1. GENOMES and VIRUSES Chapter 13

2. Genome
13.1 A genome is ALL of the genetic material of an organism transmitted from parents to offspring. Its sequence is the order of nitrogenous bases (A, T, C, G) along a DNA molecule
NOTE: Every person’s genome is unique…with the exception of identical twins!
1990 Human Genome Project started
By 2003 human and several other genomes completely sequenced
By definition, a genome is the genetic material transmitted from parents to offspring. Every person’s genome is unique (with the exception of identical twins).
In 1990, the Human Genome Project was started to sequence the human genome and some other model organisms used in research.
The Sanger method of DNA sequencing has become automated, and a machine is capable of sequencing billions of nucleotides per day. Large genomes that are to be sequenced are broken down into short fragments, and 10−50 repeats of each fragment are sequenced for accuracy.
What do you do with the fragments once you know their sequence?

3. Sequence Assembly
The short sequences are assembled by pairing their overlaps to generate the long, continuous sequence of nucleotides in the DNA molecule present in each chromosome.
This approach is called shotgun sequencing because the sequence fragments do not originate from a particular gene or region but from sites scattered randomly across the genome.
Sequences can be determined by breaking up genome into small fragments, sequencing
these fragments, and then putting them together at their overlaps.

4. Complications in Sequencing
Sequences that are repeated In
genome can make sequencing difficult.
There are a variety of types of repeated sequence in eukaryotic genomes.
Long repeated sequences may be several thousand nucleotides long and may be tandem (next to each other) or dispersed (spread throughout the genome). Long repeats are often longer than the automated sequencing machines will output, and the repeat may not be detected at all. If it is detected, it is difficult to know how many times it is repeated.
There are also short repeating sequences that can consist of two nucleotides like the AT repeat shown here. The difficulty sequencing this type of repeat is the AT can fold back upon itself to form a double-stranded structure in which A is paired with T (forming a more stable molecule). Such structures are not easily sequenced.

5. Personalized Medicine
Every individual’s genome is unique. So what is the significance of sequencing the human genome?
The sequence in the Human Genome Project is a composite from different individuals. While we do differ at millions of nucleotide sites, we do share the same genes and regulatory regions.
An individual’s genome can potentially indicate
what diseases he or she is susceptible to,
his or her drug sensitivities, and
the best treatment plans for his or her individual “blueprint.”
If every person’s genome is unique, what is the significance of sequencing the human genome?
The sequence provided by the Human Genome Project is a composite of sequences from different individuals. While individual human DNA sequences differ at millions of nucleotide sites, we do share the same genes and regulatory regions, organized the same way on our chromosomes. The differences between individuals account in part for our physical differences as well as our susceptibility to diseases or our response to medication.
Determining the differences is a step toward personalized medicine. Ev

6. Genome Annotation
13.2 Genome annotation is to identify genes and other functional elements.
A goal of biology is to identify all the component macromolecules in biological systems and to understand their individual functions as well as their interactions.
A genome sequence is merely a long list of A, T, C, and G’s that represent the order in which nucleotides occur. However, not all the DNA is transcribed into RNA and not all of the RNA that is transcribed is translated.
Genome sequencing is the first step in understanding the function of a DNA sequence.
Genomes contain many different types of sequence. There are protein-coding regions, noncoding regions, and regions that are transcribed into RNA but never translated into protein.
Genome annotation is the process by which researchers identify the various types of sequence present in a genome and where they are located.

7. Evolutionary Relationships
Closely related viruses have closely related hosts.
Often, annotations are vague and point to areas for future research. A common annotation in large genomes is “hypothetical protein,” which indicates an open reading frame that could possibly code for a protein, but at the current time, its function is unknown.
Analysis of the similarities and differences in protein-coding genes and other types of sequence in the genomes of different species is an area of study called comparative genomics. These studies help us understand how genes/genomes evolve. Sequences that are similar in different organisms are called conserved regions.
Here is an example showing the evolutionary relationship of lentiviruses, grouped according to their genome sequences. Closely related viruses have closely related hosts.

8. Annotated Genome of HIV
The annotated HIV genome indicates the functional elements of the virus.

9. Gene number is NOT a good predictor of biological complexity.
Gene Numbers
13.3 Gene numbers do not correlate with the complexity of an organism.
Gene number is NOT a good predictor of biological complexity.
Why? Human cells can do many more things with the genes they have: subtle gene regulation, different protein interactions, and gene splicing.
Gene number is not a good predictor of biological complexity. It may seem surprising to see the relatively low number of genes in humans when compared to the other species since humans are so much more complex in cell number and type and behavior.
One hypothesis is that human cells can do many more things with the genes they have, because of subtle gene regulation, different protein interactions, and gene splicing.

10. Genome Sizes
There is NO relationship between genome size and organismal complexity among eukaryotes.
There is no relationship between genome size and organismal complexity among eukaryotes.

11. Genome Sizes
The disconnect between genome size and complexity is called the C-value paradox.
Some genomes have alot of repetitive DNA
Some genomes have a lot of transposable elements
Genomes differ in amount of coding/noncoding DNA
The range of genome size is huge, even among similar organisms. The largest eukaryotic genome exceeds the size of the smallest by 500,000, and both the smallest and the largest are found in the protozoa.
Clearly, there is no relationship between the size of the genome and the complexity of the organism.
The disconnect between genome size and complexity is called the C-value paradox. C-value is the amount of DNA in a reproductive cell, and the “paradox” is the apparent contradiction between the genome size and organismal complexity.

12. Large vs. Small Genomes, Why?
Polyploidy…having more than 2 sets of chromosomes in the genome
Particularly relevant in plants
The amount of non-coding DNA varies
Highly repetitive DNA
> 100,000 copies/genome
Moderately repetitive DNA
100 – 10,000 copies/genome
Why are there such large differences in genome size among species?

13. Polyploidy
In eukaryotes, large genomes can differ from small ones for a number of reasons—one is polyploidy. Polyploidy is having more than two sets of chromosomes in the genome. Humans have two sets of 23 chromosomes, but some plants can have six or seven sets. One species of fern has 84 copies of 15 chromosomes, making 1260 chromosomes altogether.
Because of crossing between related species, this plant contains a full set of chromosomes from each parent.
In eukaryotes, large genomes can differ from small ones due to polyploidy…….having more than 2 sets of chromosomes in the genome. Humans have 2 sets of 23 chromosomes. But some plants can have 6 or 7 sets.

14. Sequence Composition of the Human Genome
Only 2.5% of the human genome
actually codes for protein
About half of human genome consists
of repetitive DNA and transposable
elements
Transposable elements are called
“selfish DNA”
DNA that replicates and inserts itself
into new positions in genome
Makes up about 45% of DNA in human genome.
Different species can have vastly different quantities of highly repetitive and moderately repetitive DNA. Since these types of sequence are almost exclusively noncoding DNA, it is the differing amounts of these noncoding sequences that in large part account for the C-value paradox.
Among the highly repetitive sequences of the human genome is the alpha satellite DNA, which consists of tandem repeats of a 171-bp sequence repeated near the centromere an average of 18,000 times. This satellite DNA is essential for attachment of spindle fibers to the centromeres during cell division.
The human genome also contains several types of sequences called transposable elements. This DNA can replicate and insert itself into new positions in the genome. Transposable elements make up about 45% of the DNA in the human genome.
One class consists of DNA transposable elements (DNA TEs) that replicate and transpose via DNA replication and repair.
The other class transpose by means of an RNA intermediate. Among these are long terminal repeats (LTR), long interspersed nuclear elements (LINE), and short interspersed nuclear elements (SINE).

15. Bacterial Genome Organization
13.4 The orderly packaging of DNA allows it to carry out its functions and fit inside cell.
Bacteria have 1 circular chromosome.
It twists on itself to form supercoils, which are anchored by proteins
Bacterial genomes are circular, and the DNA double helix is underwound. Underwinding is caused by the enzyme, topoisomerase II, that breaks the double helix, rotates the ends, and then seals the break.
Underwinding creates strain on the DNA molecule, which is relieved by the formation of supercoils. Supercoiling allows all the base pairs to form, even though the molecule is underwound.

16. Eukaryotic Genome Organization
Eukaryotic cells package
their DNA as 1 molecule/linear
chromosome
6 levels of chromosome
packing:
DNA duplex (2 nm)
Nucleosome fiber(10 nm)
Beads on a string
DNA with histone
proteins
3. 30 nm chromatin fiber
4. Coiled chromatin fiber
5. Coiled coil
6. Metaphase chromosome
DNA in the nucleus of eukaryotes is packaged differently from that of bacteria.
2. Eukaryotic DNA is first wrapped around a group of histone proteins called a nucleosome. Each nucleosome consists of two molecules, and each molecule consists of four different histone proteins. The histone proteins are rich in the amino acids lysine and arginine, whose positive charges neutralize the negative charges of the phosphates along the backbone of each DNA strand. This level of packaging is often referred to as “beads on a string.”
3. The nucleosomes and associated DNA are then coiled to form a structure called the 30-nm chromatin fiber. This coiling constitutes chromosome condensation.

17. Histones/Histones Removed
When histones are chemically removed, the DNA spreads out in loops around a supporting protein structure called the chromosome scaffold. Each loop of relaxed DNA is 30−90 kb long and anchored to the scaffold at its base.
Despite similarities between bacterial genome organization and that of eukaryotes, they evolved independently and make use of different types of proteins to bind the DNA and to form the folded structure of DNA and protein.

18. Genome Organization of Mitochondria and Chloroplasts
The genomes of mitochondria and chloroplasts are organized into nucleoids. These organelles are thought to have once been free-living bacterial cells that were engulfed by primitive eukaryotic cells. Sometime over the course of evolution, most of their DNA was transferred to the DNA in the nucleus, and the structure of their nucleoids now differ from those of free-living bacteria, and from each other.
Mitochondria and chloroplasts have their own genomes. These organelles were once free-living bacterial cells that were engulfed by primitive eukaryotic cells.
Called Endosymbiotic Theorem.

19. Do not independently fulfill characteristics of life
Viruses
13.5 Viruses have diverse genomes, but all require a host cell to replicate.
What is a virus?
Virion: Infectious, acellular particle that is virulent (able to establish infection in a host)
Obligate intracellular parasite of bacteria, protozoa, fungi, plants, animals
Do not independently fulfill characteristics of life
Not on Tree of Life
Components of a virus….consists of only 2 pieces:
Nucleic acid (either ss or ds DNA or ss or ds RNA, but not both)
Capsid (protein coat)
Sometimes a lipid envelope…..helps viruses infect their host: fools the immune system
The first genome sequenced was a viral genome. Viruses causes diseases such as influenza (“flu”), polio, and HIV. In some cases, they cause cancer. In fact, the term “virus” comes from the Latin for “poison.”
Viruses play other roles as well.
Some viruses transfer genetic material from one cell to another, in a process called horizontal gene transfer. Horizontal gene transfer has played a major role in the evolution of bacterial and archaeal genomes, as well as in the origin and spread of antibiotic-resistance genes. Molecular biologists have learned to make use of this ability of viruses to deliver genes into cells.
Viruses play vital roles in ecosystems: Scientists estimate that 1 to 100 million viruses can be found in a teaspoon of seawater. They infect and limit the number of bacteria and other microorganisms in soil and water.
A virus consists simply of nucleic acid, a protein coat called a capsid, and sometimes a lipid envelope. Viruses can reproduce only by infecting living cells and subverting cellular metabolism and protein synthesis to produce more viruses. Most viruses are tiny, some hardly larger than a eukaryotic ribosome, 25–30 nm in diameter.

20. Virus Structure Bacteriophage Tobacco mosaic virus Adenovirus
Viruses show a wide variety of shapes and sizes.
Left: The T4 virus infects cells of the bacterium Escherichia coli. Viruses that infect bacterial cells are often called bacteriophages, which literally means “bacteria eaters.” The T4 bacteriophage has a complex structure that includes
a head composed of protein surrounding a molecule of double-stranded DNA
a tail and tail fibers
In infecting a host cell, the T4 tail fibers attach to the surface, and the DNA and some proteins are injected into the cell through the tail.
Center: The tobacco mosaic virus infects plants. It has a helical shape formed by the arrangement of protein subunits entwined with a molecule of single-stranded RNA.
Tobacco mosaic virus was the first virus to be discovered: experiments showed that the infectious agent causing brown spots and discoloration of tobacco leaves was so small that it could pass through the pores of filters that could trap even the smallest bacterial cells.
Right: The adenovirus is a common cause of upper respiratory infections in humans. Many viruses have an approximately spherical shape formed from polygons of protein subunits that come together to form a polyhedral capsid. Among the most common polyhedral shapes is an icosahedron, which has 20 identical triangular faces.
Many viruses that infect eukaryotic cells, such as adenovirus, are surrounded by an envelope composed of a lipid bilayer with embedded proteins and glycoproteins that recognize and attach to host cell receptors.
Viruses have diverse shapes and sizes.

21. VIRAL STRUCTURES

22. VIRAL DIVERSITY

23. Tobacco Mosaic Virus
All known cells and organisms are susceptible to viral infection. A cell in which viral reproduction occurs is called a host cell. Some viruses kill the host cell; others do not.
A given virus can infect only certain species or types of cell.
At one extreme, a virus can infect just a single species. Smallpox infects only humans. In this case, we say that the virus has a narrow host range.
Other viruses have a broad host range.
Rabies infects many different types of mammal, including squirrels, dogs, and humans.
Tobacco mosaic virus infects more than 100 different species of plants.
Plant viruses cannot infect bacteria or animals, and viruses that infect bacteria cannot infect plants or animals.
Host specificity relates to the way that viruses gain entry into cells.
Proteins on the surface of the capsid or envelope (if present) bind to proteins on the surface of host cells. The presence of the host protein on the cell surface determines which host cells a virus can infect.
Following attachment, the virus gains entry into the cell, where it can replicate by using the host cell machinery to make new viruses. In some cases, the viral genome integrates into the host cell genome.
All known cells are susceptible to viral infection: bacteria, archae, and eukaryotes.

24. VIRAL REPLICATION
Viruses are obligate intracellular parasites: they can only express their genes and reproduce only within a living cell
They cannot reproduce independently
Viral N.A. does not code for cellular proteins, but for the protein needed to make more viruses
They lack enzymes/other molecules needed to express their genes and reproduce
May cause diseases within host or kill host
Viral DNA - viral RNA - viral protein
Problem: host stops synthesizing their own molecules; the virus hijacks the host

25. HOST RANGE Each virus has a specific host range
Host range: spectrum of host cells that a virus can infect
Cell has to have a specific receptor on its plasma membrane surface for viral attachment
May be one host species or many
HIV (only humans) versus rabies (many animals)
May be one tissue or many within a host
Hepatitis (liver); HIV (specific WBCs); Cold viruses (lining of respiratory tract)

26. 2 REPRODUCTIVE CYCLES Lytic Cycle:
A viral replication cycle that results in the lysis/death of host cell
Releases new phage particles
Host either gets better or dies
Lysogenic Cycle:
A viral replication cycle that involves the incorporation of viral genome into host cell genome without killing host; “sleeping virus”
Viruses remain inactive in host cells/latent until initiate a lytic cycle (recurrence)
Stay with host forever; repeatedly cause disease when host weakened
Herpes, HIV, chicken pox/shingles

27. LYTIC VS. LYSOGENIC CYCLES

28. RETROVIRUSES Many RNA viruses are retroviruses….”backward” viruses
ssRNA viruses that make a copy of DNA
Use reverse transcriptase enzyme to transcribe DNA from viral RNA
HIV, the virus that causes AIDs, is a retrovirus
Of all viruses, HIV has highest mutation rate….makes many mistakes going backwards from RNA to DNA

29. ANIMAL VIRUSES
Classification of animal viruses is based on nature of the genome
Ss or ds DNA or RNA
Ss RNA is further classified as to how the RNA functions in host cell
Many have outer membraneous envelopes
HIV, influenza, herpes, SARs, West Nile, Ebola, etc
Replicate only inside host animal or human cells….damage or kill cells
Indirectly responsible for disease symptoms