The Genetics of Viruses and Bacteria Chapter 18 The Genetics of Viruses and Bacteria

Figure 18.1 T4 bacteriophage infecting an E. coli cell 0.5 m

Figure 18.2 Comparing the size of a virus, a bacterium, and an animal cell Animal cell nucleus

Figure 18.3 Infection by tobacco mosaic virus (TMV)

Figure 18.4 Viral structure 18  250 mm 70–90 nm (diameter) 80–200 nm (diameter) 80  225 nm 20 nm 50 nm (a) Tobacco mosaic virus (b) Adenoviruses (c) Influenza viruses (d) Bacteriophage T4 RNA Capsomere of capsid DNA Capsomere Glycoprotein Membranous envelope Capsid Head Tail fiber Tail sheath

Figure 18.5 A simplified viral reproductive cycle VIRUS Capsid proteins mRNA Viral DNA HOST CELL Entry into cell and uncoating of DNA Replication Transcription DNA Capsid Self-assembly of new virus particles and their exit from cell

Figure 18.6 The lytic cycle of phage T4, a virulent phage Attachment. The T4 phage uses its tail fibers to bind to specific receptor sites on the outer surface of an E. coli cell. Entry of phage DNA and degradation of host DNA. The sheath of the tail contracts, injecting the phage DNA into the cell and leaving an empty capsid outside. The cell’s DNA is hydrolyzed. Synthesis of viral genomes and proteins. The phage DNA directs production of phage proteins and copies of the phage genome by host enzymes, using components within the cell. Assembly. Three separate sets of proteins self-assemble to form phage heads, tails, and tail fibers. The phage genome is packaged inside the capsid as the head forms. Release. The phage directs production of an enzyme that damages the bacterial cell wall, allowing fluid to enter. The cell swells and finally bursts, releasing 100 to 200 phage particles. 1 2 4 3 5 Phage assembly Head Tails Tail fibers

Figure 18.7 The lytic and lysogenic cycles of phage , a temperate phage Many cell divisions produce a large population of bacteria infected with the prophage. The bacterium reproduces normally, copying the prophage and transmitting it to daughter cells. Phage DNA integrates into the bacterial chromosome, becoming a prophage. New phage DNA and proteins are synthesized and assembled into phages. Occasionally, a prophage exits the bacterial chromosome, initiating a lytic cycle. Certain factors determine whether The phage attaches to a host cell and injects its DNA. Phage DNA circularizes The cell lyses, releasing phages. Lytic cycle is induced Lysogenic cycle is entered or Prophage Bacterial chromosome Phage DNA

Table 18.1 Classes of Animal Viruses

Figure 18.8 The reproductive cycle of an enveloped RNA virus Capsid and viral genome enter cell 2 The viral genome (red) functions as a template for synthesis of complementary RNA strands (pink) by a viral enzyme. 3 New virus 8 RNA Capsid Envelope (with glycoproteins) Glycoproteins on the viral envelope bind to specific receptor molecules (not shown) on the host cell, promoting viral entry into the cell. 1 New copies of viral genome RNA are made using complementary RNA strands as templates. 4 Vesicles transport envelope glycoproteins to the plasma membrane. 6 A capsid assembles around each viral genome molecule. 7 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). 5 HOST CELL Viral genome (RNA) Template proteins Glyco- mRNA Copy of genome (RNA) ER

Figure 18.9 The structure of HIV, the retrovirus that causes AIDS Reverse transcriptase Viral envelope Capsid Glycoprotein RNA (two identical strands)

Figure 18.10 The reproductive cycle of HIV, a retrovirus Vesicles transport the glycoproteins from the ER to the cell’s plasma membrane. 7 The viral proteins include capsid proteins and reverse transcriptase (made in the cytosol) and envelope glycoproteins (made in the ER). 6 The double-stranded DNA is incorporated as a provirus into the cell’s DNA. 4 Proviral genes are transcribed into RNA molecules, which serve as genomes for the next viral generation and as mRNAs for translation into viral proteins. 5 Reverse transcriptase catalyzes the synthesis of a second DNA strand complementary to the first. 3 catalyzes the synthesis of a DNA strand complementary to the viral RNA. 2 New viruses bud off from the host cell. 9 Capsids are assembled around viral genomes and reverse transcriptase molecules. 8 mRNA RNA genome for the next viral generation Viral RNA RNA-DNA hybrid DNA Chromosomal DNA NUCLEUS Provirus HOST CELL Reverse transcriptase New HIV leaving a cell HIV entering a cell 0.25 µm HIV Membrane of white blood cell The virus fuses with the cell’s plasma membrane. The capsid proteins are removed, releasing the viral proteins and RNA. 1

Figure 18.11 SARS (severe acute respiratory syndrome), a recently emerging viral disease (a) Young ballet students in Hong Kong wear face masks to protect themselves from the virus causing SARS. (b) The SARS-causing agent is a coronavirus like this one (colorized TEM), so named for the “corona” of glycoprotein spikes protruding from the envelope.

Figure 18.12 Viral infection of plants

Figure 18.13 Model for how prions propagate Normal protein Original prion New prion Many prions

Figure 18.14 Replication of a bacterial chromosome Replication fork Origin of replication Termination of replication

Figure 18.15 Can a bacterial cell acquire genes from another bacterial cell? Only the samples from the mixed culture, contained cells that gave rise to colonies on minimal medium, which lacks amino acids. RESULTS EXPERIMENT Researchers had two mutant strains, one that could make arginine but not tryptophan (arg+ trp–) and one that could make tryptophan but not arginine (arg– trp+). Each mutant strain and a mixture of both strains were grown in a liquid medium containing all the required amino acids. Samples from each liquid culture were spread on plates containing a solution of glucose and inorganic salts (minimal medium), solidified with agar. Mutant strain arg+ trp– Mutant strain arg trp+ Mixture

Mixture No colonies (control) Colonies grew Mutant strain arg+ trp– Mutant strain arg– trp+ No colonies (control) Mixture Colonies grew Because only cells that can make both arginine and tryptophan (arg+ trp+ cells) can grow into colonies on minimal medium, the lack of colonies on the two control plates showed that no further mutations had occurred restoring this ability to cells of the mutant strains. Thus, each cell from the mixture that formed a colony on the minimal medium must have acquired one or more genes from a cell of the other strain by genetic recombination. CONCLUSION

Figure 18.16 Generalized transduction Phage DNA Donor cell A+ B+ Phage infects bacterial cell that has alleles A+ and B+ Host DNA (brown) is fragmented, and phage DNA and proteins are made. This is the donor cell. A bacterial DNA fragment (in this case a fragment with the A+ allele) may be packaged in a phage capsid. 1 2 3

Figure 18.16 Generalized transduction Phage DNA Donor cell Recipient cell A+ B+ B– A– Recombinant cell Crossing over Phage infects bacterial cell that has alleles A+ and B+ Host DNA (brown) is fragmented, and phage DNA and proteins are made. This is the donor cell. A bacterial DNA fragment (in this case a fragment with the A+ allele) may be packaged in a phage capsid. Phage with the A+ allele from the donor cell infects a recipient A–B– cell, and crossing over (recombination) between donor DNA (brown) and recipient DNA (green) occurs at two places (dotted lines). The genotype of the resulting recombinant cell (A+B–) differs from the genotypes of both the donor (A+B+) and the recipient (A–B–). 1 2 3 4 5

Figure 18.17 Bacterial conjugation Sex pilus 1 m

Figure 18.18 Conjugation and recombination in E. coli (layer 1) A cell carrying an F plasmid (an F+ cell) can form a mating bridge with an F– cell and transfer its F plasmid. A single strand of the F plasmid breaks at a specific point (tip of blue arrowhead) and begins to move into the recipient cell. As transfer continues, the donor plasmid rotates (red arrow). 2 DNA replication occurs in both donor and recipient cells, using the single parental strands of the F plasmid as templates to synthesize complementary strands. 3 The plasmid in the recipient cell circularizes. Transfer and replication result in a compete F plasmid in each cell. Thus, both cells are now F+. 4 F Plasmid Bacterial chromosome Bacterial chromosome F– cell F+ cell Hfr cell F factor The circular F plasmid in an F+ cell can be integrated into the circular chromosome by a single crossover event (dotted line). The resulting cell is called an Hfr cell (for High frequency of recombination). Since an Hfr cell has all the F-factor genes, it can form a mating bridge with an F– cell and transfer DNA. A single strand of the F factor breaks and begins to move through the bridge. DNA replication occurs in both donor and recipient cells, resulting in double-stranded DNA The location and orientation of the F factor in the donor chromosome determine the sequence of gene transfer during conjugation. In this example, the transfer sequence for four genes is A-B-C-D. 5 The mating bridge usually breaks well before the entire chromosome and the rest of the F factor are transferred. 6 Two crossovers can result in the exchange of similar (homologous) genes between the transferred chromosome fragment (brown) and the recipient cell’s chromosome (green). 7 The piece of DNA ending up outside the bacterial chromosome will eventually be degraded by the cell’s enzymes. The recipient cell now contains a new combination of genes but no F factor; it is a recombinant F– cell. 8 Temporary partial diploid Recombinant F– bacterium Conjugation and transfer of an F plasmid from an F+ donor to an F– recipient (a) Conjugation and transfer of part of the bacterial chromosome from an Hfr donor to an F– recipient, resulting in recombination (b) A+ B+ C+ D+ F– cell A– B– C– D– Mating bridge

Figure 18.18 Conjugation and recombination in E. coli (layer 2) A cell carrying an F plasmid (an F+ cell) can form a mating bridge with an F– cell and transfer its F plasmid. A single strand of the F plasmid breaks at a specific point (tip of blue arrowhead) and begins to move into the recipient cell. As transfer continues, the donor plasmid rotates (red arrow). 2 DNA replication occurs in both donor and recipient cells, using the single parental strands of the F plasmid as templates to synthesize complementary strands. 3 The plasmid in the recipient cell circularizes. Transfer and replication result in a compete F plasmid in each cell. Thus, both cells are now F+. 4 F Plasmid Bacterial chromosome Bacterial chromosome F– cell F+ cell Hfr cell F factor The circular F plasmid in an F+ cell can be integrated into the circular chromosome by a single crossover event (dotted line). The resulting cell is called an Hfr cell (for High frequency of recombination). Since an Hfr cell has all the F-factor genes, it can form a mating bridge with an F– cell and transfer DNA. A single strand of the F factor breaks and begins to move through the bridge. DNA replication occurs in both donor and recipient cells, resulting in double-stranded DNA The location and orientation of the F factor in the donor chromosome determine the sequence of gene transfer during conjugation. In this example, the transfer sequence for four genes is A-B-C-D. 5 The mating bridge usually breaks well before the entire chromosome and the rest of the F factor are transferred. 6 Two crossovers can result in the exchange of similar (homologous) genes between the transferred chromosome fragment (brown) and the recipient cell’s chromosome (green). 7 The piece of DNA ending up outside the bacterial chromosome will eventually be degraded by the cell’s enzymes. The recipient cell now contains a new combination of genes but no F factor; it is a recombinant F– cell. 8 Temporary partial diploid Recombinant F– bacterium Conjugation and transfer of an F plasmid from an F+ donor to an F– recipient (a) Conjugation and transfer of part of the bacterial chromosome from an Hfr donor to an F– recipient, resulting in recombination (b) A+ B+ C+ D+ F– cell A– B– C– D– Mating bridge

Figure 18.18 Conjugation and recombination in E. coli (layer 3) A cell carrying an F plasmid (an F+ cell) can form a mating bridge with an F– cell and transfer its F plasmid. A single strand of the F plasmid breaks at a specific point (tip of blue arrowhead) and begins to move into the recipient cell. As transfer continues, the donor plasmid rotates (red arrow). 2 DNA replication occurs in both donor and recipient cells, using the single parental strands of the F plasmid as templates to synthesize complementary strands. 3 The plasmid in the recipient cell circularizes. Transfer and replication result in a compete F plasmid in each cell. Thus, both cells are now F+. 4 F Plasmid Bacterial chromosome Bacterial chromosome F– cell F+ cell Hfr cell F factor The circular F plasmid in an F+ cell can be integrated into the circular chromosome by a single crossover event (dotted line). The resulting cell is called an Hfr cell (for High frequency of recombination). Since an Hfr cell has all the F-factor genes, it can form a mating bridge with an F– cell and transfer DNA. A single strand of the F factor breaks and begins to move through the bridge. DNA replication occurs in both donor and recipient cells, resulting in double-stranded DNA The location and orientation of the F factor in the donor chromosome determine the sequence of gene transfer during conjugation. In this example, the transfer sequence for four genes is A-B-C-D. 5 The mating bridge usually breaks well before the entire chromosome and the rest of the F factor are transferred. 6 Two crossovers can result in the exchange of similar (homologous) genes between the transferred chromosome fragment (brown) and the recipient cell’s chromosome (green). 7 The piece of DNA ending up outside the bacterial chromosome will eventually be degraded by the cell’s enzymes. The recipient cell now contains a new combination of genes but no F factor; it is a recombinant F– cell. 8 Temporary partial diploid Recombinant F– bacterium Conjugation and transfer of an F plasmid from an F+ donor to an F– recipient (a) Conjugation and transfer of part of the bacterial chromosome from an Hfr donor to an F– recipient, resulting in recombination (b) A+ B+ C+ D+ A– B– C– D– Mating bridge

Figure 18.18 Conjugation and recombination in E. coli (layer 4) A cell carrying an F plasmid (an F+ cell) can form a mating bridge with an F– cell and transfer its F plasmid. A single strand of the F plasmid breaks at a specific point (tip of blue arrowhead) and begins to move into the recipient cell. As transfer continues, the donor plasmid rotates (red arrow). 2 DNA replication occurs in both donor and recipient cells, using the single parental strands of the F plasmid as templates to synthesize complementary strands. 3 The plasmid in the recipient cell circularizes. Transfer and replication result in a compete F plasmid in each cell. Thus, both cells are now F+. 4 F Plasmid Bacterial chromosome Bacterial chromosome F– cell F+ cell Hfr cell F factor The circular F plasmid in an F+ cell can be integrated into the circular chromosome by a single crossover event (dotted line). The resulting cell is called an Hfr cell (for High frequency of recombination). Since an Hfr cell has all the F-factor genes, it can form a mating bridge with an F– cell and transfer DNA. A single strand of the F factor breaks and begins to move through the bridge. DNA replication occurs in both donor and recipient cells, resulting in double-stranded DNA The location and orientation of the F factor in the donor chromosome determine the sequence of gene transfer during conjugation. In this example, the transfer sequence for four genes is A-B-C-D. 5 The mating bridge usually breaks well before the entire chromosome and the rest of the F factor are transferred. 6 Two crossovers can result in the exchange of similar (homologous) genes between the transferred chromosome fragment (brown) and the recipient cell’s chromosome (green). 7 The piece of DNA ending up outside the bacterial chromosome will eventually be degraded by the cell’s enzymes. The recipient cell now contains a new combination of genes but no F factor; it is a recombinant F– cell. 8 Temporary partial diploid Recombinant F– bacterium Conjugation and transfer of an F plasmid from an F+ donor to an F– recipient (a) Conjugation and transfer of part of the bacterial chromosome from an Hfr donor to an F– recipient, resulting in recombination (b) A+ B+ C+ D+ A– B– C– D– Mating bridge

Figure 18.19 Transposable genetic elements in bacteria Insertion sequence 3 5 3 5 A T C C G G T… A C C G G A T… T A G G C C A … T G G C C T A … Inverted repeat Transposase gene Inverted 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. Transposon Insertion sequence Antibiotic resistance gene Insertion sequence 5 3 5 3 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 18.20 Regulation of a metabolic pathway (a) Regulation of enzyme activity Enzyme 1 Enzyme 2 Enzyme 3 Enzyme 4 Enzyme 5 Regulation of gene expression Feedback inhibition Tryptophan Precursor (b) Regulation of enzyme production Gene 2 Gene 1 Gene 3 Gene 4 Gene 5 –

Figure 18.21 The trp operon: regulated synthesis of repressible enzymes (a) Tryptophan absent, repressor inactive, operon on. RNA polymerase attaches to the DNA at the promoter and transcribes the operon’s genes. Genes of operon Inactive repressor Protein Operator Polypeptides that make up enzymes for tryptophan synthesis Promoter Regulatory gene RNA polymerase Start codon Stop codon trp operon 5 3 mRNA 5 trpD trpE trpC trpB trpA trpR DNA mRNA E D C B A

DNA mRNA Protein Tryptophan (corepressor) Active repressor No RNA made Tryptophan present, repressor active, operon off. As tryptophan accumulates, it inhibits its own production by activating the repressor protein. (b)

Figure 18.22 The lac operon: regulated synthesis of inducible enzymes DNA mRNA Protein Active repressor RNA polymerase No RNA made lacZ lacl Regulatory gene Operator Promoter Lactose absent, repressor active, operon off. The lac repressor is innately active, and in the absence of lactose it switches off the operon by binding to the operator. (a) 5 3

lacl mRNA 5' DNA mRNA Protein Allolactose (inducer) Inactive repressor lacz lacY lacA RNA polymerase Permease Transacetylase -Galactosidase 5 3 (b) Lactose present, repressor inactive, operon on. Allolactose, an isomer of lactose, derepresses the operon by inactivating the repressor. In this way, the enzymes for lactose utilization are induced. mRNA 5 lac operon

Figure 18.23 Positive control of the lac operon by catabolite activator protein (CAP) 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. (a) CAP-binding site Operator Promoter RNA polymerase can bind and transcribe Inactive CAP Active CAP cAMP DNA Inactive lac repressor lacl lacZ

lacl lacZ Promoter DNA CAP-binding site Operator RNA polymerase 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. Inactive lac repressor Inactive CAP DNA RNA polymerase can’t bind Operator lacl lacZ CAP-binding site Promoter

Unnumbered figure p. 358 Time A B Number of bacteria Number of viruses