Unit 7 Notes – Cell Cycle and Genetics

DNA

I.  DNA IS THE GENETIC MATERIAL

·  There are several famous experiments that proved that DNA is responsible for inheritance.

A.  Bacterial Transformation Experiment (Griffith’s Experiment)

·  Griffith used two strains of the Streptococcus pneumoniae bacterium that causes pneumonia in mammals. One strain was diseases-causing (pathogenic) while the other was a non-pathogenic strain.

·  Griffith could change the harmless strain of the bacterium into a disease causing strain just by mixing dead harmful bacteria with live harmless bacteria.

·  Transformation – 1. A change in genotype and phenotype due to the assimilation of external DNA by a cell.

·  Question remains: What is the factor that is responsible for transformation?

B.  Avery, McCarty and MacLeod’s Experiments:

·  They purified various molecules (proteins, carbohydrates, lipids and nucleic acids) from heat-killed bacteria and tried to use the purified molecules to transfer pathogenic characteristics into non-pathogenic bacteria.

·  They used bioassays to determine the pathogenicity of various molecules. A bioassay is determining the activity of various molecules by testing their effects on living organisms and comparing the activity to various known molecules’ activity.

·  Result: DNA is responsible for inheritance

C.  Bacteriophage Experiment (Hershey and Chase Experiment)

·  Bacteriophages – viruses that kill bacteria (viruses are mostly composed of DNA or RNA and proteins)

·  In this experiment, T2 phages were used that infect E. coli bacteria

·  Results: Phage proteins remained outside of the bacterial cells while phage DNA entered the cells – radioactive DNA was detected inside of infected bacteria.

·  Conclusion: DNA, not proteins are responsible for inheritance

D.  Chargaff’s Experiment:

·  He analyzed the base composition of DNA from various organisms.

·  Results:

a.  DNA composition varies from one species to another – evidence of molecular diversity among species

b.  In each species, the number of adenine bases approximately equaled the number of thymine bases; the number of cytosine bases equaled the number of guanine bases. (Chargaff’s rule)

E.  X-ray Crystallography (Franklin):

·  Her X-ray crystallography photograph of DNA leads to the discovery of the structure of DNA.

F.  Watson and Crick

·  Built the first model of the DNA molecule and described the structure of the molecule.

·  They also predicted the process of DNA replication but did not have experimental evidence to support it.

G.  Messelson-Stahl Experiment

·  Designed an experiment to determine DNA replication by tracing the origin of the replicated DNA.

·  To tell the difference between the parental (template) DNA and the newly synthesized complementary DNA they developed a tagging system by using two different kinds of nitrogen isotopes. One isotope (14N) is the normal, light nitrogen, while the other (15N) is a heavy isotope. They could use centrifugation to separate heavy, medium weight and light DNA molecules from each other.

·  Once they used heavy DNA as templates and allowed light DNA nucleotides to assemble with them, they got medium weight DNA molecules that showed that one half of the newly synthesized DNA was old (heavy) and the other half was new (light).

II.  DNA STRUCTURE

Review this section from biochemistry or from Module 45 from your book.

III.  DNA REPLICATION:

A.  The Basic Ideas on DNA Replication

·  Base-pairing rules apply when the DNA bases pair up

·  The two strands are complementary, so each strand serves as a template for ordering nucleotides into a new complementary strand

·  The process starts with one double helix and ends up with two DNA molecules, both double stranded and identical to the parent DNA

·  Enzymes link the nucleotides together at their sugar-phosphate groups.

·  The replication is semiconservative because every new DNA molecule contains one new strand and one old strand.

http://www.sumanasinc.com/webcontent/animations/content/meselson.html

B.  A Closer Look at DNA Replication

·  DNA replication is extremely accurate and efficient

·  Although we know more about DNA replication in prokaryotes than in eukaryotes, we also know that the two processes are very similar.

·  Six major steps of replication:

  1. Origins of replication: The site where DNA replication begins. In prokaryotic cells there is only one origin of replication, in eukaryotic cells there are hundreds or thousands to speed up replication. An enzyme (helicase) is used to untwist the DNA molecule.
  2. Initiation proteins recognize the origins of replication and open up the DNA double helix forming a replication bubble. Than replication proceeds in both directions until the entire DNA molecule is copied. Each opened DNA molecule where the replication takes place forms a replication fork. RNA nucleotides (primer) are used to mark the start of replication on each DNA polynucleotide chain.
  3. Elongating a new strand: Elongation is catalyzed by enzymes called DNA polymerases.
  4. Individual nucleotides align with complementary nucleotides along a template strand of DNA. DNA polymerase adds them one by one to the growing end of the new DNA molecule. DNA polymerize identifies the starting point by attaching to the prime of the RNA nucleotides In prokaryotes, there are two different DNA polymerases while in eukaryotes there are at least 11. The added nucleotides are actually nucleoside triphosphates (ATP, GTP, TTP, CTP but with deoxyribose sugar not ribose). When the Pi groups are broken down of the nucleotides, energy is released. This energy release fuels DNA replication.
  5. Antiparallel elongation: Because the two strands of the DNA molecule are antiparallel, so they are oriented in the opposite direction. The new DNA molecules also have to orient in the same direction. However, DNA polymerase can attach nucleotides only to the 3’ end and grows the chain toward the 5’end of the original chain. The original 3’- 5’ strand is called the leading strand because the DNA polymerase simply attaches new nucleotides by using the template of the old DNA chain and forms a new polynucleotide chain. To elongate the other new strand in the 5’- 3’ direction, the DNA polymerase works away from the replication fork, backwards. It forms small segments of the new polynucleotide chain that is going to be attached together later. These small segments are called Okazaki fragments. Another enzyme, (DNA ligase), attaches the Okazaki fragments together later.

  1. Only one primer is required to start the 3’ end but each Okazaki fragment requires a primer on the lagging strand. DNA polymerase I replaces the RNA primers with DNA molecular segments. An enzyme joins the sugar-phosphate backbones of the Okazaki fragments to form a continuous DNA polynucleotide chain.

http://www.wiley.com/college/pratt/0471393878/student/animations/dna_replication/index.html -- more detailed animation of DNA replication

http://highered.mcgraw-hill.com/sites/0072437316/student_view0/chapter14/animations.html -- many things on DNA replication and the experiments

http://207.207.4.198/pub/flash/24/menu.swf -- DNA replication, also very good

http://www.fed.cuhk.edu.hk/~johnson/teaching/genetics/animations/dna_replication.htm -- basic DNA replication

C.  Proofreading and Repairing DNA

·  In general there is only 1 error out of 10 billion nucleotides when the DNA molecule is being assembled. But the initial error rate is higher. DNA polymerases proofread the DNA molecule as it is being made and they replace the incorrectly placed nucleotide.

·  Cells also have special repair enzymes to fix incorrectly paired nucleotides later.

·  Most common factors that can result in the damage of DNA are: chemicals from metabolic reactions of the cell or from the environment, radioactive emissions, X-rays, UV light, spontaneous chemical changes of the DNA molecule.

IV.  Replicating the Ends of DNA Molecules:

·  Because the end of the DNA molecule on the lagging end runs out of 3’ ends, it cannot be copied. As a result, each repeated round of replication produce shorter and shorter DNA molecules. The part of chromosomes that get lost at each DNA replication is called the telomere. Telomeres do not contain genes, they only have multiple repetitions of one short nucleotide segments (TTAGGG in humans).

·  An enzyme called telomerase catalyzes the lengthening of telomeres in eukaryotic germ cells with the help of a short RNA segment.

THE CELL CYCLE

I. Overview

·  Cells divide to replace old, dead or damaged cells, to grow the organism, the heal wounds, to reproduce the organism and maintain the species.

·  Cells can divide by different methods:

o  Binary fission – the division type of bacteria and archaea

o  Mitosis – the type of cell division that produces identical copies of the same type of cell. This division is used in all cases when any somatic cell (body cell) forms.

o  Meiosis – produces haploid daughter cells that are genetically unique. These cells become the gametes (reproductive cells) of the organism.

II. Binary Fission

·  A simple process in which bacteria increase in size, replicate its simple circular DNA, grows some more and splits into two new identical cells.

·  Scientists use fluorescent molecules to view this process under microscope.

·  During harsh environments, bacteria switch to a division that produces endospores – resistant, dormant cell forms. Some of these forms can survive for thousands of years.

·  Some bacteria produce spores normally as part of their life cycle. Others can perform budding, growing a new cell on the tip of the parent cell.

III. Eukaryotic Cell Division

This is your spring break assignment

IV. Cell Cycle Control

·  Different types of cells divide at different schedules. Skin cells divide continuously, liver cells divide only if some other cells died around them, muscle cells don’t divide, they can only repair themselves.

·  Cell cycle control mechanisms determine if the cell will remain in the G1 or G0 phase of the cell cycle or continue to other phases.

·  There are several different control mechanisms.

·  Major cell cycle controls include:

o  Checkpoints – Protein signaling pathways are used to interact with molecules that are part of the cell cycle. These protein signaling pathways allow the cell cycle to continue only if the molecules that they interact with are complete and healthy. There is a checkpoint in the G1 phase, that check the size of the cell and the condition of the DNA. If this checkpoint is not passed, the cell remains in G0 phase and will not divide. There is another checkpoint in G2 to check if the DNA replicated properly in S phase and if the cell is large enough to divide. There is a last checkpoint during mitosis in metaphase to check if all the chromosomes attached to the spindle fibers.

o  Cyclin and kinase complexes – The cell cycle is also regulated by proteins called cyclins that increase in concentration at the end of each phase of the cell cycle. Cyclins bind with cyclin-dependent kinase (Cdks) molecules that are molecules with a constant concentration in the cell. The cyclin-cdk complexes are activated together to help the cell to switch to another phase of the cell cycle. After a new phase begins, the cyclins break down and the cdks become inactive again until the next increase in cyclin concentration. There are different types of cyclins produced in different phases of the cell cycle. These cyclin-cdk complexes trigger various cell signaling pathways.

·  Cell cycle controls respond to various internal and external cues to determine when the cell needs to divide.

·  External cues to regulate the cell cycle include:

o  Availability of nutrients

o  Growth factors – paracrine signals that are released by all kinds of cells to stimulate the division of certain area cells

o  Density-dependent inhibition – cells don’t divide if they touch other cells on all sides. There are surface proteins that bind cells to other cells. When all surface proteins are connected to other cells, the cell division will be inhibited until space opens up around the cell. Cancer cells don’t stop dividing and don’t recognize density-dependent inhibition. As a result, they form tumors.

o  Anchorage dependence – most animal cells don’t divide if they are not anchored to surface tissues. Cancer cells may not follow anchorage dependence, so they can divide during metastasis.

·  Internal factors include the age of the cell. Normal cells divide only about 20-50 times before they die. Cancer cells can divide indefinitely so they are considered immortal.

V.  What Happens When the Regulatory Mechanisms Fail?

·  Cancer is uncontrolled cell division of abnormal cells in the body.

·  Cancer cells are different from normal cells because they don’t stop dividing when they fill up the available space or when they are not anchored to surfaces. They are also immortal in the sense that they don’t stop dividing.

·  Cancer is caused by a set of mutations. These mutations make the cell recognizable for the immune system and in most cases cytotoxic T cells and natural killer cells destroy cancer cells. But in some cases, the immune system fails to detect these cells. Than after further mutations the following stages of cancer occur:

o  Stage 1 – benign tumors – increased cell division forms small tumors

o  Stage 2 – benign tumors – small tumors increase in size and grow blood vessels to get nutrient and oxygen supply

o  Stage 3 – malignant tumors – tumor cells get into the blood stream and move to other parts of the body (metastasis)

o  Stage 4 – malignant tumors – secondary tumors grow all over the body.

·  Cancer cells may look different under the microscope from normal cells and various changes occur in them.

·  Genes that cause cancer are called oncogenes – the normal forms of these genes are called proto-oncogenes. These genes are normally responsible for controlling the cell cycle by for example producing cyclin proteins.

·  Other genes called tumor-suppressor genes normally inhibit cell division. These genes stop cell division or stimulate apoptosis if the cell has mutations in their oncogenes. The most frequently altered tumor-suppressor gene is p53, which is mutated in more than half of all tumors in humans.

Biointeractive – p53 activity

Use your honors biology notes for the description of the cell cycle, mitosis and meiosis.

THE SEXUAL LIFE CYCLE

I. The Organization of Chromosomes