DIFFERENTIATION
Differentiation
The process by which cells or parts of an
organism become different from one and
other and also from their previous state.
The process by which cells or tissues of an
organism acquire the ability to perform their
special functions.
How is this accomplished?
Selective gene transcription
In any given cell, only the genes necessary for
basic metabolism and that cell’s special functions
are active.
muscle cells - actin, myosin
pancreatic acinar cells - digestive enzymes
neurons - tubulin, neurotransmitters
Mechanisms of selective gene transcription
Ascaris suum, Parascalas univalens - nematode (roundworm) parasites
found in pig intestine.
http://cal.vet.upenn.edu/paraav/labs/lab_4.htm
*Chromosome diminution - loss of pieces of chromosomes during
mitosis
*In Ascaris, pieces of chromosomes are lost in certain cells during th
2nd - 5th cleavage divisions (the figure above which shows
diminution during the 2nd cleavage is from the original work of
Boveri (1925) and actually is for another roundworm, Parascalas
univalens).
*About 25% of the DNA is lost. This is mostly repetitive (non-gene
sequence) DNA; however, a few functional genes are lost.
*At the 32 cell stage only one cell remains with a complete genome.
*This cell will give rise to the germ cell line. The other 31 cells give
rise to the somatic tissues.
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Mechanisms of selective gene transcription
Post-transcriptional control - mechanisms that act after the
transcription event. e.g.,
Mechanisms that block the activity of specific mRNAs or
proteins until a later time when they become active (e.g.
maternally derived cytoplasmic information - formation of the
primordial germ cells in amphibians and insects).
Ultimate vs. Proximal controls of differentiation
DNA transcription is the ultimate controlling factor in
differentiation.
However, the DNA is not always active in
transcription at the time of the differentiation event.
Proximal controls are the factors that are active at
the time of the differentiation event. These could be
genes or stored cytoplasmic mRNA that is activated
at the time of differentiation.
Acetabularia
Acetabularia crenulata
Acetabularia mediterrania
http://www.biologe.de/Nuetzliches/botanik_online/d44/acetabul.htm
http://dragonja.nib.si/RtMadona/Slike_3.html
http://imbi.uwc.ac.za/courses/ifaa/jpg/Berger/Berger_02.html
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Differentiation - some genes are turned off
while others are allowed to function.
Common dogma - most cells in the body
contain a complete genome despite the fact
that most of the genes in the genome are not
necessary for each cells specific function.
Is differentiation reversible?
Is there some point in a cells differentiation
where unnecessary genes become
irreversibly inactive ( are irreversibly turned
off)?
Not too many years ago, the answer for
mammalian cells would have been YES however, things have changed.
Spemann, 1928
Gurdon, 1966
Somatic cell nuclear transfer.
Illmensee and Hoppe, 1981
Cell growth and division cycle
Three cloning methods:
1. Embryo splitting
1. Separate cells during early cleavage (2 cell - 4 cell)
2. Culture separate cells to blastocyst stage
3. Introduce cells in to uterus of hormonally prepared female
4. If successful, pregnancy and eventually birth
2. The Roslin Technique - used to create Dolly (Wilmut, 1997).
1. Collect oocyte from female
2. Enucleate oocyte
3. Induce adult cells in another culture to enter Go state
4. Fuse Go cell with enucleated oocyte with electrical shock
Introduces adult cell nucleus into oocyte cytoplasm
Electrical shock also activates egg -> Zygote
5. Cultures embyros to blastocyst stage
6. Introduce cells into uterus of hormonally prepared female
7. If successful, pregnancy and eventually birth (1 successful clone in
277 tries to get Dolly)
The Roslin Technique
Wilmut et al., 1997
Cell induced to
enter G o phase.
http://science.howstuffworks.com/cloning2.htm
Hello Dolly - Dolly the sheep, the
world's first clone from the nucleus of an
adult mammal.
http://www.guardian.co.uk/pictures/image/0,8543,-10904328739,00.html
Three cloning methods:
3. The Honolulu technique - Developed at University
of Hawaii. This method was developed for cloning
mice.
1. Use cells of the individual to be cloned that are
naturally in the Go state (e.g. Sertoli cells, Neurons,
Cumulus cells) to provide the nucleus. (somatic cell
nuclear transfer)
2. Transfer the Go nucleus to an enucleated egg by
injecting it into the egg.
3. Activate the egg using appropriate chemicals.
4. Culture to blastocyst stage and introduce
into uterus of hormonally prepared female
5. If successful, pregnancy and eventually
birth
(3 successful clones for every 100 tries)
Mooving on - A pair of new-born cloned calves in a cowshed
in Ishikawa Japan, on July 5 1998. Second adult mammal clones.
Roslin Technique
http://www.guardian.co.uk/pictures/image/0,8543,-10904328739,00.html
Idaho Gem The world's first cloned mule. The first clone to be
born in the equine family.
http://www.guardian.co.uk/pictures/image/0,8543,-10904328739,00.html
Copycat The world's first cloned kitten, named Cc. It was created by
scientists in Texas using a cell taken from an adult tortoiseshell female.
http://www.guardian.co.uk/pictures/image/0,8543,-10904328739,00.html
South Korea (Dr. Hwang Woo-Suk) unveils first dog
clone
Snuppy, whose name stands for Seoul National
University puppy.
Cloned using a cell taken from the ear of a threeyear-old male Afghan hound.
Scientists took nucleus from the ear cell and placed
it into an enucleated egg cell. Honolulu Technique.
This egg was then stimulated to start dividing and
develop into an embryo.
Once growing, it was transferred to Snuppy's
surrogate mother, a yellow labrador. The Afghan
pup was born by caesarean section after a full 60
days of pregnancy.
Scandal followed - made false claims about
producing human stem cell lines from cloned human
embryos.
http://news.bbc.co.uk/2/hi/science/nature/4742453.stm
Cloned goat Yangyang turns
four
Has given birth to twins and is now a great
grandmother.
Five little piggies Five cloned female piglets, named Noel,
Angel, Star, Joy and Mary.
Scottish-based firm PPL Therapeutics says is a major step towards
successfully producing animal organs and cells for use in human
transplants.
The pigs lack a gene to which the human immune system reacts
aggressively.
http://www.guardian.co.uk/pictures/image/0,8543,-10904328739,00.html
ANDi (inserted DNA spelled backwards)
The first rhesus monkey cloned by embryo splitting.
In 2007, Shoukhrat Mitalipov at the Oregon
Primate Research Center produced stem cells
from cloned monkey embryos using the
Honolulu technique.
http://www.guardian.co.uk/pictures/image/0,8543,-10904328739,00.html