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.

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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 Tumor- Suppressor 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