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Introduction to Genetics

Historical landmarks in Genetics

1865

Mendel - published the work on inheritance but was overlooked for many years until 1900

1871

Fredrich Miescher - extracted a substance he called "nuclein" composed of protein and nucleic acid from the cell nuclei

1895

Edmund Wilson - suggested that inheritance might result from the transmission of chemical compounds - proposed nucleic acid or protein as the candidateThe lack of knowledge of the nucleic acids and their versatility, combined with the advances in protein chemistry, led to the belief that "protein" is the right candidate

1900

Landsteiner discover ABO blood group

1902

Garrod described alkaptonuria as the first “inborn error of metabolism”

1909

Johannsen coined the term “gene” as unit of heredity

1928

Fredrick Griffith - reported the phenomenon which came to be known as TRANSFORMATION - transformed non-virulent Streptococcus pneumonia to a virulent form by cell-free materials from virulent bacteria

1930

Phoebus A. Levene - indicated nucleic acids contained four nitrogenous bases - thymine, cytosine, guanine and adenine (RNA contained uracil, but not thymine)

1944

Avery, McLeod and McCarthy - demonstrated that the transforming factor in bacterial transformation of non-virulent to virulent form was DNA

1952

Hershey-Chase - identified DNA (rather than protein) as the genetic material of viruses, further evidence that DNA was the molecule responsible for inheritanceExperiment with E.Coliand bacterial chromosome plus phage and phage chromosome1. Phage attaches to E. Coli and injects chromosme2. Bacterial Chromosome break down and the phage chromosome replicates3. Expression of Phage genes produces Phage structural component4. Progeny Phage particles assemble5. Bacterial wall lyses releasing progeny phages.

1956

the correct number of human chromosomes were specified – led to the discovery in 1959 that Down syndrome is caused by an extra chromosome 21

1972

Paul Berg - and co-workers create the first recombinant DNA molecule

1977

Allan Maxam and Walter Gilbert (Harvard) and Frederick Sanger (U.K. Medical research Council) independently develop methods for sequencing DNA

1990 - 2003

The Human Genome Project

was a molecular genetics project thatbegan in 1990 and was projected to take fifteen years to complete

the project finished in April 2003, taking only thirteen years (50th years anniversary from DNA double helix structure by Watson & Crick)

The project was started by the National Institutes of Health and the U.S. Department of Energy in an effort to reach six set goals:

Identifying all genes in human DNA

(initial estimate were approximately 100,000 genes which was reduced as research progressed)– only 25,000 genes have true known functions.

Determining sequences of chemical based pairs in human DNA

Storing all found information into databases

Improving the tools used for data analysis

Transferring technologies to private sectors

Addressing the ethical, legal, and social issues (ELSI) that may arise from the project

The Study of Genetics

Genetics

study of the mechanism by which “hereditary” information is passed from generation to generation

is concerned with diversity, replication, mutation & translation of information in the genes of all living organisms

Classification of Genetics

Transmission Genetics

G. J. Mendel

July 20 1822 to Jan 6 1884 - FATHER OF GENETICS

Experiments on Plant Hybridization – presented in 1865

Procedure

Homozygous Dominant and Recessive Parental Generation (P) crossed to make F1 progeny which was all dominant phenotype

e.g. Yellow pea (YY) x Green (yy) = 100% Yy, yellow progeny

F1 was crossed with self to reveal ¾ dominant and ¼ recessive phenotypes in the F2.

e.g. Yy yellow x Yy yellow = ¼ YY yellow, ½ Yy yellow, and ¼ yy green

F1 crossed with P1 homozygous recessive to reveal ½ dominant and ½ recessive phenotypes in the F2

e.g. Yy yellow x yy green = ½ Yy yellow, ½ yy green

Mendelian Conclusions

The inheritance of each trait is determined by "units" or "factors" (now called genes) that are passed on to descendents unchanged

An individual inherits one such unit from each parent for each trait

A trait may not show up in an individual but can still be passed on to the next generation

Significance

Demonstrated the transmission of genetic traits from one generation to the next

Encompasses the basic heredity and how traits are passed from one generation to another

Originally formulated for cultivated peas, Mendelian laws can also be applied to determine the pattern of transmission of inherited human diseases

Mendelian Laws

The principle of segregation

two alleles of a locus on homologous chromosomes separate when gametes are formed

e.g. R, round, r, wrinkled

The principle of independent assortment

when two alleles separate, their separation is independent of the separation of alleles at other loci

Experiment: Do alleles coding different traits separate independently Methods P generation Round Yellow Seeds (RRYY) x wrinkled Green seeds (rryy) F1 – Round, Yellow Seeds (RrYy) crossed with self F2 – 9 round yellow: 3 round green: 3 wrinkled yellow: 1 wrinkled green; Conclusion: the allele encoding colour separated independently from the allele encoding seed shape, producing a 9:3:3:1 phenotypic ratio in the F2 progeny;

The Principle of Incomplete Dominance

when the heterozygote has a phenotype intermediate between the phenotypes of the two homozygotes

e.g. Fruit Colour in Eggplanta) P generation: PP purple x pp whiteb) F1: Pp violet crossed with selfc) F2: 1 PP purple: 2 Pp violet: 1: pp whiteConclusion: genotypic ratios and phenotypic ratios remain the same.Fruit colour in eggplant is inherited as an incompletely dominant characteristic

Mendel’s work is still recognized as the foundation of modern genetics

Population Genetics

Charles Darwin

Feb 12 1809 to Apr 19 1882

Involved with species variations and survival risks

Explores the genetic composition of individual members of the same species(population) and how that composition changes over time and geographic space

The voyage of the beagle led to the origin of species

Theory of Evolution

Came up with Theory of Evolution which rests on 3 principles:

Principle of Variation

Among individuals within any population, there is a variation in morphology, physiology and behavior

Principle of Heredity

Offspring resemble their parents more than they resemble unrelated individuals

Principle of Selection

Some forms are more successful at surviving and reproducing than other forms in a given environment

Molecular Genetics

Watson & Crick

1953-62

Established the molecular structure of the "gene" encoded by DNA ,

the basic chemical units of hereditary

Concerns the chemical nature of the gene itself: how genetic information is encoded, replicated and expressed

Types of Genetics applicable to Humans

Human Genetics:

Study of variation & heredity in human beings (refers to normal hereditary patterns).

Medical Genetics:

Deals with the subset of human genetic variation that is of significance in the practice of medicine & in medical research (refers to abnormal hereditary patterns/variations that lead to pathological conditions).

Major areas of Specialization within Human and Medical Genetics

Cytogenetics

Study of chromosomes, their structure & inheritance

Definitions

Clinical Cytogenetics

the study of the chromosomes and their abnormalities

Karyotype

test to identify and evaluate chromosomes in different cell types for their

Number

Size

Shape

Chromosomes morphology

p=petitq=queue

Centromeres

movement during cell division

Divides the chromosome into

short (p=petit) arms

long (q=queue) arms

Telomeres

tip of each chromosome

seal chromosomes and retain chromosome integrity

telomere consists of tandem repeats TTAGGG

maintained by enzyme – telomerase

reduction in telomerase and decrease in number repeats important in aging and cell death

Chromosomes nomenclature

International System for Human Cytogenetic Nomenclature (ISCN) – nomenclature reports

Banding

Basic terminology for banded chromosomes - Paris 1971

-specific chromosome bands

correct identification 
of individual chromosomes

detection of deletions, 
duplications

Example - Idiogram

Chromosomes classification

Metacentric

Submetacentric

Acrocentric

Chart of Chromosome Nomenclature

Chromosomal abnormalities

Chromosomal disorders

Occur in ~1 of every 150 live births

Observed in

~50% of spontaneous first-trimester abortions

20% of second trimester spontaneous abortions

2% of pregnancies of women older than 35 years

Clinical indications for cytogenetic test

Abnormalities of early growth and development

Multiple congenital malformations

Developmental delays

Failure to thrive

Mental retardation

Ambiguous genitalia

Stillbirth and neonatal deaths

Important for genetic counseling and prenatal diagnosis in future pregnancies

Fertility problems

Couples with history of infertility

Couples with recurrent miscarriages

Women presenting amenorrhea

Advanced age of a pregnant woman

Increased risk of chromosomal aberrations in women older than 35 years

Should be offered as a prenatal test

Family history

Chromosome abnormality in a first-degree relative

Cancer

Chromosomal aberrations in almost all cancers

Useful for diagnosis or prognosis

Cell types used for chromosomal analysis

Sources include:

Peripheral blood ( T lymphocytes)

Amniotic fluid

Chorionic villus

Fibroblast cultures

Bone marrow

Solid tumors

Cytogenetic analysis

Performed in cells capable of growth and rapid division in culture (e.g. T lymphocytes)

Subtopic

Molecular & Biochemical genetics

Study of the structure & function of individual genes

Genomics

Study of the genome, its organization & functions

Population genetics

Study of genetic variation in human population & the factors that determine allele frequencies

Developmental genetics

Study of the genetic control of development

Clinical genetics

Application of genetics to diagnosis & patient care

Pharmacogenetics

Study of genetic basis for variation in drug response

Nutrigenomics

Study of molecular relationships between nutrition and the response of genes

Organization of the Human Genome

Consists of large amounts of the chemical deoxyribonucleic acid (DNA) – contains the genetic information needed for all aspects of a functional human organism what are dna ratios ? 1 chromosome per mitoch? and 23 pairs per nucleus?

Nuclear Genome

Features

46 chromosomes:

22 autosomes

x and Y sex chromosomes

2% of the nuclear genome – coding DNA

~27,000 protein-encoding genes (DNA to mRNA to protein)

~ 3000 RNA-encoding genes (DNA to RNA; no translation to protein)

98% non-coding DNA

Karyotype

Chromosomes complement

Specific number and morphology for each species

Contain the genes aligned at specific position or locus

are studied in Cytogenetics for their clinical relevance with regards to

-Clinical Diagnosis

-Gene Mapping and Identification

-Cancer Cytogenetics

-Prenatal Diagnosis

Human Chromosomes

Chromosomes

found in nucleus and nucleolus

Structure

Human chromosomes showing the centromeres and well-defined chromatids

Active or inactive centromeres surrounded by flanking heterochromatin

Each chromosome consists of a single, continuous DNA double helix

– 46 DNA molecules = 6 billion nucleotides

Minor and Major Grooves along length

2 nm in diameter

1 helical turn is 3.4nm

3 Forms

A-DNA

B-DNA

This form is the one usually found under physiological conditions

Z-DNA

Chromatin

Genomic DNA - complexed with chromosomal proteins (histones and nonhistones)

distributed throughout the nucleus, relatively homogenous under the microscope

during cell division – chromatin condenses –visible microscopically as discrete structures

Chromatin Packing

Chromosomes pass through stages of condensation and decondensation into a DNA mitotic chromosome that is 10,000x shorter than its extended length.

Stages of Condensation

Double Helix

2nm in width

Nucleosome Fibre

beads-on-a-string

Five types of histones –packing of chromatin:

H2A, H2B, H3 and H4

2 copies of H2A, H2B, H3 and H4 form an octamer around which a segment of DNA double helix winds

nucleosomes

55A tall

110 A across

~140 base pairs of DNA associated with each histone core

Core dna has linker dna on both ends around the histone

There is 20-60 base pair “spacer”

Variation

There are specialized histones – can substitute for H3 and H2A

– specific characteristics to the DNA

Histones H3 and H4 can be modified – post-translational modifications

– can change the properties of nucleosomes that contain them

Histone Code

The pattern of major and specialized histones + their modifications

Specific to cell types

binds to DNA in the internucleosomal spacer region
and participates in compaction

140 bp of DNA

-10 to 11 nm

Solenoid Fibres

Nucleosomes compacted into a secondary helical structure

-30 nm diameter

Most condensed phase

Loops

Solenoids packed into loops attached at intervals of about 100,000 base pairs to a protein scaffold (H1)

Condensed section of chromosome

700nm

Whole Mitotic Chromosome

Subtopic

Decondensed

Interphase Nucleus

Each loop contains -10,000bp of DNA

most decondensed stage – interphase

Chromatin loops decondense at loop domain when the genes are expressed

Achieved by

Histone modifying enzymes

CHromatin remodelling complexes

RNA polymerase

Distinctive chromosome puffs arise and old puffs recede as new genes are expressed and old ones are turned off

Subtopic

Organization of the Human Genome

The Genome includes

Unique Sequences

Makes up about half of the DNA in the genome

Most single-copy DNA is found in short stretches, interspersed with various repetitive DNA families

Types

Genes

a sequence of DNA that is required for production of a functional product (polypeptide or RNA molecule)

Features

Adjacent nucleotide sequences – provide “start” and “stop” signals for the synthesis of mRNA transcribed

Promoters and regulatory elements can be sites of mutation

(either at 5’ or 3’ of the gene or in its introns)

enhancers, silencers, locus control regions

Size can be variable

Examples

1.7 kb

Insulin Gene

45 kb

LDL gene

2400kb

Dystrophin gene

Components

Exons

Protein/RNA encoding regions

200bp

small proportion of whole genome

ii.

Introns

Non-coding sequences

highly variable in size

typically several are found in most genes

total size of which frequently exceeds that of exons

There is a direct correlation between gene size and intron size

Some genes lack introns

e.g. histones

Types

Protein-encoding genes

-gene encodes the message transcribed to pre-RNA, to mRNA, which is translated into a product protein

-mutation or alteration in the DNA sequence results in an altered trait or disease

ii.

RNA-encoding genes

produce non-protein translated RNAs

not a protein

can profoundly alter normal gene expression and hence produce an altered trait or disease

can alter gene expression, and also produce altered traits or disease

3 major classes

Ribosomal (rRNA) and splicesomal (snRNA)

Transfer (tRNA)

Regulatory

Regulatory (usually inhibitory)

Micro RNA (miRNA; block mRNA)

Anti-sense RNA (block mRNA)

Riboswitch RNA (site on mRNA)

Gene Families

Many genes belong to families of closely related DNA sequences

Gene families thought to have arisen by duplication of primitive precursor genes as long as 500 million years ago

Types

Classic gene family

a group of genes that exhibit a high degree of sequence homology over most of the gene length

Gene superfamily:

a group of genes that exhibit low sequence homology, but they are similar in the encoded protein function and structure

(e.g. Ig super family, globin super family, myosins, G-protein receptor super family)

Clusters

Single Clusters

genes are organized as tandem repeated array

close cluster-

genes are controlled by a single expression control locus

e.g. Ig H chain

Compound Clusters

related and unrelated genes are
clustered on a single chromosome

E.g. Major Histocompatibility Complex - HLA genes

Multiple Clusters

e.g. Immunoglobulin superfamily

hundreds of genes encoding a conserved domain of ~ 70 aa which provide a distinct structural protein motif;

located throughout the genome

Dispersed

Genes encoding related functional proteins can be dispersed on a single chromosome or on 2 or more different chromosomes

Two clusters code for closely related globin chains expressed at different development stages from embryo to adult

Genes within each cluster are more similar in sequence -

evolved by duplication over last 100 million years

Exon-intron pattern is closely conserved –

more base changes have accumulated in the intron sequences than in the exons

Epigenetics & Epigenetic Control

Gene expression can be altered (to produce altered trait or disase) by chemical modification of

DNA (via methylation)

Methylation of cytosine nucleotides

decreases the probability of that segment being transcribed

induces silencing

involved in genomic imprinting

Genomic imprinting involved in normal development and several diseases

ii.

Histones (via acetylation, phosphorylation, ubiquitinization)

Methods

Acetylation

Acetylation of histone increases the likelihood of neighboring DNA segments to be transcrib

Methylation

Phosphorylation

Ubiquitination

works in conjunction with DNA methylation

Chromatin remodeling complexes

large multiprotein complexes containing helicases, which can unwind DNA double helices

control the local structure of chromatin

Affects the transcription of protein-encoding and RNA-encoding genes

Human Epigenome Project (HEP)

aims to identify, catalogue and interpret genome-wide DNA methylation patterns of all human genes in all major tissues

more base changes have accumulated in the intron sequences than in the exons

methylation variable positions (MVPs) are common epigenetic markers

Methylation is the only flexible genomic parameter that can change genome function under exogenous influence

it constitutes the main and so far missing link between genetics, disease and the environment that is widely thought to play a decisive role in the etiology of human pathologies

ii.

Non-repetitive DNA that is neither intron nor coding

Repeated Sequences

Types

Clustered Repeated Sequences

10-15% of the genome

-=tandem repeats

Satellite DNAs

can be several million base pairs

several percent of the DNA content of an individual chromosome

Examples

α-Satellite DNA

copies of 171-base pair unit

at the centromere of each human chromosome

ii.

long arrays of repeats

found in large inert regions on:

Chromosome 1

Chromosome 9

Chromosome 16

Chromosome 7

iii.

Minisatellites

iv.

Microsatellites

Interspersed Repeated Sequences

SINEs (short interspersed nuclear elements): Alu family

are about 300 base pairs in length

more than a million Alu family members in the genome

10% of human DNA

ii.

LINEs (long interspersed nuclear elements) family

-are up to 6-7kb in length

found in about 850,000 copies per genome

20% of the genome

Segmental duplications

can span hundreds of kilobase pairs

5% of the genome

Mitochondrial Genome

a circular chromosome, 16.5 kb

Mitochondrial DNA (mtDNA

37 genes

All mtDNA genes contain only exons

All mtDNA genes are maternally inherited in humans

13 Protein-encoding genes

The 13 protein-encoding genes are subunits of enzymes of oxidative phosphorylation (OXPHOS):

7 subunits of the NADH dehydrogenase complex

3 subunits of the cytochrome oxidase complex

2 subunits of the F0 ATPase complex

1 subunit (cytochome b) of the cytochrome c oxidoreductase complex

All other mitochondrial proteins (enzymes of the citric acid cycle, DNA and RNA polymerases)

encoded by nuclear DNA,

synthesized on cytoplasmic ribosomes

imported into mitochondria via chaperone proteins

(hsp60, hsp 70)

24 RNA-encoding genes

2 rRNAs (16S and 23S)

22 tRNAs

corresponding to each AA

Genetic Variation

e.g. polymorphisms giving rise to blue eyes versus brown eyes, straight hair versus curly hair

Genetic Polymorphisms

A difference in DNA sequence among individuals, groups or populations

Variant more common than 1% in the general population

Classifications

Inherited variation and polymorphism in DNA

Single Nucleotide polymorphisms (SNPs)

a single base difference in DNA

Transition substitution

between purines (A,G) or between pyrimidines (C,T)

Transversion substitution

between a purine and pyrimidine

1 in 1000 bp differ in any two randomly chosen humans

Since the haploid genome is 3 × 109 bp, 3 million differences between any two randomly chosen individuals

A subset of ~10% of the most frequent SNPs chosen to serve as the markers for a high-density map of the human genome (hHapMap)

A source of variation in a genome

Variable Number Tandem Repeats

Insertion-deletion polymorphisms

A short nucleotide sequence is organized as a tandem repeat

These sequences are found on different chromosomes

can have different length between individuals

Each variant acts as an inherited allele

can be used as genetic markers in

linkage analysis of genomes

forensic investigations,

DNA fingerprinting

DNA sequences in the VNTR polymorphs, scattered throughout the genome are somewhat homologous to each other

ii.

Probes can be designed to detect many of these VNTR loci simultaneously

iii.

Each probe would generate a complex, unique pattern based on the VNTRs it picks up

iv.

The most informative markers have several alleles, so that no two unrelated individuals would exhibit the same pattern upon electrophoresis

Probes are selected that identify VNTRs at many
different loci

vi.

Only identical twins show the
same pattern

vii.

DNA fingerprinting is widely
used for individual identification

suspects in criminal cases

the remains of victims and military
personnel

testing of paternity

Classifications

Microsatellites

Short segments of 2, 3 or 4 to 6 bp repeatedAre prone to deletion, insertion, or duplication

Are repeating sequences of 1-5,6 base pairs of DNA

One common example - (CA)n repeat, where n is variable between alleles

are sections of DNA composed of short motifs (e.g. CA, GTG, TGCT etc) arranged in tandem

Have high mutation rate

ensures high level of polymorphism

attributed to relatively high rates of error during during

DNA replication (slippage) and

recombination (unequal crossover)

become useful for examining relationships among individuals and breeding groups within populations

Are used as molecular markers for kindship and in population studies

Stable Polymorphisms

Inherited variation and polymorphism translated in polymorphic proteinsAre clinically important:TransplantationBlood TransfusionHemolytic Disease of the Newborn

Types

Blood groups and their polymorphisms

ABO Blood Groups

Determine by one gene on chromosome 9

– that encodes a galactosyl transferase

ABO polysaccharide antigens are exquisitely immunogenic

Multiallelism:

Three alleles, two of which (A and B) are codominant and the third (O) is recessive (silent)

There is a four-nucleotide sequence difference between A and B alleles resulting in amino acid changes that alter the specificity of the galactosyl transferase encoded by the ABO gene

The O allele has a single base-pair deletion which causes a frame-shift mutation that eliminates the transferase activity in type O individuals

4 phenotypes

Group A

Antibodies Present: Anti-B

Antigens Present: A antigen

Distribution: Europe, Australia, Northern Canada

Group B

Antibodies Present: Anti-A

Antigens Present: B Antigen

Distribution: Asia

Group AB

Antibodies Present: None

Antigens Present: A and B Antigens

Distribution: South America; North America

Group O

Antibodies Present: Anti-A, Anti-B

Antigens Present: None

Blood Donation

Preferably a patient receives blood of his or her own ABO groupABO compatibility of donor and recipient – essential to graft survival

Rh System

The name comes from Rhesus monkeys – used for the experiments leading to the this system discovery

Rh factors are glycoproteins encoded by alleles at 3 loci (C,D,E) exhibiting dominant/recessive expression

The D locus products are the most significant in terms of immune response

THey are encoded by the gene RHD which is found on chromosome 1

Phenotypes

Rh-positive individuals

who express, on their red blood cells, the antigen Rh-D, a polypeptide encoded by a gene RHD on chromosome 1

Rh-negative individuals

do not express the antigen

Frequency of Rh negative varies enormously: and

~17% in Whites,

7% in African Americans

0.5% among Japanese

Hemolytic Diseases of the Newborn

If a Rh-negative pregnant woman – is carrying an Rh-positive fetus, hemolytic disease of the newborn can result:

Small amounts of fetal blood cross the placental barrier and reach the maternal blood stream

The mother forms antibodies that will return to the fetus and damage the fetal red blood cells

Treatment:

Rh immune globulin at 28 to 32 weeks of gestation and again after pregnancy

Major Histocompatibility complex

MHC – composed of a large cluster of genes located on the short arm of chromosome 6

3 Classes of Clusters (I, II, III)

class I and class II genes correspond to the human leucocyte antigen genes (HLA)class III unrelated to HLA genes

Class I Genes

class I and class II genes correspond to the human leucocyte antigen genes (HLA)

3 polymorphic class I α chain
genes:

HLA-A

767 alleles

HLA-B

1178 alleles

HLA-C

439 alleles

Encode proteins that are integral part of the plasma membrane of all nucleated cells

a variable heavy chain

another polypeptide, β2-microglobulin

2 polypeptide subunits

Class II

3 polymorphic Class II molecules, each consisting of an α and β chain

HLA-DP

27 α, 133 β alleles3591 combinations

HLA-DQ

34 α, 96 β alleles 3264 combinations

HLA-DR

3 α loci, 9 β loci,each with multiple alleles - many combinations

Encode integral membrane cell surface proteins

Heterodimers

alpha subunit

Beta subunit

class III unrelated to HLA genes

Genes present within the MHC complex but are functionally unrelated to HLA class I and II

Some genes are associated with diseases

congenital adrenal hyperplasia

hemochromatosis

Clinical Relevance

Rare Variant

Allele frequency of less than 1%

Genetic Mutation

A change (associated with disease or risk of disease) in the nucleotide sequence of a DNA molecule

e.g. changes associated with disease or risk of diseasee.g. changes resulted from damage by external agents like radiation or viruses

Genotypes & Phenotypes

Genotype

exact description of the genetic constitution of an individual, with respect to a single trait or a larger set of traits

Genotyping Analysis

detection of the genotypes of individual SNPs

High-throughput SNP genotyping

process of quickly and cost-effectively identification of the SNP in many individuals in a given population

Used in association studies

HapMap

Derived from Genetic Sequences

Similar among 2 individuals but about every 1,000 nucleotides, the sequences will differ

Changes in the DNA sequences – may increase the risk to high blood pressure, cancer, other diseases

SNPs

There are about 10 million common SNPs in a person's chromosomes

Haploptypes

Genetic variants that are near each other tend to be inherited together

E.g. all of the people who have an A rather than a G at a particular location in a chromosome can have identical genetic variants at other SNPs in the chromosomal region surrounding the A

These regions of linked variants are known as haplotypes

Origin of Haplotypes

Over the course of many generations,
segments of the ancestral chromosomes
in an interbreeding population are
shuffled through repeated recombination
events

Some of the segments of the ancestral
chromosomes occur as regions of DNA
sequences that are shared by multiple
individuals

all humans today are descended from
ancestors who lived in Africa about
150,000 years ago

A Hapmap is catalog of common genetic variants

what these variants are,

where they occur in our DNA

how they are distributed among people within populations and among populations in different parts of the world

Hapmap Project is designed to provide information that other researchers can use to link genetic variants to the risk for specific diseases

Construction of the HapMap

Single nucleotide polymorphisms (SNPs) are identified in DNA samples from multiple individuals

Adjacent SNPs that are inherited together are compiled into “haplotypes”

“Tag” SNPs within haplotypes are identified that uniquely identify those haplotypes

Selected Populations

The DNA samples for the HapMap have come from a total of 270 people

The Yoruba people of Ibadan, Nigeria, provided 30 sets of samples from two parents and an adult child

In Japan, 45 unrelated individuals from the Tokyo area provided samples

In China, 45 unrelated individuals from Beijing provided samples

Thirty U.S. trios provided samples - collected in 1980 from U.S. residents with northern and western European ancestry

Health Benefits of the HapMap

Identifying haplotypes can help in association studies (compare the haplotypes in individuals with a disease to the haplotypes of a comparable group of individuals without a disease )

Cancer, stroke, heart disease, diabetes, depression, and asthma - result from the combined effects of a number of genetic variants and environmental factors

Medical treatments could be customized, based on a patient's genetic make-up, to maximize effectiveness and minimize side effects

Genetic variants contributing to longevity or resistance to disease could be identified, leading to new therapies with widespread benefits

Phenotype

observable characteristics of an individual that have developed under the combined influences of the individual’s genotype and the effects of environmental factors

Cell Cycle: Checkpoints and Control

Cell Cycle

Phases

G0 - Gap

Cells in Go often called “quiescent

still have secretion

still attack pathogens

active repression of genes needed for mitosis

quiescence can be either:

Temporary

Permanent

terminally differentiated

never reenter the cell cycle, but carry their function until they die

Example:

Most lymphocytes in human blood are in Go

but with proper stimulation reenter the cell cycle in G1

Nerve cells

Red blood cells too?

G1 - Gap

growth and preparation of the chromosomes for replication

immediately after mitosis or G0

some cells pass through this stage in hours, others in days or years

Examples

Liver cells

may enter G0 but return to G1

Cell damage?

S - Synthesis

follows G1 phase

synthesis of DNA and duplication of the centrosomes

DNA synthesis

begins at hundreds-thousands of sites

origins of DNA replication

each chromosome replicates

– two sister chromatids

identical copy of the original DNA double helix

chromatids have at the end the telomeres

chromatids held together by the centromeres

associate with proteins=kinetochore

- DNA content of the cell doubled

G2 - Gap

at the end of S phase

preparation for mitosis

- RNA and protein synthesis continues

no DNA synthesis

cells enlarge

individual chromosomes begin to condense and become visible under microscope

M - Mitosis

Prophase

Begins mitosis

Gradual condensation of the chromosomes

Beginning of the formation of the mitotic spindle

Pair of microtubule organizing center = centrosomes form foci

Centrosomes gradually move to the poles of the cell

Prometaphase

Nuclear membrane breaks up

Chromosomes disperse within the cell

Chromosomes attach by the kinetochores to microtubules of the mitotic spindle

Chromosomes move towards the midway between the poles

= congression

Continue condensation of the chromosomes

Metaphase

Chromosomes reach maximum condensation

Chromosomes arranged in the equatorial plane of the cell

Balanced by equal forces from the kinetochore

Facilitate the analysis of chromosomal abnormalities

Anaphase

Begins suddenly

Chromosomes separate at the centromere

The chromatids become daughter chromosomes, moving at the poles

Telophase

Chromosomes decondensate

Nuclear membrane forms

Two daughters nuclei are formed

The nuclei resume interphase appearance

Subtopic

Meiosis

Meiosis I

Prophase I

Leptotene

chromosomes replicated in S phase become visible and condensate

Sister chromatids closely aligned

Zygotene

homologous chromosomes begin to align along their length

pairing/synapsis

Chromosomes held together by a synaptonemal complex

Pachytene

Chromosomes tightly coiled

Homologues appear as a bivalent (tetrad)

Meiotic crossing over takes place

Diplotene

Synaptonemal complex begins to break down

2 components of each bivalent now begin to separate

The 2 homologs are held together at chiasmata (crosses)

- ~ 50 in spermatocytes

Diakinesis

Chromosomes reach maximal condensation

Metaphase I

-nuclear membrane disappears

-spindle forms

-paired chromosomes align themselves on the equatorial plane

Anaphase I

disjunction takes place

2 members of each bivalent move apart and the centromeres are drawn to opposite poles

-chromosome number is reduced in half

-original paternal and maternal chromosome sets are sorted into random combinations (223)

-e.g a typical chromosome will have 3 to 5 segments alternately paternal and maternal in origin

-errors can occur

meiotic arrest or cell death

missegregation of chromosomes

nondisjunction

Telophase I

-the 2 sets of chromosomes are grouped at the opposite poles

Cytokinesis

spermatogenesis

the cytoplasm is more or less equally distributed between the new cells

-oogenesis

secondary oocytes receives almost all the cytoplasm and the reciprocal product becomes the first polar body

there is no S phase between the first and second meiotic divisions

-cell divides into 2 daughter cells and enters mitotic interphase

Meiosis II

Similar to an ordinary mitosis
except the chromosome number
is haploid

The 2 daughter cells from
meiosis I divide to form 4
haploid cells with 23
chromosomes

Segregation of the different
paternal or maternal alleles of
each gene

Meiosis Summary

Subtopic

Cell Cycle Control

Cell cycle controlled by proteins (Cyclins and Cyclin-Dependent Kinases (CDKs)) in the cytoplasm:

G1 Control

Proteins Involved

Cyclin D

CD4, 6

Mechanism

G1 Cyclins bind to their Cdks

cell prepares for replication

REgulation of the G1-S transition by p53

S Control

Proteins Involved

Cyclins E and A

CD2

Mechanisms

Reactions leading to S phase and M phase

Cdk activation/inactivation activity

Cyclin E binds to Cdk2

S-phase promoting factor (SPF)

cyclin A bound to Cdk2

enters the nucleus

prepare the cell to duplicate its DNA

DNA replication continues

- cyclin E is destroyed

mitotic cyclins begin to rise (in G2)

M Control

Proteins Involved

Cyclin B

CD1

Mechanism

M-phase promoting factor

mitotic B cyclins bound to M-phase Cdk (Cdk1) lead to

Assembly of the mitotic spindle

Breakdown of the nuclear envelope

Cessation of all gene transcription

These events take the cell to the metaphase of mitosis

Anaphase-promoting complex (APC)

Is activated by the M-phase promoting factor

Allows sister chromatids at the metaphase plate to separate and move to the poles (anaphase)

Separation of chromatids depends on the breakdown of the cohesins

Cohesin breakdown is caused by a protease called Separase

Separase is kept inactive by the securin (inhibitory chaperone)

Anaphase begins when the APC destroys securin

Destroys B cyclins

attaching to ubiquitin-target for degradation by proteasomes

Checkpoints: Quality control of the cell cycle

DNA Damage Checkpoints

-sense DNA damage before the cell enters S phase (G1 checkpoint)

Inhibition of Cdk2- stops cell cycle progression until the damage is repaired

Damage too severe – cell enters apoptosis

-sense DNA damage after S phase (G2 checkpoint)

Inhibition of Cdk1 – prevent the cell to go from G2 to mitosis

Spindle Checkpoints

-detect any failure of spindle fibers to attach to kinetochores

-detect improper alignment of the spindle itself and block cytokinesis

-arrest the cell in metaphase until all the kinetochores are attached correctly (M checkpoint)

-trigger apoptosis if the damage is irreparable

DNA Replication Checkpoints

-during the replication of the millions or billions of DNA base pairs

damaged template, protein complexes bound to DNA, poor supply of NTPs = barriers in replication

stall replication forks

insert stalled fork

can rearrange, break, collapse through disassembly of the replication complex

ATM Role in Cell Cycle Control

ATM (Ataxia telangiectasia mutated)

the same disease name

patients very high risk of cancer

(~100 fold increase)


ATM protein:

-detects DNA damage

especially double-strand breaks


interrupt cell cycle when damage is detected

-maintain normal telomere length

Regulation of G1-S transition by p53

P53 protein senses
DNA damage

-blocks Cdk2
stops progression of cell
cycle in G1

P53 induces apoptosis-

cells with defects are 
forced to commit suicide

P53 role in apoptosis - intrinsic pathway

BCL-2 Family

contain signature domains of homology called BCL-2 homology (BH) domains

(termed BH1, BH2, BH3, and BH4)

Protein Classes

Pro-apoptotic proteins:

Several BH domains (Bax and Bak)

those that only have the BH3 domain, such as Bid, Bad, Bim, Bmf, PUMA, and NOXA

– permeabilization of the mitochondrial membrane

Upon membrane permeabilization, cytochrome c and the pro-apoptotic proteins (SMAC/DIABLO) are then able to translocate from the inter-membraner space of the mitochondria into the cytosol

Anti-apoptotic proteins:

BCL-2 and BCL-XL act to prevent permeabilization of the outer mitochondrial membrane by inhibiting the action of the pro-apoptotic Bcl-2 proteins Bax and/or Bak

Intrinsic (Mitochondria) pathway

Cytochrome c binds the adaptor apoptotic protease activating factor-1 (Apaf-1), forming a large multiprotein structure known as the apoptosome

The primary function of the apoptosome seems to be multimerization and allosteric regulation of the catalytic activity of caspase 9

activation of caspase 9

activation of the downstream effector caspases 3, 6, and/or 7

Caspase Cascade

destroy essential cellular proteins, leading to controlled cell death

2 tiers of caspase activation during apoptosis:

Initiator caspases

(caspases 2, 8, 9, and 10)

activated through the apoptosis-signaling pathways

Effector caspases

(caspases 3, 6, and 7)

carry out apoptosis

Evasion of apoptosis as a hallmark for cancer

The balance between cell proliferation and apoptosis is influenced by

Oncogenes

genes that contribute to the development of cancer

Tumour Suppressor Genes

genes that encode proteins that normally suppress tumor formation

Regulation of p53 activity

Role of RB in the cell cycle regulation

Genetic Diseases

Classification

(and prevalence; total prevalence = 31.5-73 per 1000)

Single gene disorders

Caused by mutant genes - either a single allele or a pair of alleles e.g. sickle cell trait/sickle cell anemia

Autosomal

Dominant

(P= 2-2.5 per 1000)

Familial Hypercholesteremia

Neurofibromatosis

Recessive

(P= 3-9.5 per 1000)

Tay Sachs

Cystic Fibrosis

X-Linked

P= 0.5-2 per 1000

Dominant

Vitamin-D resistant Rickets

Incongenentia Pigmenti

Recessive

Hemophilia A

Duchenne’s Muscular Dystrophy

Chromosomal disorders

(P= 6-9 per 1000)Defective chromosome (number and/or structure)

Down syndrome (Trisomy 21)

Multifactorial disorders

Gene/Chromosome disorder plus other factors, like environmental factorsPrevalence of congenital malformations = 20-50 per 1000

birth defects (cleft lip and palate),

heart disease

hypertension

obesity

diabetes

Mitochondrial disorders

Diseases caused by alterations in the small cytoplasmic mitochondrial chromosome