CELL CYCLE

I. Reproduction is a unique feature of life and cellular reproduction is the basis of that process.

A. For a cell to reproduce it must go through certain steps:

1. There must be a cue that tells the cell to divide. We know this is not simply a lapse of time but involves internal and/or external signals.

2. All cellular components must increase to accommodate two cells instead of one. This includes the fact that the DNA must replicate.

3. That DNA must be divided into two new regions (segregation). The cellular components must also be segregated.

4. Cytokinesis: The cell membrane (and cell wall, if existent) must reorganize (may involve growth) to create two new cells.

B. Eukaryotic cell cycle is different than prokaryotic.

1. Prokaryotic DNA replication starts at the åoriπ site at the membrane attachment and proceeds around the circular DNA forming two interlocking bands that must separate (more on this later). Cytokinesis occurs by fission.

2. Eukaryotic DNA is much larger and occurs in linear strands (chromosomes) with lots of associated protein . It replicates at several simultaneous sites (more on this later). To segregate into two regions, the DNA is so large that it must first ≥condense≤ during a process known as mitosis.

C. Medical importance of understanding the eukaryotic cell cycle:

1. What causes cancer? How can we prevent and cure cancer?

2. How can we make wounds heal faster?

3. Can we induce damaged body parts to regenerate?

4. Can we induce undifferentiated cells to divide into organ specific cells? (e.g. stem cell research).

II. The eukaryotic cell cycle involves the events of completely reproducing a eukaryotic cell.

A. Interphase is all of the cycle except mitosis. The chromosomes are disperse in the cell during interphase.

B. Mitosis is the condensation and separation of the DNA. It is associated with four events: prophase, metaphase, anaphase, and telophase.

C. Cell division involves the separation of the chromosomes (mitosis) plus the actual dividing of the cells (cytokinesis). Cytokinesis actually usually overlaps with mitosis.

III. More on interphase:

A. G1 (gap1)

1. If the cell is designed to do something other than divide, this is the time it would usually do such functions.

2. This is the most variable of all of the stages in terms of time. It can last from minutes to years depending upon the cell type and the conditions.

3. Some cells, such as nerve cells and muscle cells never leave G1.

4. Cells that stay in G1 are said to stay at the R point (restriction).

5. Why is it important to know what triggers a cell to leave G1 and enter DNA replication (S)? Most cells have strong control over this step so that they divide when they are supposed to, and do not divide when they are not.

a) Cancer is a situation where this control is lost and cell division occurs continually. The cell does not stop in G1 when it should.

b) Organ regeneration is not uncommon in a lot of organisms where, when an organ is lost, cells again start dividing to make a new organ. If we could control the G1 to S phase, we may be able to regenerate new arms or surgically removed livers.

c) Nerves and muscles normally can not be replicated in vivo or in situ (because they are stuck in G1). It would be valuable to induce new nerve and muscle cells to grow. Some recent progress has been made in this area.

d) It is simply an important part of the living process that we are curious to understand.

6. What triggers a cell to leave G1 and enter DNA replication (S)? Cell size? Cyclic nucleotides? pH? histones doing something funny? All of these things have been implicated, but the exact answer is yet unknown. However we have learned some detail about some of the middle parts of the biochemistry of this process:

a) Cyclin dependent kinase (CdK) must bind with cyclin (a protein) for this enzyme to be activated. The activation of this enzyme is critical in moving a cell form G1 to S.

(1) ( a kinase is an enzyme that removes a phosphate from ATP and adds it to another molecule. Kinase reactions are very often controlling reactions that turn things on and off).

X + ATP -> XP +ADP

(2) Some cancers have been found to have problems with CdK or cyclins.

(3) There are actually many forms of Cdkπs and cyclins that each play their specific roles.

b) p53 is a protein known to inhibit Cdkπs and thus preventing (controlling) cell division. Over half of all human cancers have defective p53.

c) Growth factors (e.g. interlukins) can stimulate cell growth.

d) Many cells experience contact inhibition, the situation where the cell stops dividing when it comes in contact with other cells. Cancer cells usually lose their contact inhibition ability.

B. S (synthesis) DNA duplicates.

1. DNA replication (detail on this later).

2. The histones will also be duplicated during the S phase.

3. Centrioles, if present, duplicate (sort of).

C. G2 (gap2).

1. Once S occurs, then G2 and mitosis seem to be obligated to occur.

2. During this stage there are no obvious changes (microscopically) happening in the nucleus.

3. But, obviously, there must be biochemical changes going on in preparation for mitosis.

4. Duplication of the centrioles is usually completed.

IV. Mitosis - the segregation of the duplicated DNA molecules into two new nuclei. Although the process is quite continuous, it is often broken down into stages. Again note that we are talking only about eukaryotes. Prokaryotes do not do this DNA condensation. The time required for mitosis ranges from 10 min. to 3 hours.

A. Prophase

1. During this phase each long DNA molecule becomes highly infolded into itself in a highly defined way; a process called condensation. This apparently is important in untangling and separating the subsequent chromosomes.

a) They become so bunched that we can visually see each individual DNA molecule in a light microscope!

b) In interphase, eukaryotic DNA is already wrapped around histones forming nucleosomes. During prophase, this nucleosome DNA will further fold upon itself; presumably the histones are intimately involved in this process of condensation.

c) Such a condensed strand of DNA is known as a chromosome. Since a chromosome at this stage is made up of two replicated DNA molecules stuck together, then each DNA molecule is referred to as a chromatid. Chromatin is the combination of DNA and protein. The definitions are not well established, and the terminology is used in a variety of sloppy ways.

d) Each chromosome folds in such a unique and distinct fashion that each one can be identified by its size, shape and bumps! Humans have 23 pairs of chromosomes.

2. Nucleolus disappears.

3. Each of the duplicated centrioles, if present, move to each of the two aster poles. Microtubules develop near the centrioles in a star like shape called asters.

a) It is now thought that the asters are microtubules that develop simply to move the new centrioles into the two new cells that will be developing.

b) It is now thought that the centrioles have nothing to do with making the asters or the spindle (The removal of the centrioles does not prevent the development of this, nor does it prevent proper cell division.). It is believed that the centrioles are merely there to make basal bodies for the formation of cilia or flagella.

4. Microtubules may develop all of the way around the nucleus even before the nuclear membrane disintegrates.

5. The nuclear membrane 'disappears'. This marks the end of prophase and the beginning of metaphase.

a) It is thought that the nuclear membrane is actually just endoplasmic reticulum, and at this stage it blends back into the cell with the regular endoplasmic reticulum.

B. Metaphase is marked by the 'disappearance' of the nuclear membrane.

1. The nuclear membrane disappears.

2. Microtubules develop from the poles (centrosomes) and the microtubules (spindle) attaches to the chromosomes at the centromere to a protein material called the kinetichore.

3. An organized set of microtubules moves in toward the chromosomes. This organized set of microtubules is called the spindle or spindle apparatus.

4. The chromosomes line up in the center, marking the end of metaphase. They are perpendicular to the flow of microtubules.

C. Anaphase.

1. The microtubules start to pull the two chromatids apart.

2. Some microtubules go from the centrosome pole to the chromatid. These shorten.

3. Other microtubules go from pole to pole. These lengthen.

4. Together these two actions separate the chromatids toward their respective poles.

5. Probably works by sliding filaments. But it seems assembly-disassembly must also be involved.

6. Although usually quite accurate, mistakes can be made in this separation process.

D. Telophase.

1. Chromosomes, now at their respective poles, decondense.

2. The spindle starts to disappear and then completely disappears.

3. The nuclear membrane reappears, probably from endoplasmic reticulum.

Cytokinesis

I. Animal cell cytokinesis

A. Furrowing splits the two cells. - microfilaments.

B. All organelles are roughly divided in two. A nucleus goes to each cell.

C. The furrow position depended upon the midpoint of the spindle.

II. Plant cell cytokinesis

A. Prophase-telophase are basically the same.

B. The Golgi lays down vesicles filled pectin, a protein. Pectin is fairly flexible.

1. The cell way is initiated in this way.

2. Vesicles fuse, thus effecting a separation of the cells.

3. Plasmodesmata remain, thus there remains a cytoplasmic connection between the two daughter cells, and in a way the cells never actually divided.

III. Prokaryotic cell cycle

A. DNA replicates.

1. There are no histones.

2. The DNA is circular (it is a closed loop).

B. The two daughter strands are attached to the membrane at two closely spaced points.

C. Membrane grows between attachment points to separate the 2 DNA molecules.

D. A furrow is formed by inward growth of the plasma membrane and the cell wall. (Presumably microfilaments, as such, are not involved.)

GAMETEOGENISIS

I. There are three possible themes of a life cycle for an organism depending upon the dominance of the haploid versus diploid stage:

A. The organism spends its dominant time as a 2N organism.

1. When it undergoes meiosis, it is a small percentage of cells that do so.

2. Meiosis directly leads to the formation of gametes (these cells differentiate [specialize] into sperm and ova).

3. When sperm and ova fuse (fertilization) from two different organisms of the same species, a new diploid organism is formed (zygote).

4. By mitosis, the 2N zygote develops into the dominant form of the organism.

5. Most animals and plants follow this pattern.

B. The organism has a dominant 2N stage (sporophyte), followed by a dominant 1N stage (gametophyte).

1. Certain cells of the sporophyte (2N) organism undergo meiosis to form haploid (1N) cells.

2. These haploid cells, at this stage, can not undergo fertilization with other haploid cells. These asexual haploid cells are called spores.

3. The spores can undergo mitosis to form more haploid cells. In fact they can form an entire organism! This haploid organism would be said to be in its gametophyte stage.

4. Eventually, certain of the haploid cells of the gametophyte will differentiate and become gametes (ova and sperm).

5. When sperm and ova fuse from two different organisms of the same species, a new diploid organism is formed. Thus, this again initiates the sporophyte stage.

6. A good example of an organism that does this is the fern.

C. The organism has a dominant 1N stage followed by a brief 2N stage.

1. The diploid cells formed after fertilization, quickly undergo meiosis to again form haploid cells.

2. These haploid cells, at this stage, can not undergo fertilization with other haploid cells. These asexual haploid cells are called spores.

3. The spores can undergo mitosis to form more haploid cells. In fact they can form an entire multicellular organism.

4. Eventually, certain of the haploid cells of this organism will differentiate and become gametes, but they do not usually look different (thus, instead of saying they have formed sperm and ova, we say they are gametes in the form of + and -).

5. An example of such an organism would be the mold Neurospora.

II. Spermatogenisis in animals:(9-10 weeks in humans)

A. Meiosis produces four cells, called spermatids. At this stage, the spermatids have little visual difference from any other ordinary cell.

B. Then the cells undergo dramatic differentiation to form the final sperm structure.

1. A long flagellum is formed from a centriole.

2. Mitochondria move to the base of where the flagellum is forming.

3. The Golgi forms a lysosome type structure full of lytic enzymes. This structure (called the acrosome) moves to the tip of the sperm cell.

4. The nucleus greatly reduces in size and takes an elongated shape. The nucleus becomes very highly concentrated in DNA.

5. Most of the cytoplasm and its organelles disappear until there is only a thin layer of cytoplasm between the nucleus and the cell membrane.

6. The goal of this cell seems to be a design for streamlining for locomotion, yet still carry the full compliment of genetic information.

7. During fertilization, only the nucleus of the cell enters.

a) Thus mitochondria do not enter. All human mitochondria originate from the egg. Thus, all mitochondria DNA comes from the mother. Any changes seen in mitochondrial DNA are strictly due to mutation, since sexual variation is not possible. Thus, mitochondrial DNA is an excellent tool for the study of human evolution.

III. Oogenisis in animals (egg or ova cell generation):

A. During meiosis, the cytokinesis process is unequal such that only one of the four resulting cells ends up with most of the cellular cytoplasm. Only this cell (oocyte) will be fertilizable. The other three small cells (polar bodies) will eventually disintegrate.

B. Then the oocyte undergoes dramatic differentiation to form the final ova cell structure.

1. The nucleus moves back toward the center of the cell.

2. The cell becomes very active. Many ribosomes develop. Proteins, lipids and polysaccharides are made in large quantities, most of the protein and lipid being stored in the membrane bound structures called yolk bodies.

3. Many new organelles are made to accommodate the large increase in size of the cell.

4. Other surrounding cells may contribute to the efforts of 1. and 2!

5. The cell becomes big.

6. An outer coat is made (by the oocyte and/or surrounding cells). Useful for protection and helps in eventual acceptance of the sperm.

7. The goal of the egg cell, besides providing its complement of DNA, seems to be to build up a large resource of food, organelles etc. for the eventual zygote and embryo.

CELLULAR SEXUAL REPRODUCTION

I. What is cellular sexual reproduction?: The process whereby the genes (DNA, chromosomes) from two different members of the same species mix to yield an offspring that is not identical to the parent.

A. Thus, the offspring have at least two sets of chromosomes (diploid); one set from each parent.

B. Every organismic feature coded for on one chromosome will also have a corresponding location on a second chromosome. Two chromosomes that match in this way are called homologous. Although, both locations are involved in the same characteristic, they do not have to have the exact same code. The actual form that the characteristic takes is called an allele. For example, consider chromosome A and chromosome B as homologous chromosomes:

Map position	Allele of Chromosome 12f	Allele of Chromosome 12m
10	yellow eyes	flamingo eyes
22	fat toes	fat toes
31	ATP synthase	ATP synthase
41	green ears	white ears

C. Sexually produced offspring must have at least two sets of chromosomes, but may in fact have more (triploid, tetraploid, polyploid). Plants are particularly good at ploidy greater than 2. For the purposes of discussion, we will only consider the diploid situation.

D. If the offspring has two sets of chromosomes, then the two cells that joined must have only had one set each (i.e. they were haploid).

1. The cells with half of the regular chromosomes that join in a sexual union are called gametes. Thus to make a diploid offspring the gametes must be haploid (one set of chromosomes). An egg (or ova) cell is a gamete that accepts a sperm cell, another gamete. When the nuclei of these two cells join the unionized cell is called a zygote. The zygote would now be diploid.

2. How did the gametes get only one set of chromosomes if they themselves are a product of a diploid offspring. Somewhere along the line the homologous chromosomes split into separate cells by a process of meiosis, thus forming the gametes (gametogenisis).

II. What are the advantages of sex.

A. Diploid cells have a back up gene.

1. In case one gene fails via mutation, the other would act as backup. Such a mutation is usually not a problem because usually plenty of protein is made by the second allele. We say the gene is recessive.

a) A failed/recessive gene is usually not a problem unless the allele is also a failed gene, which can happen during fertilization of sexual reproduction from two different organisms. (brings up issues of recessive versus dominant, hybrid vigor, incest).

b) Occasionally the failed gene makes a functional protein, but it is usually bad for the organism. This type of disease is what we call dominant. The bad protein will be a problem irrelevant of the allele. (e.g Huntington's disease).

c) Very occasionally the altered gene makes a better! protein. This could give the organism with this gene an advantage and an evolutionary step may be in progress!

2. The failed gene can further mutate to something even better! The diploid condition gives more leeway for mutation without intervening failure. (on the other hand, it reduces its chance of expression).

3. A second allele can replicate, or jump to a new position, while a primary allele keeps the organism alive. The replicated or jumped gene can mutate into a new function!

B. Sexual reproduction offers genetic variability.

1. In mitotic reproduction, the offspring are identical to the parents (except for mutations). In sexually produced offspring, the offspring are not identical to their parents nor to their siblings. If an environmental crisis occurs, there are lots of different gene combinations that have a chance of surviving.

2. Crossing over (to be discussed later) causes mixing within chromosomes.

C. Two beneficial alleles produced in two different organisms can be brought together for the benefit of the species.

III. In mammals the female is XX and male is XY. The Y chromosome carries the sex determining gene that causes maleness. Without the gene, the default condition is femaleness (an XY person with a failed sex determining gene will be female).

MEIOSIS

I. Meiosis - the process:

A. Overall

1. The number of chromosomes in the resultant cells is half of that in the beginning cells.

2. Two nuclear divisions occur (each overtly appears much like that of mitosis: prophase, metaphase...). In division I, the two sets of chromosomes are actually split.

B. Division I

1. Meiotic Interphase I:

a) S: DNA replication occurs. It may take longer than the S phase of mitosis.

2. Meiotic prophase I:

a) Condensation and nucleolus disappears.

b) Pairing. Homologous chromosomes find each other by some mechanism and line up with each other. This coming together is called synapses or pairing. Thus, at this stage there are four closely related chromatids. Such a unit is called a tetrad. This stage can be very lengthy, or not.

c) Recombination = crossing over. The DNA breaks at a location and refuses with an adjacent homologous chromosome. This is a significant way of mixing genes. This happens fairly frequently. (Maybe three to four locations per chromosome during each meiosis). Text has some very good diagrams.

d) Transcription stage. The chromatids can decondense and the DNA can make m-RNA (transcription)! A large amount of cell growth occurs, particularly in eggs. In humans, the oocytes reach this stage at the 5th month of fetal life and stays there until ovulation, whereby one cell a month is triggered into the subsequent stages.

e) Recondensation.

f) the centrioles have replicated and separated (if present).

g) nuclear envelope breakdown signals the beginning of metaphase.

3. Meiotic metaphase I.

a) nuclear membrane disappears.

b) a mitotic apparatus of microtubules reaches from

(1) pole to pole

(2) and pole to chromosomal centromere.

c) The homologous pairs are lined up perpendicularly to the spindle apparatus.

4. Meiotic anaphase I. The homologous pairs are separated. Which goes to which side is completely random. From here, meiotic telophase may not be performed at all, and the system would directly skip into Meiotic metaphase II. All possible scenarios in between can occur depending upon the species of organism.

5. Meiotic telophase I. Decondensation and new nuclear membranes are formed.

II. Division II

1. Meiotic interphase II. DNA is NOT replicated.

2. Meiotic prophase II. Condensation

3. Meiotic metaphase II. Nuclear membrane disappears and chromosomes line up. Spindle apparatus appears.

4. Meiotic anaphase II. DNA split into two new regions.

5. Meiotic telophase II. Decondensation and new nuclear membranes formed.