1. Define the following terms and
utilize these terms in an appropriate context:
active regulator
attenuation
constitutive mutation
inducer
lac operon
negative control
operon model
positive control
prosttranscriptional regulation
promoter
repressible system repressor
structural gene
operator region
enhancer
inducible system
anabolic
catabolic
2. Describe differentiation and relate
this to the concept that all cells possess all genes
at all times yet are
not actively using them.
3. Appreciate that a system of genetic
regulation must exist if the complete genome
is not to be continuously
active in transcription.
4. Differentiate between inducible
and repressible operons, illustrated by the lac and
trp gene complexes,
respectively.
5. Contrast the need for the enzymes
involved in the metabolism of lactose and tryptophan
in bacteria in the
presence and absence of lactose and tryptophan, respectively.
6. Contrast positive and negative control systems.
7. Compare the normal activity and
control mechanisms of genes for anabolic (synthesis)
enzymes with those
for catabolic (respiration) enzymes.
8. Contrast the role of the represor in an inducible system and in a repressible system.
9. Explain the occurrance of an additional
regulatory mechanism, called attenuation,
illustrated by the
trp operon.
10. Outline the genetic control mechanisms of the lac and trp operons.
11. Outline differences between genetic regulation in prokaryotes vs. eukaryotes.
12. Distinguish between the regulatory elements referred to as promoters and enhancers.
13. List and define various levels
of regulation in eukaryotes including posttranscriptional
control and translational
control mechanisms.
14. Describe the impacts on regulatory
mechanisms of eukaryotes when dealing with
the properties of multicellularity,
expanded genome size, and the spatial and temporal
separation between
transcription and translation.
15. Describe the nature of chromosome
puffs and lampbrush chromosomes, and relate
them to gene activity.
Resources: Text Chapter 15, Cartoon
Guide pgs. 164-178
Regulation of Gene Activity- Prokaryotes & Eukaryotes
Basic assumption: -when
dealing with regulation, not all cells need all enzymes at all times
-all
cells contain some genetic information but not all cells need and use all
the info
(cellular differentiation)
-single
celled prokaryotes need less control than multicellular eukaryotes
Experiments in differentiation
Totipotency- "all powerful", ultimate control. Can any cell with full genetic
information
produce a new individual?
Are same genes present
in all cells?
Differentiation ---> past assumption: only genes specific for digestive
functions will be
found in cells of digestive system (no other genes needed, therefore, no
other genes
present)
Different cells get different genes.
1. Hypothesis #1- same genes present in all cells
-Totipotency experiments
Frog egg --> nucleus destroyed ---> nucleus of intest. epithelial cell
injected into
cell ---> tadpole ---> mature frog (sometimes)
**mechanism of control still unclear - don't know how to turn on all inactive genes.
Conclusion: epithelial nucleus contained info to produce mature frog.
-Adult plant tissue
1. dissociated into single cells
2. cultured in nutrient media
3. produce cell mass (callus)
4. add hormones (growth) adult can form.
**differentiation due to regulation not to genes being seperated according to job.
2. Hypothesis #2- differentiation due to different genes being active (regulated)
in different cells.
-mRNAs will only be produced by "active" genes
-Hybridizing experiments- create DNA/RNA hybrid to determine activity
A. Mouse liver cells
1) DNA denatured and cut (all DNA, 100%)
2) Add cut DNA to Nuclear mRNA (mRNA would indicate activity of gene,
tells how many genes are being transcribed)
3) 4.5% of DNA hybridize, therefore, 4.5% of genes, (DNA) are active in
mouse liver cells. Other 95.5% inactive
B. Mouse kidney cells
1) DNA denatured and cut.
2) add cut DNA to nuclear mRNA
3) 4.0% of DNA hybridize, therefore, 4.0% of DNA active 96% inactive.
C. Are these the same genes?
If no then: DNA + Kidney mRNA + Liver mRNA should equal 8.5%
If yes then: DNA + Kidney mRNA + Liver mRNA should equal 4.5%
*actual findings 7.5%: indicates there may be some genes in
common
but most are not
*supports hypothesis that all DNA present but some are "turned on"
others "turned off"
Regulation in Prokaryotes
Promoter- AUG (met) start, short
sequence of DNA where RNA polymerasebegins
transcription
repressor- prevents RNA polymerase from attaching to a promoter
operator- where repressor binds
preventing RNA polymerase from attaching to promoter
(called on/off switch of transcription
regulator- gene that codes for repressor, seperate from gene it regulates
structural gene- one to several
genes that are transcribed as a unit, determine structure
of proteins.
Operon Model
A. Lac Operon
Jacob and Monod 1961
-experiments show E. coli capable of regulating genes necessary for lactose
metabolism
*genes for proteins that function together should be controlled together
*metabolic pathways require numerous enzymes (proteins) in roughly equal
amounts
*one mechanism that allows coordination of a group of functionally regulated
genes
would be to group them so they can be regulated together
- E. coli ordinarily uses glucose, because E. coli uses glucose, lac operon
usually
turned off! (therefore, operator repressed)
promoter
lac z lac y
lac a
DNA----------------------------------------------------------------
r
o+
repressor- acts on operator gene to turn operator off, prevents RNA poly from working
lac z- codes for B galactosidase (breaks down lactose)
lac y- codes for enzyme that takes lactose into cell
lac a- utilization of lactose function
still unclear
When lactose present:
lac z lac y
lac a
DNA-----------------------------------------------------------------
r
o+
repressor + lactose (produces inactive repressor)
Operon induced ans allowed to function
(RNA polymerase can read message)
Negative control of lac Operon
-operon turned on unless
something intervenes to stop it (turn it off)
*when lactose present operon is on, repressor can't repress
Positive control- operon turned off until something intervenes to turn it on.
Negative control- brakes of car
Positive control- ignition switch
*removing repressor from operator (releasing brake) is not enough to turn
of the
operon. Positive factor, turning on ignition, is also needed.
*mutant regulator- cannot produce functional repressor therefore, pathway can't stop
oc- constitutive operator- repressor can't attach therefore, can't turn system off
oo- mutant regulator- combines irreversibly w/repressor, therefore, genes
can't be
turned on
Genes for enzymes of synthetic or anabolic pathways are controlled by repression
(negative control). They are turned on all the time except when specifically
repressed
Tryp operon- repressed by presence of tryp
Genes for enzymes of catabolic (respiration and energy release) pathways
are
inducible. They are normally turned off except when specifically
induced to turn on
(positive control)
lac Operon- induced in presence of lactose
Regulation in eukaryotes Pg. 314-321
1. Transcriptional control- involves organization of chromatin and use of regulatory genes.
A. Organization of chromatin
Heterochromatin- highly condensed (before mitosis)
Euchromatin- diffuse (interphase)
-histones are basic proteins, synthesized in nucleolus, become complexed
w/DNA to form coiled chromatin.
-histone complexed DNA cannot transcribe to mRNA, therefore, it is genetically
inactive
Evidence
Barr Bodies- one of the X chromosomes in females is condensed and inactive.
Polytene Chromosomes- genetically inactive (euchromatin)
Chromosome puffs- gene regions, actively synthesized mRNA, hormone induced
*prokaryotes- no histone
2. Post-transcriptional
Control
-differential mRNA processing, effects size of mRNA, therefore, time it
takes to leave
nucleus and in effect, the time it takes to make a protein.
3. Translational
control- dependent on the life expectancy of mRNA (can vary)
-causes stock pile of mRNA, when time is right there will be a large burst
of
translation and protein production.
4. Post-translational Control- effects activity of protein produced
-occasionally other alternatives are necessary
-feed back mechanisms
Cancer- Failure of Genetic Control
*exhibit uncontrolled and disorganized growth
Tumor- growth of cells that invades and destroys neighboring tissues
- fail to differentiate into organ cells and never help function or organ (heart, lung . . .)
Vascularization- growth
of blood vessels into cancerous tissues supplying nutrients
and O2
*cancer cells break away from tumor and spread throughout body
Metastasis- tumor cells begin growth in other parts of body
Benign Tumor- prior to metastasis (non cancerous), basis for early
detection
programs
Malignant tumor- tumors that have metastasized
Causes- Mutagens (mutagens/carcinogens)
Carcinogens- mutagenic agents which cause changes in DNA
1st step in development ---> Initiation
Radiation- breaks, disruption of DNA bonding patterns
Chemicals- base sequence changes
2nd step in development ---> Promotion
any influence that triggers cell to divide uncontrollably(either a second
change in DNA or cumulative
effects of DNA changes)
*Oncogenes (onco= cancer)
-mutated versions of normal (proto-oncogenes) genes already incorporated into DNA
-mutated version turns on gene normally turned off
-mutation leads to greater
expression of the gene or to inappropriate expression of gene
-proto-oncogene induced
structurally by crossing over placing inactive structural gene next to
active promoter
-proto-oncogene induced by the introduction of its oncogene by a virus
-Cancer develops if immune system fails to attack the cancerous cell.
**active area of research
-may be genetic basis for some cancers which would predispose individuals to develop cancer
Tp53 genes- p53 proteins: Tumor supressor genes
The "anti" oncogene
Father- smoker w/ lung cancer Smoker- cancer
-offspring
may inherit gene
Mother- non-smoker
Healthy- no cancer