CLASS: 1st hour (10:00-11:00)
DATE: 9.21.10
PROFESSOR: Pittler
Eukaryotic Chromosome Organization
Scribe: Ashley Russell
Proof: Abby Northcutt
Page 1 of 6
I.
Intro [S1]
a.
b.
c.
II. Chromosomes [S2]
a. Read Slide.
b. Heterchromatin is what we used to call the “junk DNA”
c. Human genome is thought to contain 35,000 genes and are coded by the euchromatic regions.
d.
III. No title [S3]
a. An actual representation of a giemza (sp?) stained chromosome
b. This is a metacentric chromosome with the centromere roughly in the middle
c. Each portion of the chromosome is called a chromatid; sister chromatids are joined at the centromere
d. By definition, short arm is called the “p” arm and the longer arm is called the “q” arm; the shorter arm will be
show facing up and long arm down. This is strictly convention
e. At the end are telomeres; without telomeres, the chromosomes would shorten with every round of replication;
telomeres allow the ends of chromosomes to be maintained.
IV. Multiple Levels of Packing are Required… [S4]
a. Start out with a short region of dsDNA (2nm from top to bottom)
b. beads on a string form of chromatin 11nm histone complexes
c. Condense into 30nm fiber (packed nucleosomes)
d. 300nm form condenses to the 700 nm form
e. Can condense a large amount of DNA into a very small portion.
f. Ultimately will be condensed into a portion of the chromosome.
g. Each DNA molecule has been packaged into a mitotic chromosome that is about 10,000 times shorter than
its extended length
h. Open it up and extend all DNA in all the cells could extend out to Jupiter and back multiple times
V. Basic Unit of Chromatin… [S5]
a. Used to think DNA just had its role in expression of genes; but it is dynamic
b. Basic unit of chromatin is the nucleosome
c. 200bp make up nucleosome
d. a nuclease can digest the “linker portions” of the beads on a string and you will still maintain the individual
nucleosomes
e. This nuclease process occurs naturally in cell death
f. When we are looking for DNA fragmentation, this is one of the tests that is done.
g. High concentration of salt will separate DNA from proteins
VI. The nucleosome consists of 146bp of DNA… [S6]
a. Nucleosome consists of 146 bp of DNA that wraps around
b. Octameric histone core is made of histone H2A, H2B, H3 and H4.
c. These are multimeric complexes of proteins that make that core.
VII. Histon H1 helps compact the nucleosome into a 30nm fiber [S7]
a. H1 is the linker histone; it holds the histone complex so it stays compact and tight
b.
c.
VIII. THE HISTONE TAILS ARE A CRITICAL DETERMINANT OF CHROMATIN STRUCTURE [S8]
a. Each one has a portion of the protein that extends out (the Histone tail)
b. They are quite visible in the DNA and can be modified
c. This is part of the process of genomic imprinting
d.
IX. SPECIFIC MODIFICATIONS ARE ASSOCIATED WITH SPECIFIC FUNCTIONS [S9]
a. Read table
b. It changes the appearance of the DNA and allows different TF to bind and promote opening of the chromatin
and subsequent gene expression
c. Can have same DNA in all cells, but only certain genes are expressed in certain cells. The differences are
the modifications in the histone tail regions.
CLASS: 1st hour (10:00-11:00)
Scribe: Ashley Russell
DATE: 9.21.10
Proof: Abby Northcutt
PROFESSOR: Pittler
Eukaryotic Chromosome Organization
Page 2 of 6
d. Don’t have to memorize the different types of things and what they might be able to do. Just know that basic
changes of the histone tails is what leads to genomic imprinting and regulation of the initial phases of
expression of a gene.
X. THE NORMAL KARYOTYPE [S10]
a. 22 autosome pairs
b. 1 XY
c. XX= female
d. XY= male
e. We each have a diploid genome (2 copies of each chromosome)
f. 2 hapoild gametes together, one from egg one from sperm = a diploid
g. Aneuploid  can have one extra or one fewer chromosome
h. Euploid true chromosome content
XI. KARYOTYPE ANALYSIS [S11]
a. Isolate DNA from an individual
b. immortalize with Epstein Barr Virus
c. Freeze cells in mitosis with Cholchicine
d. Chromosome is most opened up during metaphase – get the best view of the chromosome at this point
e. Stain and physically cut these chromosomes out and rearrange them so you have the chromosome set.
f. Y chromosomes are relatively short; this is a female because 2 X chromosomes
g. Banding pattern on chromosome pair is essentially the same
h. Chromosomes are separated based on size – from largest to smallest
i. Arbitrary classification based on the apparent length
XII. “HISTONE CODE” HYPOTHESISS12]
a. Read slide
b. Can have various types of post-transcription or post-replication modifications: acetylation, methylation,
phosphorylation, etc.
c. Coding may be heritable; different TF that lead to specific pattern; Can have very similar patterning within
families that you wouldn’t see otherwise.
d. If there is no modification, probably a silent gene
e. If there is extensive acetylation, probably an active gene.
f. Only one explanation for how you can have some genes turned on in one type of cell but not in another, even
though ALL DNA is in ALL cells.
XIII. EPIGENETICS [S13]
a. Read Slide.
b. In order for a gene to express, chromosome has to be opened up (transcriptionally active). Gene has to be
opened up so you can make RNA then protein
c. First, have to decide if gene is supposed to be opened up or not
XIV.
EPIGENETICS AND DISEASE [S14]
a. Can be involved in disease
b. read slide
c. Lyon’s hypothesis: Why don’t females have double the expression of their X chromosome? Because one of
their chromosomes is inactivated.
d. Same thing is seen in autosomes.
e. Unexpressed allele is called an autosome
XV. WILMS TUMOR (S15)
a. You get an over expression of both alleles; leads to cancer
b. Read slide.
XVI.
DNA METHYLATION (S16)
a. CpG islands  regions that are important for promoter activity.
b. Even if the chromosome opens, there are other mechanisms of gene silencing
XVII.
EPGENETICS AND DISEASE (S17)
CLASS: 1st hour (10:00-11:00)
DATE: 9.21.10
PROFESSOR: Pittler
Eukaryotic Chromosome Organization
a. Read slide
b. A number of things get turned on at once, and this is deleterious to the system.
Scribe: Ashley Russell
Proof: Abby Northcutt
Page 3 of 6
XVIII.
THE KARYOTYPE SHOWS THE CHROMOSOME COMPLEMENT OF A NORMAL______ (S18)
a. Female because two sex chromosomes of the same length.
b. You know that karyotype is normal because all of the autosomes and sex chromosomes show pairs that are
relatively matched
c. Must know the typical banding pattern in order to determine if it is normal or not
XIX.
LARGEST, METACENTRIC (S19)
a. metacentric  centromere in the center of the chromosome
b. acrocentric  The centromere is near the top so they fold over, very few in humans (more in mice), drew
representation on board
XX.
ANATOMY OF A CHROMOSOME (S20)
a. read slide
b. telocentric  centromeres at the end; no human chromosomes are telocentric
c. acrocentric  close to the tip
d. submetracentric  not so close to the end, but not the middle either
e. metacentric  centromere in the middle
XXI.
ANATOMY OF A CHROMOSOME (S21)
a. read slide
b. can have a p and q arm on every type of chromosome besides telocentric.
c. Sister chromatids are connected at the centromere. They are not necessarily identical, but are very close.
XXII. ANATOMY OF A CHROMOSOME (S22)
a. Read slide.
b. Large region at end of chromosome that is there to prevent shortening of chromosomes over time
XXIII.
XXIV.
XXV.
XXVI.
SUBTELOMERES (S23)
a. Subtelomeres have some similarities to protein coding genes
b. Telomere is made of repeats
c. Occur at both ends
d. Telomere and sub-telomere region.
ANATOMY OF A CHROMOSOME (S24)
a. Read slide
b. Spindle fibers help separate the sister chromatids during mitosis
c.
TABLE 13.1 (S25)
a. Moving towards the morbid?? anatomy
b. Read slide
c. Table lists several diseases that are known to be associated with different regions of the genome.
d. Diseases have been mapped to certain regions or particular genes
CHROMOSOMAL SHORTHAND (S26)
a. From that, can put together the anatomy of the human genome
b. Can know exact gene and gene defect that causes specific diseases
c. Bands are ID with letter and numbers. Numbers are based on the band pattern that shows up during
staining. Numbers start at centromere and go out.
d. For 10-15 years, linkage analysis was done, had markers every 10 megabases in the genome. Could find
out if a particular disease segregates with a particular marker
e. The more tightly it segregates, can limit the region to say that this region must contain the gene. Then, you
identify some candidate genes within that region and try to ID which particular gene causes the disorder.
f. Now, we can limit it much further; we know many of the genes and their functions. Can use PCR.
g. Could take a sample of blood, then use PCR and within a month tell you if you are likely to have a particular
disorder or not. Before, would have taken years to do previously.
h. Major banding can be separated into subregions (ie. 3q.26.1, 3q.26.2, 3q.26.3)
i. This representation is called an ideogram
j. Giemza staining is the most common type of staining used to ID bands on the chromosome.
k. Sucrose intolerance is at 3q.26.
CLASS: 1st hour (10:00-11:00)
DATE: 9.21.10
PROFESSOR: Pittler
Eukaryotic Chromosome Organization
Scribe: Ashley Russell
Proof: Abby Northcutt
Page 4 of 6
XXVII. CHROMOSOMES CARRY DIFFERENT GENES (S27)
a. Ideograms for several chromosomes and the diseases associated with them.
b. There are diseases associated with every chromosome.
XXVIII. VISUALIZING THE CHROMOSOMES (S28)
a. How do we look at the chromosome?
i. Obtain fetal or adult tissue
ii. Take cells, prepare them on a slide.
iii. Get rid of excess material with a wash
iv. Put a cover slip on and tap it to disperse the chromosomes
1. “good tappers make good spreads”
v. DNA dyes or probes
vi. Today, we use fluorescent markers with markers for a specific chromosome.
vii. Then, we compare them. Is there change in the banding pattern? Is there a change in the size of the
chromosome?
viii. if you perturb more than 1% of the genome will likely have changes in development (polydactily) and/or
mental retardation.
XXIX.
FISHing (S29)
a. “Now we can go FISHing…”
b. Fluorescence in situ Hybridization
c. now we have DNA probes that are complementary to certain base sequences (possible because we know the
whole genome).
XXX.
FISH: fluorescence in situ hybridization (S30)
a. We can make a probe to any region that we want. Can be unique to a specific chromosome.
XXXI.
CYTOGENETICS (S31)
a. Anything that can change in the chromosome; gross changes in the chromosome.
XXXII. MUTATION AT THE CHROMOSOME LEVEL (S32)
a. Read slide.
b. Down to the specific NT is smallest change you would see. Will not be looking for point mutations at the
same time you are looking for gross abnormalities, though.
c. Definition of “mutation” will not be a test question
d. More expression is not necessarily better, but its better than less.
e. Huge amount of spontaneous abortion; huge amount of proofreading goes on during development in the
body; numbers are significantly higher than would have imagined.
f. A small percentage of newborns will make it through with chromosomal aberrations.
g. If a child is not developing normally it will spontaneously abort
h. 0.2 percent have a change that is not harmful. If the break does not effect a gene, then there will be no
changes. Very often, though, a break point will be within a gene or it will affect the expression of a gene
disease
XXXIII. CHROMOSOMAL ABNORMALITIES (S33)
a. One of the most common abnormality  Down syndrome (trisomy 21)
b. Genetically, if you have a Y, you are going to be a male, and if they have at least one X and no Y, classified
as female.
XXXIV. CHROMOSOME ANOMALIES MAY CAUSE PHENOTYPE ABNORMALITIES (S34)
a. female with Downs syndrome and her karyotype.
b. She has 3 copies of chromosome 21
c. Looks like some other issues as well.
XXXV. EXTRA AUTOSOMES (S35)
a. A number of other diseases associated with an extra autosome
i. 13, 18 and 21 are the most frequently seen autosomes.
1. Have fewer genes
XXXVI. CHROMOSOMAL SHORTHAND (S36)
CLASS: 1st hour (10:00-11:00)
Scribe: Ashley Russell
DATE: 9.21.10
Proof: Abby Northcutt
PROFESSOR: Pittler
Eukaryotic Chromosome Organization
Page 5 of 6
a. read slide
b. a normal male would be: 46, XY
c. normal female would be: 46, XX
XXXVII. CHROMOSOMAL ABNORMALITIES (S37)
a. read slide
b. Monosomy  only one chromosome
c. Trisomy  example, down syndrome (one extra)
d. Deletion  deletion of a long segment (could be deletion of a base or a long segment)
e. Duplication  addition of more DNA
f. Reciprocal Translocation  not one sided (exchange of gene segments)
g. Nonreciprocal translocation  no information coming back
h. Iso chromosome  has identical arms; a defect in meiosis
Ring  2 ends join together to form a ring
XXXVIII.
POLYPLOIDY (S38)
a. 3 copies of every chromosome
b. This person fell into the 3% of stillbirths
c. Too much DNA, you have extra gene expression and changes in the control of gene expression
i. 3 copies is not compatible with life
d. 2 sperm in one egg
e. Haploid sperm fertilizes a diploid egg
XXXIX. ANEUPLOIDY (S39)
a. Will have changes that affect more than 1% of the genome; so most are spontaneously aborted.
b. Down syndrome can occur by a translocation involving chromosome 21 or a trisomy 21.
c.
XL. ANEUPLOIDY (40)
a. What is the basis for these abnormalities?
b. Non-disjunction
c. Read slide
XLI.
NNONDISJUNCTION CAUSES ANEUPLOIDY (S41)
a. If non-disjunction happens in the 2nd division, you are a little better off because you will have half normal
gametes.
b. Non-disjunction of the first meiosis will be more problematic; will have no normal gametes. More likely to
cause disease.
XLII.
TRISOMIES AND MONOSOMIES (S42)
a. Read slide.
b. The majority of trisomies and monosomies produce enviable embryos.
c. Survival rate for Down syndrome is quite good (better than others)
XLIII.
AUTOSOMAL ANEUPLOIDS (S43)
a. Most are lethal
b. Read slide for symptoms of Down Syndrome
c. Term mongoloid is not used anymore.
XLIV.
AUTOSOMAL ANEUPOLOIDS (S44)
a. Read slide
b. A lot of issues associated with this trisomy.
XLV.
AUTOSOMAL ANEUPOLOIDS (S45)
a. Read slide
XLVI. ODDLY CLENTCHED FIST (S46)
XLVII. AUTOSOMAL ANEUPOLOIDS (S47)
a. Read slide
b. As far as information that one might want to remember, I think memorizing the particulars of the specific
defects is not the most important to focus on. – Don’t need to know every single symptom, just the genetics
XLVIII. SEX CHROMOSOME ANEUPLOIDY (S48)
a. read the slide
CLASS: 1st hour (10:00-11:00)
Scribe: Ashley Russell
DATE: 9.21.10
Proof: Abby Northcutt
PROFESSOR: Pittler
Eukaryotic Chromosome Organization
Page 6 of 6
b. look back at slide 41 to aid in understanding.
XLIX. Causes of Chromosomal Abnormalities (S68)
a. Know the basics of abnormalities, what they are & what they mean
b. Don’t need to know details of diseases
c. Go back to earlier slides to know Inversion or anything else you don’t understand from this slide
i. Two types of inversion: Pericentric & _____ (Inversion is covered on two previous slides)
[End 49:42 mins]