Chapter 5
C. Histone proteins have a lower-than-average number of lysines and arginines in their polypeptide chains. (T/F)
False. Histones have a higher number of lysines and arginines than most proteins. These amino acids are positively charged and help to increase the nonspecific affinity between the histones and the negatively charged phosphates in the DNA backbone.
A. Comparing the relative number of chromosome pairs is a good way to determine whether two species are closely related. (T/F)
False. There are several examples of closely related species that have a drastically different number of chromosome pairs. Two related species of deer—Chinese and Indian muntjac—have 23 and 3, respectively.
What portion of the nucleosome was destroyed/removed during this treatment and what function does it normally serve?
Nuclease treatment degrades linker DNA, which can be up to 80 bp in length. This region of DNA is typically bound to linker histones (H1), which are involved in higher-level packing of the chromatin.
Nucleosome formation compacts the DNA into approximately [one-third/one-hundredth/one-thousandth] of its original length.
Nucleosome formation compacts DNA into approximately (one-third) of its original length.
Describe the mechanism by which heterochromatin can spread, once it has been established in one region of the chromosome.
Once the initial H3 lysine 9 methylation is established on core histone octamers in one region, the modification attracts a specific set of proteins and other histone-methylating enzymes. These enzymes create the same modification on adjacent histone octamers, which continue to recruit more heterochromatin-specific proteins and enzymes, creating a wave of heterochromatin spreading along the chromosome.
What can you conclude about your purified remodeling complex from the results in lanes 5 and 6?
Sample 5 demonstrates that our purified complex is working. It must be moving the nucleosomes, providing access for EcoRI, such that it can now cleave at its restriction site, which was not possible in the absence of NICRC. Sample 6 shows that NICRC will function only if ATP is available as an energy source for the remodeling process.
Evidence suggests that the replication of DNA packaged into heterochromatin occurs later than the replication of other chromosomal DNA. What is the simplest possible explanation for this phenomenon?
The DNA double helix in heterochromatin may be so tightly packed and condensed that it is inaccessible to the proteins that bind replication origins, including the DNA replication machinery. It may take extra time to remodel the chromatin to make it more accessible to the proteins required to initiate and perform DNA replication.
What molecular components were identified after this treatment was complete?
The core nucleosome was revealed to contain two molecules of the histones H2A, H2B, H3, and H4, as well as a 147-base-pair (bp) fragment of DNA.
B. What are the spots representing longer lengths of DNA? Why is there a ladder of spots?
The ladder of bands representing longer lengths of DNA probably corresponds to stretches of DNA associated with increasing numbers of nucleosomes (1, 2, 3, 4, 5, and so on). In support of this proposal, adjacent bands differ in size by roughly 200 nucleotides, which is the length of DNA found in a nucleosome core particle plus neighboring linker DNA. This interpretation must mean that the M-nuclease digestion did not go to completion, because if all non-nucleosomal DNA were digested completely, the samples would contain only the 150-base-pair fragment.
D. What kinds of enzymes might have been involved in changing the chromatin structure between lanes 2 and 3?
The main candidates for enzymes that catalyzed the nucleosome alterations near Sweetie are chromatin-remodeling complexes and enzymes that covalently modify histone tails with methyl, acetyl, or phosphate groups.
A. How many centromeres are in each cell? What is the main function of the centromere?
There are 46 centromeres per cell, one on each chromosome. The centromeres have a key role in the distribution of chromosomes to daughter cells during mitosis.
B. How many telomeres are in each cell? What is their main function?
There are 92 telomeres per cell, two on each chromosome. Telomeres serve to protect the ends of chromosomes and to enable complete replication of the DNA of each chromosome all the way to its tips.
B. Chromosomes exist at different levels of condensation, depending on the stage of the cell cycle. (T/F)
True.
B.Linker histones help compact genomic DNA by influencing the path of the DNA after it has wrapped about the nucleosome core. (T/F)
True.
C. Eukaryotic chromosomes contain many different sites where DNA replication can be initiated. (t/F)
True.
Figure Q5-45 clearly depicts the nucleolus, a nuclear structure that looks like a large, dark region when stained. The other dark, speckled regions in this image are the locations of particularly compact chromosomal segments called ____________. (a) euchromatin. (b) heterochromatin. (c) nuclear pores. (d) nucleosomes.
(b)
Interphase chromosomes are about______ times less compact than mitotic chromosomes, but still are about______ times more compact than a DNA molecule in its extended form. (a) 10, 1000 (b) 20, 500 (c) 5, 2000 (d) 50, 200
(b)
Stepwise condensation of linear DNA happens in five different packing processes. Which of the following four processes has a direct requirement for histone H1? (a) formation of "beads-on-a-string" (b) formation of the 30-nm fiber (c) looping of the 30-nm fiber (d) packing of loops to form interphase chromosomes
(b)
A. Over how many miles would the total DNA from the average human stretch?
2 × 1014 m = 124,274,238,447 miles.
Define a gene.
A gene is a segment of DNA that stores the information required to specify the particular sequence found in a protein (or, in some cases, the sequence of a structural or catalytic RNA).
A nucleosome contains two molecules each of histones [H1 and H2A/H2A and H2B] as well as of histones H3 and H4.
A nucleosome contains two molecules each of histones (H2A and H2B) as well as of histones H3 and H4.
Consider two different species of yeast that have similar genome sizes. Is it likely that they contain the same number of genes? A similar number of chromosomes?
A similar genome size indicates relatively little about the number of genes and virtually nothing about the number of chromosomes. For example, the commonly studied yeasts Saccharomyces cerevisiae (Sc) and Schizosaccharomyces pombe (Sp) are separated by roughly 400 million years of evolution, and both have a genome of 14 million base pairs. Yet Sc has 6500 genes packaged into 16 chromosomes and Sp has 4800 genes in 3 chromosomes.
E. Do you think that gene expression of Sweetie is higher, lower, or the same in galactose compared to glucose? What about Salty?
As the chromatin seems to have been loosened near Sweetie, it seems likely that Sweetie gene expression is increased when cells are grown in galactose rather than glucose, whereas Salty gene expression is likely to be the same under the two conditions. Perhaps the Sweetie gene contains instructions for a protein that is required for cells to metabolize galactose but not glucose.
Methylation and acetylation are common changes made to histone H3, and the specific combination of these changes is sometimes referred to as the "histone code." Which of the following patterns will probably lead to gene silencing? (a) lysine 9 methylation (b) lysine 4 methylation and lysine 9 acetylation (c) lysine 14 acetylation (d) lysine 9 acetylation and lysine 14 acetylation
(a)
Mitotic chromosomes are _____ times more compact than a DNA molecule in its extended form. (a) 10,000 (b) 100,000 (c) 1000 (d) 100
(a)
Most eukaryotic cells only express 20-30% of the genes they possess. The formation of heterochromatin maintains the other genes in a transcriptionally silent (unexpressed) state. Which histone modification directs the formation of the most common type of heterochromatin? (a) H3 lysine 4 methylation (b) H3 lysine 9 methylation (c) H3 lysine 14 methylation (d) H3 lysine 27 methylation
(b)
The N-terminal tail of histone H3 can be extensively modified, and depending on the number, location, and combination of these modifications, these changes may promote the formation of heterochromatin. What is the result of heterochromatin formation? (a) increase in gene expression (b) gene silencing (c) recruitment of remodeling complexes (d) displacement of histone H1
(b)
The human genome is a diploid genome. However, when germ-line cells produce gametes, these specialized cells are haploid. What is the total number of chromosomes found in each of the gametes (egg or sperm) in your body? (a) 22 (b) 23 (c) 44 (d) 46
(b)
Which of the following is not a chemical modification commonly found on core histone N-terminal tails? (a) methylation (b) hydroxylation (c) phosphorylation (d) acetylation
(b)
Although the chromatin structure of interphase and mitotic chromosomes is very compact, DNA-binding proteins and protein complexes must be able to gain access to the DNA molecule. Chromatin-remodeling complexes provide this access by __________________. (a) recruiting other enzymes. (b) modifying the N-terminal tails of core histones. (c) using the energy of ATP hydrolysis to move nucleosomes. (d) denaturing the DNA by interfering with hydrogen-bonding between base pairs.
(c)
How do changes in histone modifications lead to changes in chromatin structure? (a) They directly lead to changes in the positions of the core histones. (b) They change the affinity between the histone octamer and the DNA. (c) They help recruit other proteins to the chromatin. (d) They cause the histone N-terminal tails to become hyperextended.
(c)
Nucleosomes are formed when DNA wraps _____ times around the histone octamer in a ______ coil. (a) 2.0, right-handed (b) 2.5, left-handed (c) 1.7, left-handed (d) 1.3, right-handed
(c)
The core histones are small, basic proteins that have a globular domain at the C-terminus and a long, extended conformation at the N-terminus. Which of the following is not true of the N-terminal "tail" of these histones? (a) It is subject to covalent modifications. (b) It extends out of the nucleosome core. (c) It binds to DNA in a sequence-specific manner. (d) It helps DNA pack tightly.
(c)
The process of sorting human chromosome pairs by size and morphology is called karyotyping. A modern method employed for karyotyping is called chromosome painting. How are individual chromosomes "painted"? (a) with a laser (b) using fluorescent antibodies (c) using fluorescent DNA molecules (d) using green fluorescent protein
(c)
What type of macromolecule helps package DNA in eukaryotic chromosomes? (a) lipids (b) carbohydrates (c) proteins (d) RNA
(c)
The human genome is divided into linear segments and packaged into structures called chromosomes. What is the total number of chromosomes found in each of the somatic cells in your body? (a) 22 (b) 23 (c) 44 (d) 46
(d)
The inactivation of one X chromosome is established by the directed spreading of heterochromatin. The silent state of this chromosome is _______________ in the subsequent cell divisions. (a) completed (b) switched (c) erased (d) maintained
(d)
The octameric histone core is composed of four different histone proteins, assembled in a stepwise manner. Once the core octamer has been formed, DNA wraps around it to form a nucleosome core particle. Which of the following histone proteins does not form part of the octameric core? (a) H4 (b) H2A (c) H3 (d) H1
(d)
Which of the following questions would not be answered by using karyotyping? (a) Is the individual genetically female or male? (b) Do any of the chromosomes contain pieces that belong to other chromosomes? (c) Does the individual have an extra chromosome? (d) Do any chromosomes contain point mutations?
(d)
Explain the differences between chromosome painting and the older, more traditional method of staining chromosomes being prepared for karyotyping. Highlight the way in which each method identifies chromosomes by the unique sequences they contain.
Chromosome painting relies on the specificity of DNA complementarity. Because unique sequences for each chromosome are known, short DNA molecules matching a set of these sites can be designed for each chromosome. Each set is labeled with a specific combination of fluorescent dyes and then allowed to hybridize (form base pairs) with the two homologous chromosomes that contain the unique sequences being targeted. Giemsa stain is a nonfluorescent dye that has a high affinity for DNA, and specifically accumulates in regions that are rich in A-T nucleotide pairs. This dye produces a pattern of dark and light bands, which differ for each chromosome on the basis of the distribution of AT-rich regions.
For each of the following sentences, fill in the blanks with the best word or phrase selected from the list below. Not all words or phrases will be used; each word or phrase should be used only once. Each chromosome is a single molecule of __________________ whose extraordinarily long length can be compacted by as much as __________________-fold during __________________ and tenfold more during __________________. This is accomplished by binding to __________________ that help package the DNA in an orderly manner so it can fit in the small space delimited by the __________________. The structure of the DNA-protein complex, called __________________, is highly __________________ over time. 10,000, chromosome, mitosis, 100, different ,nuclear envelope 1000, DNA, nucleolus cell cycle, dynamic proteins cell wall, interphase similar chromatin, lipids static
Each chromosome is a single molecule of (DNA) whose extraordinarily long length can be compacted by as much as (1000)-fold during (interphase) and tenfold more during (mitosis). This is accomplished by binding to (proteins) that help package the DNA in an orderly manner so it can fit in the small space delimited by the (nuclear envelope.). The structure of the DNA-protein complex, called (chromatin), is highly (dynamic) over time.
D. The telomere is a specialized DNA sequence where microtubules from the mitotic spindle attach to the chromosome so that duplicate copies move to opposite ends of the dividing cell. (T/F)
False. The telomere is a specialized DNA sequence, but not for the attachment of spindle microtubules. Telomeres form special caps that stabilize the ends of linear chromosomes.
C. Notice the faint spots and extensive smearing in lane 3, suggesting the DNA could be cut almost anywhere near the Sweetie gene after growth of the cells in galactose. This was not observed in the other lanes. What probably happened to the DNA to change the pattern between lanes 2 and 3?
Given the ability of M-nuclease to cut anywhere near Sweetie after growth in galactose, it seems that the DNA is no longer protected from digestion by binding to histones. Perhaps the wrapping of DNA within the nucleosomes has been loosened considerably. This change in the nucleosomes must be specific to the Sweetie gene, because it is not seen at the Salty gene or throughout the genome.
Specific regions of eukaryotic chromosomes contain sequence elements that are absolutely required for the proper transmission of genetic information from a mother cell to each daughter cell. Which of the following is not known to be one of these required elements in eukaryotes? (a) terminators of replication (b) origins of replication (c) telomeres (d) centromeres
(a)
The chromosomes we typically see in images are isolated from mitotic cells. These mitotic chromosomes are in the most highly condensed form. Interphase cells contain chromosomes that are less densely packed and __________________________. (a) occupy discrete territories in the nucleus. (b) share the same nuclear territory as their homolog. (c) are restricted to the nucleolus. (d) are completely tangled with other chromosomes.
(a)
The classic "beads-on-a-string" structure is the most decondensed chromatin structure possible and is produced experimentally. Which chromatin components are not retained when this structure is generated? (a) linker histones (b) linker DNA (c) nucleosome core particles (d) core histones
(a)
A nucleosome core particle contains a core of histone with DNA wrapped around it approximately [twice/three times/four times].
A nucleosome core particle contains a core of histone with DNA wrapped around it approximately (twice).
D. Interphase chromosomes represent a physical state of the chromatin with the highest order of packaging. (T/F)
False. When cells enter mitosis, the interphase chromosomes undergo at least one more level of packaging, which facilitates the segregation of sister chromatids.
Nucleosomes are aided in their formation by the high proportion of [acidic/basic/polar] amino acids in histone proteins.
Nucleosomes are aided in their formation by the high proportion of (basic) amino acids in histone proteins.
Which of the following best describes the mechanism by which chromatin-remodeling complexes "loosen" the DNA wrapped around the core histones? (a) They use energy derived from ATP hydrolysis to change the relative position of the DNA and the core histone octamer. (b) They chemically modify the DNA, changing the affinity between the histone octamer and the DNA. (c) They remove histone H1 from the linker DNA adjacent to the core histone octamer. (d) They chemically modify core histones to alter the affinity between the histone octamer and the DNA.
(a)
When there is a well-established segment of heterochromatin on an interphase chromosome, there is usually a special barrier sequence that prevents the heterochromatin from expanding along the entire chromosome. Gene A, which is normally expressed, has been moved by DNA recombination near an area of heterochromatin. None of the daughter cells produced after this recombination event express gene A, even though its DNA sequence is unchanged. What is the best way to describe what has happened to the function of gene A in these cells? (a) barrier destruction (b) heterochromatization (c) epigenetic inheritance (d) euchromatin depletion
(c)
A. The histone proteins that constitute the core nucleosome include tetramers of histones H2A, H2B, H3, and H4. (T/F)
False. When the core nucleosome is analyzed, it is revealed that there are H2A/H2B tetramers and H3/H4 tetramers in solution. Each of the tetramers has two subunits of the respective histone proteins.
How can nucleosome core particles be isolated from chromatin?
In a test tube, the nucleosome core particle can be released from chromatin by treatment with a nuclease that degrades the exposed, linker DNA, but not the DNA wrapped around the nucleosome core.
-For each of the following sentences, fill in the blanks with the best word or phrase selected from the list below. Not all words or phrases will be used; each word or phrase should be used only once. -In eukaryotic __________________, DNA is complexed with proteins to form __________________. The paternal and maternal copies of human Chromosome 1 are __________________, whereas the paternal copy of Chromosome 1 and the maternal copy of Chromosome 3 are __________________. Cytogeneticists can determine large-scale chromosomal abnormalities by looking at a patient's __________________. Fluorescent molecules can be used to paint a chromosome, by a technique that employs DNA __________________, and thereby to identify each chromosome by microscopy. bands extended kinetochore chromatin homologous nonhomologous chromosomes hybridization condensation karyotype
In eukaryotic (chromosomes), DNA is complexed with proteins to form (chromatin). The paternal and maternal copies of human Chromosome 1 are (homologous), whereas the paternal copy of Chromosome 1 and the maternal copy of Chromosome 3 are (nonhomologous). Cytogeneticists can determine large-scale chromosomal abnormalities by looking at a patient's (karyotype). Fluorescent molecules can be used to paint a chromosome, by a technique that employs DNA (hybridization), and thereby to identify each chromosome by microscopy.
For each of the following sentences, fill in the blanks with the best word or phrase selected from the list below. Not all words or phrases will be used; each word or phrase should be used only once. Interphase chromosomes contain both darkly staining __________________ and more lightly staining __________________. Genes that are being transcribed are thought to be packaged in a __________________ condensed type of euchromatin. Nucleosome core particles are separated from each other by stretches of __________________ DNA. A string of nucleosomes coils up with the help of __________________ to form the more compact structure of the __________________. A __________________ model describes the structure of the 30-nm fiber. The 30 nm chromatin fiber is further compacted by the formation of __________________ that emanate from a central __________________. 30-nm fiber heterochromatin linker active chromatin histone H1 loops axis histone H3 more beads-on-a-string histone H4 synaptic complex euchromatin less zigzag
Interphase chromosomes contain both darkly staining (heterochromatin) and more lightly staining (euchromatin). Genes that are being transcribed are thought to be packaged in a (less) condensed type of euchromatin. Nucleosome core particles are separated from each other by stretches of (linker) DNA. A string of nucleosomes coils up with the help of (histone H1) to form the more compact structure of the (30-nm fiber). A (zigzag model) describes the structure of the 30-nm fiber. The 30 nm chromatin fiber is further compacted by the formation of (loops) that emanate from a central (axis).
Nucleosomes are present in [prokaryotic/eukaryotic] chromosomes, but not in [prokaryotic/eukaryotic] chromosomes.
Nucleosomes are present in (eukaryotic )chromosomes, but not in (prokaryotic) chromosomes.
Explain the results in lanes 1-4 and why it is important to have this information before you begin to test your remodeling complex.
Sample 1 confirms the location of the EcoRI restriction site and shows what those fragments should look like when separated on the gel. The cleavage is the readout that will tell us whether the remodeling complex is working. Sample 2 demonstrates that DNA that is not assembled into nucleosomes can be cut into many small fragments by DNaseI. That is why we do not see discrete bands. Sample 3 demonstrates that when nucleosomes are assembled on the DNA, DNase I cuts only in one place, presumably in the linker region between two assembled nucleosomes. Sample 4 demonstrates that EcoRI cannot access its cleavage site when nucleosomes are assembled over it.
B. How many times would the total DNA from the average human wrap around the planet Earth at the Equator?
The Earth's circumference at the Equator is 24,902 miles. The length of DNA from the average human body could wrap around the Earth 4,990,532 times.
C. How many times would the total DNA from the average human stretch from Earth to the Sun and back?
The average distance from the Earth to the Sun is 93,000,000 miles. So, the round-trip distance is 186,000,000 miles. The length of DNA from the average human body could stretch from the Earth to the Sun and back 668 times.
D. How many times would the total DNA from the average human stretch from the Earth to Pluto and back?
The distance from the Earth to Pluto is, on average, about 39 × 93,000,000 miles. So, the round trip distance is 78 × 93,000,000 miles. The length of DNA from the average human body could stretch from the Earth to Pluto and back 17 times.
A. The lowest spot (as observed in lanes 2, 4, and 5) has a length of about 150 nucleotides. Can you propose what it is and how it arose?
The lowest spot represents DNA of a length similar to that of the segment of DNA found in a nucleosome core particle. Partial digestion with an enzyme such as M-nuclease causes breaks in the DNA backbone primarily within the linker DNA or other DNA segments not bound tightly to histones. Thus, this band probably comprises the DNA bound tightly to a single histone octamer and it arose by cutting the linker DNA outside a single nucleosome core particle.
C. How many replication origins are in each cell? What is their main function?
There are far more than 200 replication origins in a human cell, probably about 10,000. These DNA sequences direct the initiation of DNA synthesis needed to replicate chromosomes.
