surrounding levels of what substances?
Lactose;
cAMP (glucose-dependent);
glucose
cAMP binds what modifier to the lac operon?
Lactose binds what modifier to the lac operon?
The activator (CAP) (inducing gene transcription);
the repressor (removing it and inducing gene transcription)
What will be the state of transcription of the Lac operon in the following situation?
Lactose low
Glucose high
Low cAMP
No transcription
What will be the state of transcription of the Lac operon in the following situation?
Lactose high
Glucose high
Low cAMP
Some transcription
What will be the state of transcription of the Lac operon in the following situation?
Lactose high
Glucose low
high cAMP
High transcription
What scenario would lead to no lac operon transcription?
No lactose
Yes glucose
Low cAMP
What scenario would lead to a small amount of lac operon transcription?
Yes lactose
Yes glucose
Low cAMP
What scenario would lead to high lac operon transcriptional activity?
Yes lactose
No glucose
High cAMP
What is the eukaryotic equivalent of the prokaryotic promoter?
The proximal promoter
Where is the eukaryotic equivalent of the prokaryotic TATAAT sequence?
What is it called?
-30 BP;
TATA box
What eukaryotic protein binds the TATA box to organize the transcriptional process and identify the strand for usage?
TATA-binding protein (TBP)
How does most gene activation occur?
Which is more common, gene activating proteins or gene silencing proteins?
Heterochromatin to euchromatin changes;
activating (most silencing is simply the genome remaining as heterochromatin)
What is the name of the eukaryotic helicase that unwinds the DNA for transcription to take place?
What other activity does this enzyme have that allows it to activate the preinitiation complex and induce transcription?
TFIIH;
serine kinase activity
Phosphorylation primarily occurs on which histone’s tail?
Acetylation primarily occurs on which histone’s tail?
Methylation primarily occurs on which histone’s tail?
H2a
H2b;
H3
(remember PAM 2,2,3)
What is the important region of the prokaryotic promoter region to know?
If the gene of interest is +1, what is the region specified above?
TATAAT;
-10
Describe eukaryotic transcription initiation.
TFIIB and TATA-binding protein (TBP) bind what sequence and recruit what?
The TATA unit (-30);
the PIC (at the INR [-10])
What part of RNA polymerase II is phosphorylated/activated by TFIIH and is also involved in hnRNA processing into mRNA?
The carboxy-terminal domain (CTD)
How can repressing proteins block eukaryotic transcription?
Inhibiting: the TATA-binding protein (TBP),
the preinitiation complex (PIC),
recruiting: histone deacetylation complexes (HDACs)
The template strand is also known as the ________ strand or the ________ strand.
noncoding;
antisense
(Templates Are Nonsense)
The noncoding strand is also known as the ________ strand or the ________ strand.
antisense;
template
(Templates Are Nonsense)
The antisense strand is also known as the ________ strand or the ________ strand.
template;
noncoding
(Templates Are Nonsense)
The TATA box is how many base pairs prior to the gene of interest (+1)?
And the initation region (INR)?
- 30;
- 10
True/False.
RNA primers are used in DNA synthesis in vivo and in vitro.
True.
Western blots probe what?
Northern blots probe what?
Southern blots probe what?
Protein;
RNA;
DNA
Which blot is useful for probing RNA?
Which blot is useful for probing protein?
Which blot is useful for probing DNA?
Northern;
Western;
Southern
When initiating transcription, what enzymes are actively ensuring the gene is open and available?
What enzymes are inactivated at this time?
HATs;
HDACs
What protein is responsible for the bending of DNA and the gene sequence in question into hairpin turn when transcription is being initiated?
HMG1
What transcription factor is responsible for recruiting the transcription helicase (TFIIH)?
What factor identifies and binds the TATA box?
What transcription factor is responsible for phosphorylating the CTD of RNA polymerase II and also unwinding the DNA strands?
TFIIE;
TATA-binding protein (TBP);
TFIIH
What is the central dogma?
DNA — (transcription) —> RNA — (translation) —> Protein
Addition of the _________ cap to the 5’ end of hnRNA and a poly-A tail to the 3’ end are both associated with what portion of the RNA polymerase II that coded the hnRNA strand?
7-methylguanosine;
the carboxy-terminal domain (CTD)
What is the name for the raw RNA strand transcribed from DNA?
This is modified to become what?
hnRNA;
mRNA
What must occur for hnRNA to become mRNA?
5’ cap;
3’ poly-A tail;
splicing out of of introns
___________ are made of various proteins including snRNPs.
Spliceosomes
Via what shape do spliceosomes remove hnRNA introns?
Lariats
What molecule provides the methyl group to 7-MeGuanosine?
This molecule is replenished by what vitamins?
S-adenosylmethionine;
folate, B12 (via methionine)
What enzyme adds the poly-A tail to the 3’ end of an hnRNA?
What nucleotide sequence does it follow?
Poly-A polymerase;
AAUAAA
Will nucleopores allow hnRNA to leave the nucleus?
Will nucleopores allow mRNA to leave the nucleus?
No;
yes
What are the two portions of a eukaryotic ribosome?
Which binds to the mRNA first?
60s, 40s;
40s
What proteins are responsible for connecting the 40s ribosomal subunit to an mRNA strand?
What is their energy source?
Eukaryotic initiation factors (eIFs);
GTP
What are the four main eukaryotic initiation factors (eIFs)?
What is their overall goal?
eIF2, eIF3, eIF4a, eIF4b;
unite the 40s ribosomal subunit with an mRNA
What does the eIF2 (eukaryotic initiation factor 2) do?
What does the eIF3 (eukaryotic initiation factor 3) do?
What does the eIF4a (eukaryotic initiation factor 4a) do?
What does the eIF4b (eukaryotic initiation factor 4b) do?
Facilitates 40s ribosomal subunit binding;
binds 40s;
removes 2° mRNA structures (it is an RNA helicase);
locates AUG
What does the eIF2 (eukaryotic initiation factor 2) do?
Facilitates 40s ribosomal subunit binding
What does the eIF3 (eukaryotic initiation factor 3) do?
Binds 40s
What does the eIF4a (eukaryotic initiation factor 4a) do?
Removes 2° mRNA structures (it is an RNA helicase)
What does the eIF4b (eukaryotic initiation factor 4b) do?
Locates AUG
After reaching the end codon (UGA, UAA, or UAG), what protein family remove the 40s and 60s ribosomal subunits from the mRNA strand?
Eukaryotic release factors (eRFs)
Which end of a tRNA holds the amino acid?
The 5’ end
The initial tRNA holds what amino acid?
On which end?
Methionine;
5’
In what order of binding sites does a tRNA pass through the ribosome?
A –> P –> E
What is the A site of the ribosome?
Where the aminoacyl-tRNA enters the ribosome.
What is the P site of the ribosome? What two things happen here?
Where the aminoacyl-tRNA anticodon binds the mRNA codon;
the amino acid is joined to the growing chain via peptidyl transferase
What is the E site of the ribosome?
The exit point for the empty tRNA
What proteins bind recently translated mRNA to protect them and help them fold?
Chaperone proteins
(e.g. heat-shock proteins)
Proteins can be heavily phosphated and then tagged with what marker to target them for degradation?
What is this process called?
What enzyme attaches the marker?
Ub;
ubiquination;
ub-ligase
What complex degrades ubiquinated proteins?
The 26S proteasome
What is a silent mutation?
What is a frameshift mutation?
What is a nonsense mutation?
No change in coded amino acid (often due to RNA wobble);
Insertion or deletion (change in reading frame);
premature stop codon (truncated protein)
What are the stop codons?
For what amino acid do they code?
What is the start codon?
For what amino acid does it code?
UAA, UAG, UGA,
none;
AUG,
methionine
The wobble hypothesis involves which of the three nucleotides of an RNA codon?
The third
What enzyme normally cleaves APP extracellularly?
What enzyme normally cleaves APP intracellularly?
What enzyme is pathological in its extracellular cleavage of APP?
Alpha-secretase;
gamma-secretase (PSEN1);
beta-secretase
G-proteins hydrolyze what substrate for energy?
GTP
What does the Gα subunit of a GPCR do?
Binds GDP (or GTP);
modifies the effector molecule (e.g. adenylyl cyclase)
After a ligand binds the receptor portion of a G-protein coupled receptor, what occurs?
The Gα subunit exchanges GDP for GTP,
dissociates from the GPCR,
and binds to an effector molecule
Upon ligand binding, which of the trimeric subunit of a GPCR dissociates from the GPCR and binds to an effector molecule?
What subunits are there?
Gα;
Gα, Gβ, Gγ
To what is the Gα subunit bound when a ligand binds the GPCR receptor and what occurs next?
GDP;
the GDP is exchanged for GTP;
the Gα dissociates from the GPCR
True/False.
A single effector molecule (such as adenylyl cyclase) can have both stimulatory and inhibitory GPCRs.
True.
After leaving the GPCR and binding the effector molecule, how does the GTP-bound Gα dissociate from the effector to return to the GPCR?
It hydrolyzes the GTP
Describe the pathway of a GPCR that acts on adenylyl cyclase.
Ligand binds GPCR –>
Gα (trades GDP for GTP) dissociates –>
binds adenylyl cyclase –>
creates cAMP –>
activates protein kinase A
What secondary messenger is created by adenylyl cyclase?
What enzyme does it activate?
cAMP;
protein kinase A (PKA)
Describe the pathway of a GPCR that acts on guanylyl cyclase.
Ligand binds GPCR –>
Gα (trades GDP for GTP) dissociates –>
binds guanylyl cyclase –>
creates cGMP –>
activates protein kinase G
What secondary messenger is created by guanylyl cyclase?
What enzyme does it activate?
cGMP;
protein kinase G (PKG)
Describe the pathway of a GPCR that acts on phospholipase C.
Ligand binds GPCR –>
Gα (trades GDP for GTP) dissociates –>
binds phospholipase C –>
creates inisitol triphosphate (IP3) –> releases stored Ca2+
and diacylglycerol (DAG) –> activates protein kinase C
What secondary messengers are created by phospholipase C?
What substances do they activate and release, respectively?
Diacylglycerol (DAG) –> protein kinase C (PKC);
inisitol triphosphate (IP3) –> Ca2+
What secondary messenger is elevated in cases of adenylyl cyclase activation?
cAMP
What secondary messenger is elevated in cases of guanylyl cyclase activation?
cGMP
What secondary messengers are elevated in cases of phospholipase C activation?
diacylglycerol (DAG),
inisitol triphosphate (IP3),
Ca2+
How does cAMP exert an effect on protein kinase A?
It binds the 2 regulatory subunits,
these activate the 2 catalytic subunits,
these enter the nucleus
What sorts of extracellular signals will activate GPCRs that eventually activate PKA?
What are some of the effects in adipose tissue, liver tissue, and skeletal muscle?
Norepinephrine, epinephrine, ACTH, glucagon;
increased serum fatty acids, amino acids, and glucose
(increased lipolysis, decreased glycogenolysis, increased glycogenesis, decreased amino acid uptake, increased gluconeogenesis etc.)
Will cAMP-PKA signaling have the same effect on each tissue in which it is utilized?
No
(liver responses are very different from ovarian responses which are very different from kidney responses, etc.)
Phospholipase C acts on what plasma membrane substrate to produce DAG and IP3?
PIP2
(phosphatidylinositol + 4-,5-phosphates)
(PI from the P-leaflet)
After phospholipase C acts on PIP2 to form DAG and IP3, what happens next?
IP3 triggers release of Ca2+ from the ER;
Ca2+ then helps PKC to go out towards the plasma membrane where DAG activates it
Smooth muscle relaxation is a GPCR-stimulated response that is an example of what sort of cell-to-cell signaling?
Paracrine signaling
Acetylcholine binds a GPCR on a smooth muscle cell. What effector enzyme is affected and how?
What is the effect?
Phospholipase C, activated;
increased Ca2+ binds calmodulin and activate nitric oxide synthase
(which diffuses and thus relaxes surrounding myocytes)
After acetylcholine binds a smooth muscle GPCR, phospholipase C is activated and calcium released from the sarcoplasmic reticulum.
This calcium then binds to what?
The union of calcium and this other molecule then have what effect?
This nitric oxide then does what?
Calmodulin;
activate nitric oxide synthase;
binds a GPCR for guanylyl cyclase (causing smooth muscle relaxation)
Nitric oxide binds to what receptor on smooth muscle cells?
GPCR –> guanylyl cyclase –> cGMP –> PKG
What are the three main categories of single pass receptor signaling pathways?
(C, RTK, T)
Cytokine receptors;
receptor tyrosine kinase;
TGFβ receptors
What type of single pass receptor is responsible for JAK/STAT signalling?
Cytokine receptors
Do cytokine receptors have kinases that are intrinsic (part of) or extrinsic (just nearby or associated) to the receptor itself?
Extrinsic (the JAK kinase)
What is the name of the kinase that is extrinsic and closely associated with cytokine receptors?
JAK
When a ligand binds to a cytokine receptor monomer, what happens?
Two monomers are brought together, and their extrinsic (but intracellular) kinases are united
When cytokine receptors dimerize (upon meeting a ligand) and their extrinsic JAK kinases come together, what happens?
The JAK kinases phosphorylate and activate one another;
they then phosphorylate tyrosine residues on the intracellular portion of the receptor itself
Why is it important that JAK kinases phosphorylate the intracellular portions of activated cytokine receptors?
So, the phosphorylated proteins can serve as scaffolding/cascade for signalling proteins
What is the STAT (of the JAK/STAT cytokine receptor pathway) part of JAK/STAT?
A transcription factor
(SH2 + a DNA-binding domain)
Phospholipase C activation has what effect on smooth muscle cells?
Guanylyl cyclase activation has what effect on smooth muscle cells?
Nitric oxide production and subsequent guanylyl cyclase activation;
relaxation and blood vessel dilation
When is the Gα subunit (of a GCPR) active?
When bound to GTP
Often, single pass receptors (cytokine receptors, RTKs, TGFβ receptors) exist as single ___________, but, after ligand binding, unite to become ____________.
Monomers;
dimers
(Image: example cytokine receptor)
Describe the pathway of a receptor tyrosine kinase.
- Ligand binding and receptor dimerization
- Kinase activation (of each other)
- Receptor phosphorylation; signal cascade activation
Which has an extrinsic kinase, receptor tyrosine kinases or cytokine receptors?
Cytokine receptors
(extrinsic = associated but not directly part of the receptor)
A monomeric G-protein is active if it is bound to:
A monomeric G-protein is inactive if it is bound to:
GTP;
GDP
A monomeric G-protein is active if it is bound to GTP.
What activating enzyme exchanges G-protein GDP for GTP?
A monomeric G-protein is inactive if it is bound to GDP.
What inactivating enzyme exchanges G-protein GDP for GTP?
GEF
(guanine nucleotide exchange factor);
GAP
(GTPase activating protein)
What enzyme activates monomeric G-proteins?
Through what mechanism?
GEF (guanine nucleotide exchange factor);
trading G-protein GDP for GTP
What enzyme inactivates monomeric G-proteins?
Through what mechanism?
GAP (GTPase activating protein);
promoting the hydrolysis of G-protein GTP to GDP
The following cascade of proteins describes the effects of what ligand type?
Sos (a GEF, a GDP to GTP exchanger) activates Ras;
Ras –> Ref –> MEK –> MAPK (ERK)
A receptor tyrosine kinase (RTK)
What mutation in the receptor tyrosine kinase pathway leads to overexpression of MAP kinase activity and increased likelihood of cancer development?
Ras (a monomeric G-protein) mutates and is perpetually active
What is the JAK (of the JAK/STAT cytokine receptor pathway) part of JAK/STAT?
The kinase associated with cytokine receptors
What transcription factors are activated via a typical TGF-β receptor mechanism after the receptors (I, II, and III) have united and autophosphorylated?
SMAD3, SMAD4
What are some examples of substances that interact with cytokine receptors?
Interferons,
prolactin,
EPO
Cytokine receptors and the JAK/STAT pathway are typically pro- what?
Pro-growth
(and sometimes differentiation)
Besides being pro-growth, what other effect does the JAK/STAT pathway of cytokine receptors have on cell responses to noxious stimuli?
It is anti-inflammatory
An inactivating mutation in the SMAD4 transcription factor (or receptor I or II) of the TGF-beta pathway would have what effect on cellular activity?
Increased growth and cancer risk
Via which G-protein is a signal transferred to the effector adenylyl cyclase to stimulate increased cAMP production?
Gα stimulatory (Gαs) subunit
(turn on the Gαs)
What is another term for homologous recombination?
What is the term for the anatomic point of DNA ‘swapping’ between chromosomes?
True/False.
This involves the RecBCD complex and RecA enzyme.
Crossing over;
chiasma;
true
What is here described: reciprocal chiasmatic exchange of genetic information between homologous chromosomes during gametogenesis.
Homologous recombination
True/False.
The further apart two loci are, the more likely it is that recombination will occur.
True.
Homologous recombination can be used to ‘skip’ past what type of DNA error?
One that stalls replication
(e. g. a thymidine dimer)
* (Note: in lefthand portion of image, pay attention to the location of the lesion in each step)*
How can homologous recombination help DNA replication to ‘skip’ past large DNA lesions such as thymidine dimers?
By regressing and using the newly synthesized strand opposite it as a template
(Note: in lefthand portion of image, pay attention to the location of the lesion in each step)
Homologous recombination can be used to repair small DNA nicks by uniting what two strands?
The leading and lagging strands
(Note: righthand portion of image)
The Sos protein is activated by receptor tyrosine kinases to do what to what what G-protein?
Activate (Sos is a GEF);
Ras
Ras is a __________ and is part of what signalling pathway?
JAK is a __________ and is part of what signalling pathway?
Monomeric G-protein, receptor tyrosine kinase;
kinase, cytokine receptor
How are retrotransposons ‘copy and pasted’ throughout the genome?
RNA polymerase makes an RNA template;
reverse transcriptase then turns it back into DNA
(much like a retrovirus)
Via what two category of enzymes are DNA transposons ‘cut and pasted’ throughout the genome?
Endonucleases and ligases
TGFβ signalling causes modified gene transcription with what two main effects regarding the extracellular matrix?
- Increased ECM secretion
- Secretion of plasmogen activator inhibitor 1 (PAI1)
(basically, more extracellular protein fibers and less degradation of the fibers –> more stable tissues)
What are the three segments of antibody light-chain DNA sequences?
V segments (many which can be swapped in and out);
J segments (a few, out of which one type is usually selected);
C (the constant portion)
What sequences and enzymes are responsible for this process of antibody light-chain transposon recombination?
Recombination signal sequences (RSSs);
RAG1, RAG2
TGFβ signalling causes modified gene transcription with what main effect regarding cellular growth?
TGFβ signalling is anti-growth
(9CDKI p15 and PAI1 expression)
Inactivating mutations in the TGFβ signalling pathway would have what effect?
Excess growth and increased cancer risk
(e.g. SMAD or receptor I or II mutations in RB and cancers of the colon, pancreas, stomach)
What is an example of a serum protein with the opposite effect of tissue plasminogen activator (TPA)?
Plasminogen activator inhibitor 1 (PAI1)
Oncogenic mutations involving SMAD4 are related to what single pass transmembrane receptor pathways?
Oncogenic mutations involving Ras are related to what single pass transmembrane receptor pathways?
Mutations involving JAK are related to what single pass transmembrane receptor pathways?
TGFβ receptors;
receptor tyrosine kinases;
cytokine receptors
What is the purpose of DNA recombination?
Mixing and rearranging the genome
(creates more genetic diversity and variability among humans)
When does homologous recombination occur?
What are its purposes?
Gametogenesis,
somatic replication;
increase genetic diversity, DNA repair
True/False.
Homologous recombination sometimes leads to tumor formation.
True.
When during the cell cycle does homologous recombination occur in a gamete?
Prophase I
Homologous recombination can be used to map what on a chromosome?
Genetic loci
What is the term for chromosomes where no crossing over has occurred?
What is the term for chromosomes where crossing over has occurred?
Parental;
recombinant
Homologous recombination (crossing over) can have what effect on an individual heterozygous for a particular allele?
Is this true for meiosis or mitosis?
Loss of heterozygosity
(the alleles replicate for mitosis [2 –> 4], and then the gene is swapped so each daughter cell is homozygous for one of the original two alleles);
mitosis
True/False.
Mitotic (or meiotic) nondisjunction is a form of homologous recombination.
False.
What category of transposons are ‘copy-and-pasted’ around the genome?
What category of transposons are ‘cut-and-pasted’ around the genome?
Retrotransposons;
DNA transposons
Are many DNA transposons inactive?
Yes;
nearly all
Excision of various transposons is essential to the diverse variability of what type of cellular immune product?
Antibodies
What is ‘exon shuffling?’
Switching of exons among various genes
(transposition recombination)
How can the frequency with which two genes are swapped in homologous recombination be useful to mapping out genetic loci on a chromosome?
The farther apart two genes are, the more likely it is that there will be some crossing over between them
(the frequency is proportional to the distance between the loci)
Can multiple eukaryotic ribosomes be actively translating a single mRNA strand simultaneously?
Yes
(polyribosomal activity)
What post-translational protein modifier converts proline from the trans configuration to the cis configuration?
What post-translational protein modifier creates disulfide bonds between cysteine residues?
Note: These enzymes, along with chaperones and HSPs, are both important to proper protein folding.
Peptidyl prolyl cis-trans isomerase;
protein disulfide isomerase
What is added to a substrate during carboxylation?
A carboxyl (COO-) group
What is the term for an inactive enzyme that is lacking its specific prosthetic group?
What is the term for an active enzyme that is accompanied by its specific prosthetic group?
Apoprotein, apozyme;
holoenzyme
Which of the following signal types is(are) generally pro-growth?
Which of the following signal types is(are) generally anti-growth?
Cytokine receptors
Receptor tyrosine kinases
TGFβ receptors
Cytokine receptors,
Receptor tyrosine kinases;
TGFβ receptors
