Download Gene Expression in Bacteria

Gene Expression
• Second way to carry out therapeutic cloning is
to use adult stem cells.
• Found in many organs
– Skin, bone marrow, umbilical cord
• Goal is to turn any adult stem cell into any
type of specialized cell
Gene Expression in Bacteria
• Simpler than in Eukaryotes
• E. coli
– Lives in intestines and can quickly adjust its
enzymes according to what you eat.
– If you drink milk, immediately begins making 3
enzymes needed to metabolize lactose
Gene Expression in Bacteria
• Operon: A cluster of structural and regulatory
genes that acts as a unit.
– Promoter: A sequence of DNA where transcription
begins
– Operator: A sequence of DNA where a repressor
binds
Lac Operon in E. coli
• When lactose is absent:
– The regulatory gene codes for a repressor that is
normally active
– A repressor protein binds to the operator
– RNA polymerase cannot transcribe the three
structural genes of the operon (the structural
genes are not expressed)
Lac Operon in E. coli
– RNA polymerase cannot transcribe the three
structural genes of the operon (the structural
genes are not expressed)
Lac Operon in E. coli
• When lactose is present:
– Lactose binds with the lac repressor
– Repressor is unable to bind to the operator
– Structural genes are transcribed
• Enzymes are produced
– Binding of RNA polymerase to the promoter is
further ensured by another gene regulatory
protein, called CAP.
• https://www.youtube.com/watch?v=h_1QLdt
F8d0
Lac Operon in E. coli
• Enzymes are produced
– Binding of RNA polymerase to the promoter is
further ensured by another gene regulatory
protein, called CAP.
Gene Expression
• Every cell in the body gets a copy of all the
genes, therefore every cell in the body has the
potential to become a complete organism
• And yet the human body contains many types
of cells that differ in structure and function.
• Each cell type contains its own mix of different
proteins that make it different from all other
cell types.
• Only certain genes are active in different cells.
Gene Expression in Eukaryotes
• Chromatin: complex of DNA and proteins that
forms chromosomes
• Nucleosome: two turns of DNA around a set
of eight proteins called histones
Gene Expression in Eukaryotes
• Eukaryotic cells exhibit control of gene
expression at various levels
• 1. Chromatin packing keeps genes turned off.
– Some inactive genes are located within darkly
staining portions of chromatin, called
heterochromatin.
Heterochromatin
• Barr body in mammalian females
– Females have a small, darkly staining mass of
condensed chromatin adhering to the inner edge
on the nuclear membrane.
– This structure is an inactive X chromosome
Heterochromatin
• X-inactivation in cats (females)
– Alleles for black and orange carried on X
chromosomes.
– Random X-inactivation occurs
Heterochromatin
– In heterozygous females, 50% of the cells have an
allele for black coat colour and 50% have allele for
orange coat.
– Result is a tortoiseshell or calico cat with patches
of orange and black
Heterochromatin
• Can occur in humans
– Ex. Women who are heterozygous for hereditary
absence of sweat glands have patches of skin
lacking sweat glands.
Euchromatin
• Euchromatin: loosely packed areas of active
genes
• When DNA is transcribed, a chromatin
remodeling complex pushes aside the histone
portion of the nucleosome so that
transcription can begin
• Only after DNA unpacking is it possible for a
gene to be turned on and expressed.
Unpacking of DNA
Euchromatin
• Housekeeping Genes
– Required for the maintenance of basic cellular
function
– Not finely regulated
– Products are always needed to some degree
• Ex. red blood cells, muscle cells, pancreatic cells
Gene Expression in Eukaryotes
1. Transcription
• Transcription factors: proteins that help RNA
polymerase bind to a promoter
– If one is defective, can have serious effect –
Huntington Disease
Gene Expression in Eukaryotes
Transcription activators: proteins that speed up
transcription
– Bind to enhancer region on DNA
– Enhancer and promoter may be far apart
– DNA forms a loop to bring them close together
Gene Expression in Eukaryotes
2. mRNA processing (post-transcription)
– Follows transcription
– Removal of introns and splicing of exons
– One segment of mRNA can be processed in
several different ways therefore can produce
several different proteins
– The speed at which the mRNA leaves the nucleus
can affect the amount of protein produced
Gene Expression in Eukaryotes
3. Translation
• Differences in the poly-A tails may determine how
long a mRNA is available for translation
• The longer an mRNA remains in cytoplasm before
broken down, the more gene product there will be.
• Specific hormones may also effect longevity of mRNA
Gene Expression in Eukaryotes
4. Protein Activity (post-translation)
• Some proteins must be activated after
synthesis
• Many proteins function only for a short time
before they are degraded or destroyed by the
cell.
SO HOW THE HECK DOES A GENE GET
TURNED ON????
• Signaling Between Cells
– Cells are in constant
communication
– Cell produce a signaling
molecule that binds to a
receptor protein on a target
cell
SO HOW THE HECK DOES A GENE GET
TURNED ON????
• Initiates a signal
transduction pathway;
series of reactions that
change the receiving cell’s
behaviour
– May result in stimulation
of a transcription activator
– Transcription activator
will then turn on a gene
Cancer: A Failure of Genetic Control
• Cancer cells are no longer able to respond
appropriately to signals from their neighbours.
Characteristics of Cancer Cells
• Contact Inhibition: When cells come into
contact with neighbouring cells, they stop
dividing (regular cells)
• Cancer cells lose contact inhibition and form
tumors.
• Benign: noncancerous if it stays as a single
mass.
• Malignant/Cancerous: when they invade
surrounding tissues.
Characteristics of Cancer Cells
• Nonspecialized cells: do not contribute to the
functioning of a body part.
• Look distinctly abnormal
• Normal cells enter the cell cycle about 50
times and die
• Cancer cells enter cell cycle repeatedly
Characteristics of Cancer Cells
• Abnormal nuclei: enlarged, may contain
abnormal number of chromosomes
• Chromosomes have mutated; deleted or
duplicated
• Apoptosis: cells with damaged DNA undergo
programmed death (normal cells)
– Cancer cells fail to undergo apoptosis
Characteristics of Cancer Cells
• Spread to new locations: cancer cells can
invade underlying tissue and also move
through the blood or lymph.
• Metastatic tumors: found elsewhere in the
body
Proto-oncogenes and TumorSuppressor Genes
• Mutations in these types of cells account for
the characteristics of cancer cells:
– Proto-oncogenes: promote the cell cycle and
prevent apoptosis.
– Tumor-suppressor genes: prevent the cell cycle
and promote apoptosis.
Proto-Oncogenes
• Mutate into cancer causing genes called
oncogenes
• An altered Ras protein is found in approximately
25% of all tumors.
• Several proto-oncogenes code for the Ras protein
that functions in the signal transduction
pathways that turn a gene on.
• Ras oncogenes are found in lung, colon and
pancreatic cancers, (just to name a few) and
essentially keep the cell cycle turned on
permanently (big problem!).
Proto-Oncogenes
• Cyclin D is a proto-oncogene that codes for cyclin
– a protein that promotes the cell cycle. If it
becomes an oncogene, it will produce cyclin all of
the time which leads to continual cellular
division.
• A transcription activator called p53 normally
prevents cell division if there is damage to the
DNA and will repair it. If the gene that codes for
p53 mutates, the cells may continue to divide
indefinitely with errors.
Tumor-Suppressor Genes
• When these mutate, they no longer inhibit the
cell cycle.
• The retinoblastoma protein (RB) controls the
activity of transcription factor cyclin D and
other genes that regulate the synthesis phase
(where DNA is replicated during interphase).
Tumor-Suppressor Genes
• When the tumor-suppressor gene p16
mutates, the RB protein is always functional
and results in too much active cyclin D.
• The protein Bax promotes apoptosis. When a
tumor-suppressor gene Bax mutates, the
protein Bax is not present; therefore,
preventing cell death (apoptosis).
Other Genetic Changes
• Telomere: sequence of bases at the end of
chromosomes keeping them from fusing with one
another or deteriorating
• Shorten with each cell division, eventually
becoming useless.
• Cell dies by apoptosis
• Cancer cells have an enzyme
telomerase that rebuilds
telomeres allowing them to keep
dividing.
Other Genetic Changes
• Angiogenesis: formation of new blood vessels
by cancer cells
• Growth factors cause new blood vessels to
form and send capillaries into centre of tumor.
• Supply tumor with nutrients and oxygen
Other Genetic Changes
• Metastasis: formation of new tumors distant
from primary tumor.
• Cancer cells travel through blood or lymphatic
vessel
• Recovery doubtful
Causes of Cancer
• Heredity
– Particular types run in families
– Ex. Li-Fraumeni cancer family syndrome, Lynch
cancer family syndrome
• Environment
– Carcinogens: environmental agent that causes
mutations leading to the development of cancer
– Radiation, organic chemicals, viruses
Diagnosis of Cancer: Screening Tests
• Pap test: sample of cells from cervix,
examined under microscope for abnormalities
• Mammograms: reveal a breast tumor too
small to be felt
• Colonoscopy: detects polyps (clump of cells)
while they are small enough to be destroyed
by laser therapy
Diagnosis of Cancer: Screening Tests
• Tumor Marker Tests: blood tests for tumor
antibodies/antigens.
– Tumors release substances that provoke an
antibody response in the body
– For someone who has already had colon cancer,
can use presence of an antigen called CEA to
detect relapses
– When CEA level rises, additional tumor growth
occurs.
Diagnosis of Cancer: Screening Tests
• Genetic Tests: test for genetic mutations in
proto-oncogenes and tumor-suppressor genes
– Detects the likelihood that cancer in present or
will develop in near future
– Tests available for colon, bladder, breast, and
thyroid cancers and melanoma
– A Ras oncogene can be detected in stool and
urine. If test for Ras oncogene in stool is positive,
colon cancer is suspected. If Ras oncogene in
urine is positive, bladder cancer is suspected
Diagnosis of Cancer: Screening Tests
• Genetic testing can be used to determine if
cancer cells still remain after a tumor has
been removed.
• The gene that codes for telomerase is turned
off in normal cells but active in cancer cells. If
the test for the presence of telomerase is
positive, the cell is cancerous.
Confirming Diagnosis
• Biopsy: removal of cells for examination
• Laparoscopy: permit viewing of body parts
• Radioactive scan: obtained after radioactive
isotope is administered, can reveal abnormal
isotope accumulation due to a tumor.
• Ultrasound: echoes of high-frequency sound
waves directed at a part of the body are used
to reveal size, shape, and location of tumors.
Treatments of Cancer
• Surgery
– Good for removal of localized tumor
– Danger that some cancer cells left behind
– Often followed by radiation or chemotherapy
• Radiation
– Will cause cancer cells to mutate and undergo
apoptosis.
– Powerful X rays or gamma rays administered through
applied beam or by implanting tiny radioactive
sources directly into body
Treatments of Cancer
• Chemotherapy: chemotherapeutic drugs kill
cancer cells
– Used to reduce the chance of recurrence after
surgery
– Taxol (type of drug) interferes with microtubules
needed for cell division
– Bone Marrow Autotransplantation: Bone marrow
prone to destruction during chemo. Patient’s stem
cells harvested and stored before chemo begins
and returned by injection post chemo
Future Therapies
• Vaccines: stimulate immune system to find
and destroy cancer cells
• Antiangiogenic drugs: confine and reduce
tumors by breaking up network of new
capillaries
• Injection of gene for p53 protein to promote
apoptosis