Questions. Chapter 18 - Genetics of Viruses and Bacteria

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1 Chapter 18 - Genetics of Viruses and Bacteria Questions 1. The proteins that encapsulate the genetic material of a virus is known as the. 2. Draw a general structure of a eukaryotic virus and label parts. 3. An individual protein of the structure mentioned in question number 1 is known as a. 4. A bacteriophage can reproduce via two different life cycles known as the and. 5. The genetic material of viruses can be,, or. 6. This general structure is found to be part of some viruses like Influenza and not part of other viruses like Adenovirus.

2 Chapter 18 - Genetics of Viruses and Bacteria NEW AIM: Viruses: Packaged Genes Viruses : Packaged Genes

3 Chapter 18 - Genetics of Viruses and Bacteria NEW AIM: Viruses: Packaged Genes What is a virus? 1. Obligate intracellular parasite - A small [20 to 250nm in diameter] infectious agent that requires a host cell to replicate (make more of itself). **1/1000 th the diameter of a eukaryotic cell. If the classroom was a cell, a virus would be about the size of a paperclip. 2. General Structure Nucleic acid enclosed in a protein coat and, in some cases, a membranous envelope 3. Host Range - Each virus can only infect a specific range of cell types Ex. HIV can only infect CD4+ Helper T-cells SEM of adenovirus

4 Chapter 18 - Genetics of Viruses and Bacteria NEW AIM: Viruses: Packaged Genes Size Comparison Virus: 20 to 250nm (.02 to.25um) Prokaryote: 1 to 10um Eukaryote: 10 to 100um

5 Chapter 18 - Genetics of Viruses and Bacteria NEW AIM: Viruses: Packaged Genes CAPSID 1. All viruses contain genetic material (DNA or RNA) encapsulated by a protein coat called a capsid. 2. An individual protein in the capsid is called a capsomere. 3. Bacteriophage (phage) have the most complex capsids Capsomere of capsid RNA Capsomere DNA Membranous envelope Capsid RNA Head Tail sheath Tail fiber DNA Glycoprotein mm nm (diameter) Glycoprotein nm (diameter) nm 20 nm 50 nm (a) Tobacco mosaic virus (b) Adenoviruses 50 nm (c) Influenza viruses 50 nm (d) Bacteriophage T4

6 Chapter 18 - Genetics of Viruses and Bacteria NEW AIM: Viruses: Packaged Genes Influenza looks different it has an envelope. What s up with that? Capsomere of capsid RNA Capsomere DNA Membranous envelope Capsid Head Tail sheath DNA RNA Tail fiber Glycoprotein mm nm (diameter) Glycoprotein nm (diameter) nm 20 nm 50 nm (a) Tobacco mosaic virus (b) Adenoviruses 50 nm (c) Influenza viruses 50 nm (d) Bacteriophage T4

7 Chapter 18 - Genetics of Viruses and Bacteria NEW AIM: Viruses: Packaged Genes Envelopes 1. Only some viruses have cell membrane-like envelopes Ex. Influenza (shown right) Membranous envelope Capsid RNA 2. The envelope is derived (comes from) the cell membrane of the host cell Glycoprotein nm (diameter) 50 nm (c) Influenza viruses

8 Chapter 18 - Genetics of Viruses and Bacteria NEW AIM: Viruses: Packaged Genes How do viruses replicate (reproduce)? Viruses Hijack Cells They gain access and use the enzymes, ribosomes, and small molecules (ATP, nucleotides, amino acids, phospholipids, etc ) of host cells. Simplified viral reproductive cycle

9 Chapter 18 - Genetics of Viruses and Bacteria AIM: Viruses: Packaged Genes Let s begin with the best understood virus: T4 Phage infecting E. coli 1. Bacterial virus (bacteriophage or just phage) How do they reproduce?

10 Chapter 18 - Genetics of Viruses and Bacteria NEW AIM: Viruses: Packaged Genes Fig Bacteriophage reproductive cycle (two methods of reproduction) Bacteriophage binds to the surface of the bacterium using the tail fibers and injects its DNA into the cell

11 Chapter 18 - Genetics of Viruses and Bacteria NEW AIM: Viruses: Packaged Genes Fig Bacteriophage reproductive cycle (two methods of reproduction) Lysogenic cycle

12 Chapter 18 - Genetics of Viruses and Bacteria NEW AIM: Viruses: Packaged Genes Fig Bacteriophage reproductive cycle (two methods of reproduction) Lytic cycle Lysogenic cycle

13 Chapter 18 - Genetics of Viruses and Bacteria NEW AIM: Viruses: Packaged Genes Lysogenic cycle - After the bacteriophage injects its DNA, it might get incorporated into the bacterial chromosome and is now called a prophage. Now when the bacterial cells replicates, the phage DNA replicates with it. Lytic cycle - After the bacteriophage injects its DNA or when the prophage jumps out of the DNA, it can hijack the cell and use it (its ribosomes and other enzymes) to make more viral DNA and proteins to in turn make more viral particles. The cell will lyse and the viruses will be released. Temperate Phages - Phages that can do both lytic and lysogenic methods of reproduction Ex. Lambda (λ) phage

14 Chapter 18 - Genetics of Viruses and Bacteria NEW AIM: Viruses: Packaged Genes What causes a temperate phage like lambda to switch from lysogenic to lytic? We observed the switch to be caused by environmental factors like radiation or certain chemicals causing DNA damage, which would promote the lytic phase as the bacterial cell will likely die soon and the phage needs to get out quick. In addition, lytic is favored when nutrients are plentiful allowing the phage to makes lots more of itself, while the lysogenic is favored when nutrients are in low concentration within the bacterium. This makes sense as the virus can lay low until better times. Can t make more of yourself if the materials are simply not available.

15 Chapter 18 - Genetics of Viruses and Bacteria NEW AIM: Viruses: Packaged Genes Can prokaryotes defend themselves against this attack? Of course. They contain enzymes that attempt to hydrolyze the viral DNA known as restriction enzymes like little molecular scissors.

16 Chapter 18 - Genetics of Viruses and Bacteria AIM: Viruses: Packaged Genes 2. Animal viruses A. Anatomy Genetic Material Can be ssdna/dsdna or ssrna/dsrna depending on the virus. Codes for polypeptides/proteins needed by the virus to enter and hijack the cell as well as the proteins of the capsid and envelope. Capsid made of proteins and surrounds the genetic material in the core. Envelope Phospholipid bilayer similar to a cell membrane with embedded proteins (protein spikes) surrounding the capsid. Not all virus types have envelopes

17 Chapter 18 - Genetics of Viruses and Bacteria AIM: Viruses: Packaged Genes 2. Animal viruses Capsid Protein spikes DNA

18 Chapter 18 - Genetics of Viruses and Bacteria AIM: Viruses: Packaged Genes 2. Animal viruses They are classified by their genetic material.

19 Chapter 18 - Genetics of Viruses and Bacteria AIM: Viruses: Packaged Genes 2. Animal viruses DNA viruses

20 Chapter 18 - Genetics of Viruses and Bacteria AIM: Viruses: Packaged Genes 2. Animal viruses B. DNA viruses DNA capsid envelope

21 Chapter 18 - Genetics of Viruses and Bacteria AIM: Viruses: Packaged Genes 2. Animal viruses B. DNA viruses Ex. Adenovirus - Causes upper respiratory infections - Symptoms range from those similar to the common cold to bronchitis or pneumonia. (Common cold is caused by rhinovirus, an RNA virus)

22 Chapter 18 - Genetics of Viruses and Bacteria AIM: Viruses: Packaged Genes 2. Animal viruses B. DNA viruses Ex2. Herpesviruses (family of related viruses) These can cause: 1. Oral herpes (cold sores) or genital herpes (an STD)

23 Chapter 18 - Genetics of Viruses and Bacteria AIM: Viruses: Packaged Genes 2. Animal viruses B. DNA viruses Ex2. Herpesviruses (family of related viruses) These can cause: 2. Chicken pox (varicella zoster virus)

24 Chapter 18 - Genetics of Viruses and Bacteria AIM: Viruses: Packaged Genes 2. Animal viruses B. DNA viruses Ex3. Poxvirus (family of related viruses) Can cause: 1. Small pox This is the only human infectious disease to ever be eradicated (removed from the face of the planet) we did this through extensive vaccination.

25 Chapter 18 - Genetics of Viruses and Bacteria AIM: Viruses: Packaged Genes 2. Animal viruses B. DNA viruses Ex4. HPV Human Papillomavirus A. Over 200 different types many are STDs (sexually transmitted) 1. Some of these STD viruses can lead to cancers of the cervix, vagina, and anus in women or cancers of the anus and penis in men. a. Nearly all cases of cervical cancer are caused by HPV 2. Others cause genital warts

26 Chapter 18 - Genetics of Viruses and Bacteria AIM: Viruses: Packaged Genes 2. Animal viruses B. DNA viruses HPV Vaccine Recommended by CDC for all females and males age 11 to 26.

27 Chapter 18 - Genetics of Viruses and Bacteria NEW: Viruses: Packaged Genes What do viruses need to accomplish to continue to exist? 1. Gain access to a cell 2. Use the cell s workers (ribosomes, RNA polymerase, etc ) to make more of itself. a. Synthesize viral proteins b. Replicate its genome c. Assemble these into new viral particles

28 Chapter 18 - Genetics of Viruses and Bacteria NEW: Viruses: Packaged Genes Life cycle of a DNA virus What is the first thing a virus must be able to do? 1. Viral Attachment and Entry a. If the virus does not have an envelope, protein spikes on the capside will act as ligands and bind cell receptors, triggering receptor mediated endocytosis.

29 Chapter 18 - Genetics of Viruses and Bacteria NEW: Viruses: Packaged Genes Life cycle of a DNA virus 1. Viral Attachment and Entry b. If it does have an envelope, the protein spikes in the envelope will act as ligands and bind to cell receptors resulting in fusion of the viral membrane and cell membrane, injecting the capsid into the cell

30 Chapter 18 - Genetics of Viruses and Bacteria NEW: Viruses: Packaged Genes Life cycle of a DNA virus 1. Viral Attachment and Entry Analogy: Cell receptors = door lock Protein spikes = the key In either case, the protein spikes on the surface need to bind receptors to gain access to the cell, which is why specific viruses can only infect specific cells with matching receptors.

31 Chapter 18 - Genetics of Viruses and Bacteria NEW: Viruses: Packaged Genes Life cycle of a DNA virus A. Viral attachment and entry B. Uncoating The capsid fall apart and the viral DNA enters the nucleus C. Transcription and translation of the viral DNA The viral DNA is transcribed and translated by our workers (our RNA polymerases, ribosomes/trnas/ etc ) using our ATP made by our mitochondria!!

32 Chapter 18 - Genetics of Viruses and Bacteria NEW: Viruses: Packaged Genes Life cycle of a DNA virus D. Replication of the viral DNA E. Viral protein sorting Capsid proteins are brought into the nucleus while envelope proteins get into nuclear membrane via endomembrane system.

33 Chapter 18 - Genetics of Viruses and Bacteria NEW: Viruses: Packaged Genes Life cycle of a DNA virus F. Viral assembly Capsid forms around DNA and then buds out of nucleus picking up its envelope H. Release How the virus, now in the cytoplasm, gets out of the cell is not understood yet.

34 Chapter 18 - Genetics of Viruses and Bacteria NEW: Viruses: Packaged Genes Life cycle of a DNA virus This process typically happens over and over and over again until the cell dies The cell is a virus producing factory. DNA integration In certain viruses, like Herpes virus, the viral DNA can integrate (become part of) the cell s DNA (your DNA), and sit quietly similar to the lysogenic cycle of bacteriophages. Almost all adults carry Herpes Simplex 1 virus (oral herpes).

35 Chapter 18 - Genetics of Viruses and Bacteria AIM: Viruses: Packaged Genes 2. Animal viruses RNA viruses

36 Chapter 18 - Genetics of Viruses and Bacteria AIM: Viruses: Packaged Genes 2. Animal viruses C. RNA viruses Ex1. Mumps virus - Member of the paramyxovirus family - Causes the mumps Extreme swelling of salivary glands Before infection After infection Contagious via respiratory secretions (coughing/sneezing/sharing glass/kissing/ etc )

37 Chapter 18 - Genetics of Viruses and Bacteria AIM: Viruses: Packaged Genes 2. Animal viruses C. RNA viruses Ex2. Rubella virus - Member of the togavirus family - Causes rubella (German measles) Rash on body Flu-like symptoms Highly Contagious

38 Chapter 18 - Genetics of Viruses and Bacteria AIM: Viruses: Packaged Genes 2. Animal viruses C. RNA viruses Ex3. Measles - Caused by a member of the paramyxovirus family like mumps - Highly contagious through respiratory secretion just like mumps Symptoms: Rash on body, cough, runny nose, red eyes, four day fevers

39 Chapter 18 - Genetics of Viruses and Bacteria AIM: Viruses: Packaged Genes 2. Animal viruses C. RNA viruses If these viruses are so easily contagious, why haven t you gotten them? You have all been vaccinated against them (MMR shot) MMR = measles, mumps, rubella

40 Chapter 18 - Genetics of Viruses and Bacteria AIM: Viruses: Packaged Genes 2. Animal viruses C. RNA viruses Ex4. Poliomyelitis (polio) - Highly contagious through fecal-oral route (feces to the mouth) It is easier than you think the chef prepares your food and didn t wash his hands - In 1% of infections, virus enter neurons and destroys motor function lose control of your muscles You are vaccinated against this one too

41 Animal RNA virus life cycle 1. Viral attachment and entry Similar to DNA virus protein spikes act as ligands for cell receptors. 2. Uncoating Capsid falls apart releasing the RNA 3. RNA synthesis A viral enzyme will make the complementary RNA strand (purple) using the genomic RNA (red) as a template 4. Protein synthesis Complementary RNA can act as mrna and your ribosomes will translate it, making new viral proteins. Fig a

42 Animal RNA virus life cycle 5. Synthesizing more genomic RNA The complementary strand (purple) can also act as a template to back synthesize the more genomic RNA (red) 6. Assembly The viral proteins and genomic RNA come together to make new viral particles. Some of the viral proteins made were sent through the endomembrane system to the cell membrane. Fig a

43 Animal RNA virus life cycle 7. Exit The capsid/rna pinch off from the cell, which is how it acquires the envelope with embedded viral proteins. -Notice that the nucleus is not involved. -This process happens again and again until the cell is dead. -There can be no integration of standard RNA viruses into our genome as RNA cannot be integrated into DNA Fig a

44 The reproductive cycle of an enveloped RNA virus Capsid RNA 1 Glycoproteins on the viral envelope bind to specific receptor molecules (not shown) on the host cell, promoting viral entry into the cell. Envelope (with glycoproteins) 2 Capsid and viral genome enter cell Template HOST CELL Viral genome (RNA) 3 The viral genome (red) functions as a template for synthesis of complementary RNA strands (pink) by a viral enzyme. 5 Complementary RNA strands also function as mrna, which is translated into both capsid proteins (in the cytosol) and glycoproteins for the viral envelope (in the ER). ER mrna Glycoproteins Capsid proteins Copy of genome (RNA) 4 New copies of viral genome RNA are made using complementary RNA strands as templates. 6 Vesicles transport envelope glycoproteins to the plasma membrane. Figure A capsid assembles around each viral genome molecule. 8 New virus

45 Chapter 18 - Genetics of Viruses and Bacteria AIM: Viruses: Packaged Genes 2. Animal viruses C. RNA viruses Ex5. Retrovirus Ex. HIV (human immunodeficiency virus) you will need to know the details on this one

46 Chapter 18 - Genetics of Viruses and Bacteria NEW: Viruses: Packaged Genes Retroviruses - A special family of RNA viruses - Retro implies Reverse - These viruses have an RNA genome, but use a special enzyme called Reverse Transcriptase to make a DNA copy of the RNA (the reverse of transcription; hence the name) Ex. HIV (human immunodeficiency virus)

47 Chapter 18 - Genetics of Viruses and Bacteria NEW: Viruses: Packaged Genes Retroviruses Fig 10.21A Attachment protein is called GP120 HIV HIV - Enveloped RNA virus - Capsid houses two identical RNA molecules and the enzyme Reverse Transcriptase as well as others needed for the virus to function. Why do you think the virus needs to carry its own Reverse Transcriptase? Because our cells do not have the gene for reverse transcriptase

48 Chapter 18 - Genetics of Viruses and Bacteria NEW: Viruses: Packaged Genes Retroviruses HIV How is HIV transmitted? The virus is transmitted through contact of a bodily fluid containing HIV like blood, semen, vaginal fluid, and breast milk with a mucous membrane or the bloodstream. A. ~33 million people are HIV positive in the world. B. Estimated 1.1 million people are HIV positive in the US. C. ~2.2 million people, 330,000 of which were children, died as a result of the virus last year 75% of deaths occurred in Sub-Saharan Africa.

49 Chapter 18 - Genetics of Viruses and Bacteria NEW: Viruses: Packaged Genes Retroviruses Fig 10.21A HIV What disease does HIV cause? - AIDS Acquired Immune Deficiency Syndrome Immune system gradually declines leaving the individual susceptible to opportunistic infections like tuberculosis (5 10% of Americans test positive for the bacterium that causes tuberculosis, but the immune system keeps it in check and the person is fine)and tumors (many cells that would have caused cancer are destroyed by the immune system). Therefore, HIV/AIDS does not kill anyone directly, it is the opportunistic infection or cancer that kills the person.

50 Chapter 18 - Genetics of Viruses and Bacteria NEW: Viruses: Packaged Genes Retroviruses HIV How does HIV cause AIDS? HIV (blue dots) infects, hijacks and in the end destroys Helper T-cells (red) (special type of cell of the human immune system required for proper function). Let s look at how HIV infects Helper-T cells

51 Chapter 18 - Genetics of Viruses and Bacteria NEW: Viruses: Packaged Genes HIV Life Cycle GP120 Attachment and Entry: HIV envelope glycoprotein GP120 (ligand) binds to the CD4 receptor on the surface of the Helper T-cell resulting in fusion of the viral envelope with the cell membrane thereby allowing the capsid to enter the cell and fall apart releasing the viral RNA and Reverse transcriptase enzymes.

52 Chapter 18 - Genetics of Viruses and Bacteria NEW: Viruses: Packaged Genes HIV Life Cycle This figure skips the attachment and entry and uncoating of the viral particle. Fig 10.21B

53 Chapter 18 - Genetics of Viruses and Bacteria NEW: Viruses: Packaged Genes HIV Life Cycle 1. Reverse Transcriptase makes a DNA copy (blue) of the viral RNA genome (red). 2. Reverse Transcriptase then removes the RNA and synthesizes the complementary DNA strand. 3. Integration: the dsdna enters the nucleus and gets integrated (inserted) into the DNA. Fig 10.21B

54 Chapter 18 - Genetics of Viruses and Bacteria NEW: Viruses: Packaged Genes HIV Life Cycle 4/5. Transcription/Translation: viral RNA and proteins are synthesized from the provirus (analogous to prophage) DNA. 6. Assembly: viral particles are assembled and bud off the cell This process happens over and over again as long as the Helper T-cell lasts Fig 10.21B

55 The reproductive cycle of HIV, a retrovirus HIV Membrane of white blood cell 1 The virus fuses with the cell s plasma membrane. The capsid proteins are removed, releasing the viral proteins and RNA. 2 Reverse transcriptase catalyzes the synthesis of a DNA strand complementary to the viral RNA. Viral RNA HOST CELL Reverse transcriptase 3 Reverse transcriptase catalyzes the synthesis of a second DNA strand complementary to the first µm HIV entering a cell RNA-DNA hybrid DNA 4 The double-stranded DNA is incorporated as a provirus into the cell s DNA. RNA genome for the next viral generation Chromosomal DNA mrna NUCLEUS Provirus 5 Proviral genes are transcribed into RNA molecules, which serve as genomes for the next viral generation and as mrnas for translation into viral proteins. 6 The viral proteins include capsid proteins and reverse transcriptase (made in the cytosol) and envelope glycoproteins (made in the ER). Figure New HIV leaving a cell 9 New viruses bud off from the host cell. 8 Capsids are assembled around viral genomes and reverse transcriptase molecules. 7 Vesicles transport the glycoproteins from the ER to the cell s plasma membrane.

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57 Chapter 18 - Genetics of Viruses and Bacteria NEW: Viruses: Packaged Genes What determines the damage a virus does? One item is the type of cell it infects Examples: HIV immune system cells Influenza respiratory cells Polio neurons (can t divide)

58 Chapter 18 - Genetics of Viruses and Bacteria NEW: Viruses: Packaged Genes Vaccinations 1. Edward Jenner A. Credited with discovering the first vaccine in 1798 B. The disease was small pox C. He observed that milk maids (people that milked cows) did not get small pox. D. Took the pus from these people infected with cow pox (a similar virus to small pox that you catch from cows) and injected it into other people. E. The cow pox pus somehow protected these people against small pox

59 Chapter 18 - Genetics of Viruses and Bacteria NEW: Viruses: Packaged Genes Vaccinations 2. How do vaccines work? - By injecting the cowpox pus, the immune system mounts an attack against the virus in the pus. - The immune system remembers the foreign substances it attacks and is prepared if it attacks again - Since the small pox virus is so similar to the cow pox virus, the immune system is prepared for the small pox virus as well...

60 Chapter 18 - Genetics of Viruses and Bacteria NEW: Viruses: Packaged Genes Vaccinations 2. How do vaccines work? - Most modern day vaccines are typically an injection of dead or weakened (attenuated) viruses or viral proteins more about this when we look into the immune system in detail.

61 Chapter 24: The Immune System NEW AIM: How does the body defend itself against MO s? I. Nonspecific vs. Specific Immunity B. Specific immunity (The Immune System) - OVERVIEW Memory T-cells are also made from T- cells activated by Helper T-cells. For a future encounter with the same antigen carrying pathogen.

62 Chapter 24: The Immune System NEW AIM: How does the body defend itself against MO s? I. Nonspecific vs. Specific Immunity B. Specific immunity (The Immune System) vii. Memory cells Primary immune response a. Memory B and T-cells are reservists for next time that specific antigen shows up: The first time the lymphocytes see the antigen. Antibodies are made, but relatively slowly due to the small number of B-cells activated and only a relatively small number of antibodies are made compared to the second time the lymphocytes see the antigen for the same reason. Secondary immune response The secondary response results upon reexposure to the antigen. You have millions of memory B-cells. Most of them will be activated and antibodies are made quickly and in large number thanks to the large number of cells. You do not get sick. It must be the same antigen. Any mutation that changes the structure of the antigen will not elicit the secondary response. Fig. 24.8

63 Chapter 18 - Genetics of Viruses and Bacteria NEW: Viruses: Packaged Genes Vaccinations

64 Chapter 18 - Genetics of Viruses and Bacteria NEW: Viruses: Packaged Genes Vaccinations

65 Chapter 18 - Genetics of Viruses and Bacteria NEW: Viruses: Packaged Genes Fig Tobacco Mosaic Virus Plants get viruses too

66 Chapter 18 - Genetics of Viruses and Bacteria Transmission Bacterial and Viral Transmission 1. Droplet Contact - coughing or sneezing on another person Ex. Chicken pox, common cold (rhinovirus), influenze (flu), Tuberculosis, Measles, Mumps, Rubella, Pertussis, Strep throat

67 Chapter 18 - Genetics of Viruses and Bacteria Transmission Bacterial and Viral Transmission 2. Direct Physical Contact - touching an infected person, including sexual contact Ex. Sexually transmitted diseases, Athlete s foot (fungal), Warts

68 Chapter 18 - Genetics of Viruses and Bacteria Transmission Bacterial and Viral Transmission 3. indirect contact - usually by touching a contaminated surface like a door knob or your desk. (ex. Rhinovirus common cold) 4. airborne transmission - if the microorganism can remain in the air for long periods (essentially droplet transmission) 5. fecal-oral transmission - usually from contaminated food or water sources (cholera, hepatitis A, polio, rotavirus, salmonella) 6. vector borne transmission - carried by insects or other animals (malaria a protist)

69 Chapter 18 - Genetics of Viruses and Bacteria Transmission Bacterial and Viral Transmission This is why surgeons look like this

70 Chapter 18 - Genetics of Viruses and Bacteria Transmission Bacterial and Viral Transmission and people working in a biosafety level 4 laboratory look like this

71 Chapter 18 - Genetics of Viruses and Bacteria Transmission Bacterial and Viral Transmission Biosafety Levels Examples Non-pathogenic E. coli (Escherichia coli) Hepatitis A, B, C, influenza Tuberculosis, West Nile Virus, Anthrax Ebola virus, small pox, Argentine hemorrhagic fevers, Marburg virus, Lassa fever, Crimean-Congo hemorrhagic fever

72 Chapter 18 - Genetics of Viruses and Bacteria Transmission Viroids 1. Circular RNA molecules that infect plants (only several hundred nucleotides long) 2. DO NOT encode proteins 3. The RNA molecules replicate inside plant cells using their machinary THEY ARE JUST SINGLE MOLECULE!! TEM of circular viroid RNA (black rings) Plants infected with varying degrees of viroid particles (control on left)

73 Chapter 18 - Genetics of Viruses and Bacteria Transmission Prions 1. Infectious Protein!! 2. Cause a number of degenerative brain diseases in various animals Ex. scrapie in sheep, mad cow disease in cows, Creutzfeldt-Jakob disease in humans 3. Transmitted through ingestion of food with these prions in them like eating beef from cattle that had mad cow disease. ALARMING CHARACTERISTICS 1. They are slow-acting - Takes about 10 years until you see symptoms 2. Virtually Indestructable - They are not destroyed (denatured) by heating to normal cooking temperatures

74 Chapter 18 - Genetics of Viruses and Bacteria Transmission Prions How can a protein, which cannot replicate itself, be a transmissible pathogen? Hypothesis: - A prion is a misfolded form of a protein normally present in brain cells - When the prion gets into a normal cell, with the normal form of the protein, it converts the normal protein to the prion form. You eat prion infected beef Prion gets into neurons in your brain and turn normal protein into prion form chain reaction.

75 Chapter 18 - Genetics of Viruses and Bacteria Transmission BACTERIAL GENETICS

76 Chapter 12 - DNA Technology and the Human Genome How can we use bacteria to manipulate DNA and protein? How do bacteria (prokaryotes) they take up DNA (it is more than just mutation that gives certain species of bacteria their genetic diversity)

77 Chapter 12 - DNA Technology and the Human Genome How can we use bacteria to manipulate DNA and protein? Reproduce by binary fission

78 Chapter 12 - DNA Technology and the Human Genome How can we use bacteria to manipulate DNA and protein? Reproduce by binary fission Replication of single, circular bacterial chromosome preceding binary fission How do bacteria maintain genetic diversity? One way is through mutation since they can reproduce so quickly leading to millions upon billions of slightly different individuals in only a days time.

79 Chapter 12 - DNA Technology and the Human Genome How can we use bacteria to manipulate DNA and protein? Reproduce by binary fission Is this the only way they maintain diversity? Replication of single, circular bacterial chromosome preceding binary fission Absolutely not let s look at other ways to do this

80 Chapter 12 - DNA Technology and the Human Genome How can we use bacteria to manipulate DNA and protein? Look at this experiment and explain what is being observed: How were these bacteria able to exchange genes (DNA)?

81 Chapter 12 - DNA Technology and the Human Genome How can we use bacteria to manipulate DNA and protein? Three major methods have evolved by which bacteria take up foreign DNA to enhance diversity: 1. Transformation 1. Bacteria can take up a free piece of bacterial DNA 2. Crossing-over will occur between exogenous DNA and the bacterial chromosome. Fig. 12.1A-C Recall Griffith s experiment

82 Chapter 12 - DNA Technology and the Human Genome How can we use bacteria to manipulate DNA and protein? There are three methods by which bacteria take up DNA in nature. 2. Transduction Bacteriophage is mistakenly packaged with bacterial DNA. Injects this DNA into another bacteria. Recall Hershey and Chase Fig. 12.1A-C

83 Chapter 12 - DNA Technology and the Human Genome How can we use bacteria to manipulate DNA and protein? 2. Transduction Bacteriophage is mistakenly packaged with bacterial DNA. Injects this DNA into another bacteria. Fig. 18.6

84 Chapter 12 - DNA Technology and the Human Genome How can we use bacteria to manipulate DNA and protein? There are three methods by which bacteria take up DNA in nature. 3. Conjugation Male (F+) bacteria extend sex pili called a mating bridge (long tube) to female (F-) bacteria. Part of chromosome is replicated and transferred. F+ F-

85 Chapter 12 - DNA Technology and the Human Genome How can we use bacteria to manipulate DNA and protein? There are three methods by which bacteria take up DNA in nature. 3. Conjugation F+ means the cell has the so-called F (fertility) factor What is an F factor? It is a special segment of DNA that can be part of: F+ F- 1. The bacterial chromosome OR 2. A plasmid Now what s a plasmid? Bacteria can have small, circular extra-chromasomal (not the chromosome) pieces of DNA.

86 Chapter 12 - DNA Technology and the Human Genome How can we use bacteria to manipulate DNA and protein? Lysed bacterium The majority of the DNA above that has spilled out of the bacterium is chromosomal, but you can see smaller circular pieces not part of the chromosome plasmids.

87 Chapter 12 - DNA Technology and the Human Genome How can we use bacteria to manipulate DNA and protein? Plasmid - Small, circular piece of DNA distinct from bacterial chromosome - has own origin of replication (ori) - carries genes in nature or humans can modify them and insert genes into the so-called polylinker region - called vectors when used by humans as tools of genetic engineering

88 Chapter 12 - DNA Technology and the Human Genome How can we use bacteria to manipulate DNA and protein? There are three methods by which bacteria take up DNA in nature. 3. Conjugation F+ means the cell has the so-called F (fertility) factor F+ F- The F plasmid A special plasmid containing the F factor plus some 25 other genes needed for the production of sex pili. ***This plasmid has the ability to integrate into the chromosome of the bacterium or remain separate (see next slide). F+ cells have the F plasmid and can form sex pili and exchange DNA with an F- cell.

89 Chapter 12 - DNA Technology and the Human Genome How can we use bacteria to manipulate DNA and protein? There are three methods by which bacteria take up DNA in nature. 3. Conjugation The F- cell is now and F+ cell because it now has the F plasmid and can form sex pili with other F- cells and pass along DNA. Fig

90 Chapter 12 - DNA Technology and the Human Genome How can we use bacteria to manipulate DNA and protein? There are three methods by which bacteria take up DNA in nature. 3. Conjugation As mentioned earlier, the F plasmid has the potential to integrate into the chromosome of the bacterium as shown above resulting in what we call an Hfr (High frequency of recombination) cell. Fig

91 Chapter 12 - DNA Technology and the Human Genome How can we use bacteria to manipulate DNA and protein? There are three methods by which bacteria take up DNA in nature. 3. Conjugation

92 Chapter 12 - DNA Technology and the Human Genome How can we use bacteria to manipulate DNA and protein? There are three methods by which bacteria take up DNA in nature. 3. Conjugation

93 Chapter 12 - DNA Technology and the Human Genome How can we use bacteria to manipulate DNA and protein? There are three methods by which bacteria take up DNA in nature. 3. Conjugation

94 Chapter 12 - DNA Technology and the Human Genome How can we use bacteria to manipulate DNA and protein? There are three methods by which bacteria take up DNA in nature. 3. Conjugation

95 Chapter 12 - DNA Technology and the Human Genome How can we use bacteria to manipulate DNA and protein? There are three methods by which bacteria take up DNA in nature. 3. Conjugation Now when the plasmid begins to replicate, it will also replicate part of the bacterial chromosome giving new genes to the recipient cell. Crossing over and therefore recombination will occur within the recipient.

96 Chapter 12 - DNA Technology and the Human Genome How can we use bacteria to manipulate DNA and protein? There are three methods by which bacteria take up DNA in nature. 3. Conjugation Complete picture of the two possibilities Fig

97 Chapter 12 - DNA Technology and the Human Genome How can we use bacteria to manipulate DNA and protein? 1. Transformation 2. Transduction 3. Conjugation Where have we observed transformation before in this class? The Griffith experiment when he mixed the R strain with the heat-killed S strain

98 Chapter 12 - DNA Technology and the Human Genome How can we use bacteria to manipulate DNA and protein? R plasmids (aside) β-lactam ring 1. R stands for resistance 2. These are bacterial plasmids that carry genes that confer resistance to antibiotics like ampicillin ampicillin 3. The gene that confers resistance is called AmpR (ampicillin resistance). What protein does is code for? It encodes the protein β-lactamase Guess what is does: β-lactamase with ampicillin bound in the active site

99 Chapter 12 - DNA Technology and the Human Genome How can we use bacteria to manipulate DNA and protein? Transposable Elements (Transposons) 1. Also known as jumping genes Insertion sequence Nobel Prize, Cold Spring Harbor 5ʹ 3ʹ A T C C G G T T A G G C C A A C C G G A T T G G C C T A Inverted Transposase gene Inverted repeat repeat (a) Insertion sequences, the simplest transposable elements in bacteria, contain a single gene that encodes transposase, which catalyzes movement within the genome. The inverted repeats are backward, upside-down versions of each other; only a portion is shown. The inverted repeat sequence varies from one type of insertion sequence to another. 3ʹ 5ʹ Figure 18.19a

100 Chapter 12 - DNA Technology and the Human Genome How can we use bacteria to manipulate DNA and protein? Transposable Elements (Transposons) 1. Also known as jumping genes Nobel Prize, Cold Spring Harbor

101 Chapter 12 - DNA Technology and the Human Genome How can we use bacteria to manipulate DNA and protein? Transposable Elements (Transposons) 1. Also known as jumping genes Transposon Insertion sequence Antibiotic resistance gene Insertion sequence 5ʹ 3ʹ 3ʹ 5ʹ Inverted repeats Transposase gene (b) Transposons contain one or more genes in addition to the transposase gene. In the transposon shown here, a gene for resistance to an antibiotic is located between twin insertion sequences. The gene for antibiotic resistance is carried along as part of the transposon when the transposon is inserted at a new site in the genome. Figure 18.19b

102 Chapter 12 - DNA Technology and the Human Genome How can we use bacteria to manipulate DNA and protein? Transposable Elements (Transposons) DNA-transposons vs Retrotransposons Almost 50% of the human genome is composed of retrotransposons

103 Chapter 12 - DNA Technology and the Human Genome How can we use bacteria to manipulate DNA and protein? Transposable Elements (Transposons) DNA transposon:

104 Chapter 12 - DNA Technology and the Human Genome How can we use bacteria to manipulate DNA and protein? Transposable Elements (Transposons) DNA-transposons Important in gene duplication during S phase of meiosis

105 Chapter 11 - The Control of Gene Expression NEW AIM: How are genes regulated (controlled) in prokaryotes? Bacteria, like all other organisms, respond to their environment by regulating gene expression and protein/enzyme activity (a) Regulation of enzyme activity Precursor (b) Regulation of enzyme production Feedback inhibition Enzyme 1 Gene 1 (a) Negative Feedback: You have already seen how the product of a biosynthesis pathway like the amino acid tryptophan (trp) can allosterically inhibit an enzyme in its production pathway thereby shutting down its own production (negative feedback). Enzyme 2 Enzyme 3 Gene 2 Gene 3 Regulation of gene expression (b) Regulating gene expression: Genes can also be turned on/off. Enzyme 4 Gene 4 Let s look at how bacteria regulate gene expression first in relation to lactose and then trptophan Figure 18.20a, b Tryptophan Enzyme 5 Gene 5

106 Chapter 18 - Genetics of Viruses and Bacteria Questions 1. Jumping genes are known as and always code for the enzyme known as. 2. An F+ cell is said to be fertile because it carries with it the. 3. SRP RNA is found where in the cell? 4. How many different aa-trna synthetases are there? 5. Amino acids are added to what end of the trna by aa-trna synthestase. 6. If the anticodon for a given trna is 3 -GCG-5, what letters would you look for on the genetic code chart to determine the amino acid attached to this trna?

107 Chapter 11 - The Control of Gene Expression AIM: How are genes regulated (controlled) in prokaryotes? In order to begin to understand this process, we will look at a set of three genes involved in lactose metabolism (the hydrolysis of lactose to ) Glucose and galactose called the Lactose (Lac) Operon

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109 Chapter 11 - The Control of Gene Expression AIM: How are genes regulated (controlled) in prokaryotes? Fig. 11.1B LacI LacZ LacY LacA Anatomy of an operon (only prokaryotes have operons) An operon typically contains a: 1. Promoter 2. Operator 3. A set of genes (3 in this specific case) A. LacZ B. LacY The terminator sequence C. LacA 4. What critical gene part is missing from this figure? The terminator sequence The regulatory gene (LacI) is found OUTSIDE of the operon.

110 Chapter 11 - The Control of Gene Expression AIM: How are genes regulated (controlled) in prokaryotes? Fig. 11.1B LacI LacZ LacY LacA The three gene products (can you guess what they might be?): 1. LacZ codes for β-galactosidase - The enzyme that hydrolyzes lactose to glucose and galactose 2. LacY codes for permease - A passive lactose transporter protein that sits in the membrane and allow lactose to diffuse into the cell. 3. LacA codes for transacetylase - Exact function not yet known

111 Chapter 11 - The Control of Gene Expression AIM: How are genes regulated (controlled) in prokaryotes? Fig. 11.1B QUESTION If lactose is present around the cell (perhaps it is one of the bacterium in your mouth and you just drank a glass of milk), should these genes be turned on or off? They should be ON since lactose is present and will need to be hydrolyzed so the glucose and galactose can be used to make ATP of for biosynthesis. Let s look at how this operon works to control expression of these three genes

112 Chapter 11 - The Control of Gene Expression AIM: How are genes regulated (controlled) in prokaryotes? 1. The regulatory gene codes for the repressor protein. A. What does repress mean? - To prevent B. What will this protein do then? - It will prevent the expression of the genes (turn them off) Fig. 11.1B - Any guess how it might do this?

113 Chapter 11 - The Control of Gene Expression AIM: How are genes regulated (controlled) in prokaryotes? 1. The regulatory gene codes for the repressor protein. C. It represses by binding to the Operator sequence and in doing so blocks the promoter sequence. Fig. 11.1B

114 Chapter 11 - The Control of Gene Expression AIM: How are genes regulated (controlled) in prokaryotes? Fig. 11.1B 1. The regulatory gene codes for the repressor protein. C. It represses by binding to the Operator sequence. -When it binds the operator, it will interfere with RNA polymerase binding to the promoter. The genes are off.

115 Chapter 11 - The Control of Gene Expression AIM: How are genes regulated (controlled) in prokaryotes? Fig. 11.1B ALL FOR ONE AND ONE FOR ALL Notice that all three genes are turned on/off together. Eukaryotes do not typically do this. They turn genes on/off individually.

116 Chapter 11 - The Control of Gene Expression AIM: How are genes regulated (controlled) in prokaryotes? Fig. 11.1B Q1. How do you suppose these genes will be turned ON when lactose is present? A1. Somehow the repressor needs to fall off. Q2. How can we get it to fall off? (HINT: you are changing its function) A2. You need to change its structure. Q3. How can we change the structure? A3. Bind something to it a ligand. Q4. What should the ligand be?

117 Chapter 11 - The Control of Gene Expression AIM: How are genes regulated (controlled) in prokaryotes? The ligand should be lactose itself since in the presence of lactose these genes should be turned ON. Fig. 11.1B

118 Chapter 11 - The Control of Gene Expression AIM: How are genes regulated (controlled) in prokaryotes? Activating the operon: 1. Lactose binds the repressor. 2. A conformational (shape) change occurs and the repressor falls off the operator. 3. RNA polymerase now binds to the promoter and begin transcription of all three genes in one long mrna. 4. Ribosomes translate the mrna into proteins.

119 Chapter 11 - The Control of Gene Expression AIM: How are genes regulated (controlled) in prokaryotes? Q1. What will happen when β-galactosidase breaks down most of the lactose? A1. Lactose will fall off the repressor and the repressor will once again bind to the operator and turn the genes off. Q2. Why not just leave these genes on all the time? A2. This would be a huge waste of resources ATP, amino acids, ribosomes, nucleotides, RNA polymerases and space.

120 Chapter 11 - The Control of Gene Expression AIM: How are genes regulated (controlled) in prokaryotes? To be more detailed about it A small amount of lactose is converted to allolactose by an enzyme in the cell. It is actually allolactose that is what we call the inducer, which simply means it inactivates the repressor (aka induces transcription).

121 Chapter 11 - The Control of Gene Expression AIM: How are genes regulated (controlled) in prokaryotes? Lac repressor protein Repressor bound to the operator sequence

122 Lac operon The video

123 Chapter 11 - The Control of Gene Expression AIM: How are genes regulated (controlled) in prokaryotes? In reality, the presence of lactose alone is not enough to induce the transcription of the lac gene why would this be logical? Because there could be other sugars in excess like glucose. Why waste ATP going after lactose if you are already overloaded. How does the bacterium sense the levels of glucose and translate this information to the genome you ask

124 Chapter 11 - The Control of Gene Expression AIM: How are genes regulated (controlled) in prokaryotes? When glucose is absent and lactose present, camp levels are high 1. camp is an allosteric activator of CAP (catabolite activator protein) 2. CAP will bind to the CAP-binding site on the promoter and recruit RNA polymerase resulting in the production of much mrna: Promoter DNA lacl lacz camp CAP-binding site Active CAP RNA polymerase can bind and transcribe Operator Inactive CAP Inactive lac repressor Figure 18.23a (a) Lactose present, glucose scarce (camp level high): abundant lac mrna synthesized. If glucose is scarce, the high level of camp activates CAP, and the lac operon produces large amounts of mrna for the lactose pathway.

125 Chapter 11 - The Control of Gene Expression AIM: How are genes regulated (controlled) in prokaryotes? When glucose is present with lactose, camp levels are low 1. CAP is inactive and RNA polymerase will not bind well to the promoter even if the repressor is not present. 2. Little mrna made DNA lacl Promoter lacz CAP-binding site Operator RNA polymerase can t bind Inactive CAP Inactive lac repressor Figure 18.23b (b) Lactose present, glucose present (camp level low): little lac mrna synthesized. When glucose is present, camp is scarce, and CAP is unable to stimulate transcription.

126 Chapter 11 - The Control of Gene Expression AIM: How are genes regulated (controlled) in prokaryotes? How exactly does glucose lower the levels of camp? 1. Obviously the activity of adenylyl cyclase needs to be lowered, but glucose does not interact directly with this enzyme Not something you should memorize, just understand Figure X. Control of adenylate cyclase via the phosphotransferase system. A. IIA, IIB, IIC, and HPr comprise the phosphotransferase system. When glucose is present, the phosphorylated forms of IIAGlc are low because glucose siphons off the phosphate. IIAGlc then interacts with and inhibits adenylate cyclase activity. B. In the absence of glucose, the phosphorylated forms of glucose-specific IIAGlc and IIBCGlc accumulate because they cannot pass the phosphate to substrate (there is no glucose). Adenylate cyclase functions in this situation to produce camp. The inset on the right shows the conversion of ATP to cyclic AMP by adenylate cyclase.

127 Chapter 11 - The Control of Gene Expression AIM: How are genes regulated (controlled) in prokaryotes? Tryptophan (Trp) operon - This operon contains fours genes whose protein products are responsible for synthesizing (making) the amino acid tryptophan.

128 Chapter 11 - The Control of Gene Expression AIM: How are genes regulated (controlled) in prokaryotes? Tryptophan (Trp) operon When would you want to turn these genes on? When tryptophan is NOT present, because that is when you need to make it when trp is present, it will bind to and activate the repressor:

129 Chapter 11 - The Control of Gene Expression AIM: How are genes regulated (controlled) in prokaryotes? Tryptophan (Trp) operon How does this compare to the lac operon?

130 Chapter 11 - The Control of Gene Expression AIM: How are genes regulated (controlled) in prokaryotes? Tryptophan (Trp) operon Inducible operon You can turn ON (induce) the operon by adding something (lactose in this case) Repressible operon You can turn OFF (repress)the operon by adding something (Tryptophan in this case)

131 Chapter 11 - The Control of Gene Expression AIM: How are genes regulated (controlled) in prokaryotes? Tryptophan (Trp) operon I do not recommend memorizing the difference. Think about is logically: 1. The repressor bind to the operator 2. When it is bound the genes are off 3. You need the lactose break down genes when lactose is present. 4. Therefore, when lactose binds to repressor, it should fall off operator 5. Likewise, when trp is present, the trp synthesis genes are unnecessary because you have it already 6. Therefore, Trp when Trp binds to the repressor, the repressor should bind the operator and shut the genes off.

132 Chapter 11 - The Control of Gene Expression AIM: How are genes regulated (controlled) in prokaryotes? Trp operon in detail Tryptophan (Trp) is a corepressor since it represses along with the repressor.

133 Chapter 11 - The Control of Gene Expression AIM: How are genes regulated (controlled) in prokaryotes? The trp repressor (with trp bound) binding to the operator sequence.

134 Chapter 11 - The Control of Gene Expression AIM: How are genes regulated (controlled) in prokaryotes? Both cases are examples of repression but there can also be activation by activator proteins as we shall see in the next slide.

135 Chapter 18 - Genetics of Viruses and Bacteria Questions 1. A functioning unit of genomic DNA containing a cluster of genes under the control of a single promoter. 2. The lac genes in E. coli are turned on when what two conditions are present in the cell? 3. The major difference between how the trp genes are regulated compared to how the lac genes are regulated. 4. When glucose concentrations are low within an E. coli cell the concentration of is causing the activation of, which is required for recruiting RNA pol. 5. What is Χ 2 analysis used to determine?