Genetics Part 1 (Review of basic genetics)

Define a Nucleotide

• BUILDING block of DNA or RNA (nucleic acids)

• Purine or pyrimidine base attached to a ribose phosphate

  • EX: Adenine, Guanine, Thymine, Cytosine, Uracil

• DON’T have the phosphate

Define a Codon

Set of three nucleotides that codes for a particular amino acid

Define an amino acid

• BUILDING blocks of PROTEIN

How many amino acids are in human proteins?

20

How many amino acids are essential in human proteins?

• 9

• Cannot be synthesized in humans

• Must be provided by dietary sources

How many amino acids are non-essential in human proteins?

• 11

• Can be synthesized in the body

• Not required in the diet

Define a Protein

Polymers of amino acids

• Enzymes

• Structural

• Carrier

• Receptor

• Regulatory

Enzymes….

carry out biochemical reactions

Structural…

holds things together (ex: collagen, keratin)

Carrier…

transfer small molecules (ex: hemoglobin, ferritin)

Receptors…

bind circulating molecules (ex: insulin, cholesterol)

Regulatory….

turn genes on and off (ex: growth factors)

Explain what DNA is?

• Double stranded

• Deoxyribose nucleotides (A, T, G, C)

• Repository of genetic information

• Replicated in cell divisions

• Stored as chromosomes

Explain what RNA is?

• SINGLE stranded

• Ribose nucleotides (A,U,G, C)

• Short half life

• mRNA, tRNA, rRNA

• Regulatory functions (snRNA)

Each chromosome is _____ molecule of ____ and associated protein

• one

• DNA

What are associated with chromosomes?

• Karyotype

• Autosomes

• Sex Chromosomes

• Homologs

• Sister Chromatids

• Alleles

How many pairs of chromosomes are there in normal content?

23

What explains how Nuclear chromosomes come in pairs?

• Humans are diploid

• ONE from MOTHER (maternal) and ONE from FATHER (paternal)

Define a Karyotype

• Graphic arrangement of chromosomes in a cell

• Arranged by size, position of centromere

• Numbered 1-22, X and Y

Define autosomes

• Chromosomes that are the same in men and women

• Numbered in order of size (sort of)

• Chromosome 1-22

• Pairs

Define sex chromosomes

• chromosomes that are different in men and women

• X and Y

• Y contains genes that determine male sex and little else

• X contains A LOT of genes that have nothing to do with SEX DETERMINATION

  • color blindness, hemophilia, muscular dystrophy

Define Homologs

• Pairs of the same chromosome

• One maternal, one paternal

Define Sister chromatids

• Duplicated copies of chromosomes

• After DNA replication, before cell division

Define Alleles

• Variants of individual genes

• Variants at a single genetic locus

• An individual has two alleles for each of their gene

Explain chromatin structure

Each chromosome is a molecule of DNA→ Coiled into chromatin→ Coiled around central cores called nucleosomes→ Associated with histone proteins →TIGHTNESS of coiling controls GENE EXPRESSION

Each Chromosome is a molecule of____

DNA

Chromosomes are numbered in order of _____

size

Chromosomes are grouped by position of ______

centromere

• Metacentric

• Submetacentric

• Acrocentric

What is a centromere

(Chromosome structure)

• Divides chromosomes into TWO parts (arms)

• Attachment point for SPINDLE FIBERS during cell division

Define what “Arms” are in the structure of chromosome

• Human chromosomes that have “arms”

  • Short arm (p)

  • Long arm (q)

• Short arms of acrocentric chromosomes contains REPETITIVE sequences for rRNA

Advanced staining techniques reveal dark and light _____ in _______

• Bands

• Chromosomes (Giemsa banding)

Each Chromosome has a unique and consistent banding pattern. What are they?

• Bands numbered from centromere (band 0) to end

  • Subbands given second and third number

  • 15q11.2

• Dark bands contains heterochromatin

• Light bands contain euchromatin

Dark bands are…

• More condensed

• Fewer expressed genes

Light bands are…

• Less condensed

• More expressed genes

Explain the components of Chromosome nomenclature

• First part is the number of chromosomes

  • 46 normal

• Second part is the sex chromosome constitution

  • XX, XY, XXY etc

• Third part is any abnormal chromones

  • 46, XX and 46, XY is normal

Is 46 chromosomes normal or abnormal?

Normal

Is 45 or 47 chromosomes normal or abnormal?

Abnormal

45, X is a _________ and associated with what disease?

Monosomy, Turner syndrome

“47, XY +21 is a male with trisomy 21” is correlated with what disease

Down syndrome

Define Trisomy

extra chromosome

• 21,13 and 18 are only seen at birth

Define Monosomy

missing chromosome

• Monosomy X (Turner syndrome) is only viable monosomy

Explain Translocations

• Piece of one chromosome attached to another

• RECIPROCAL, balanced (pieces of chromosomes interchanged)

Individuals with balanced translocation at risk for having children with _________

Unbalanced rearrangements

• Missing part of one chromosome, trisomic for a piece of another chromosome

Explain Deletions and the types of chromosomal deletions

Part of a chromosome is missing, Involve many genes, Cause of syndromes (Prader-Willi, Angelman, Williams, DiGeorge)

• Terminal

• Interstitial

Terminal deletion

at end of chromosome

Interstitial deletion

in middle of chromosome

Define insertions

• COUNTERPART of deletion

• Part of chromosome is inserted of duplicated

What is the interphase in the cell cycle?

• “Resting” part of the cycle

• Cell metabolism and growth (G1)

• DNA synthesis (S)- chromosome duplication

• Cell metabolism and growth (G2)

Describe the Mitotic phase (M) in the cell cycle

• Mitosis (division of chromosomes)

• Cytokinesis (cell division)

Describe the process of Mitosis in Cell Division

• Binary division of a cell

• DNA replication → sister chromatids of each chromosome, 92 sister chromatids

• Chromosomes condense (prophase)

• Chromosomes align along cell midline (metaphase)

• Chromosomes split at centromere→ One chromatid drawn to each side of cell →Spindle fibers →Anaphase

•Cell divides →Telophase→ Cytokinesis →46 chromosomes in each cell

Describe the process of Meosis in Cell Division

• REDUCTION division of a cell

• Occurs in gonadal tissues

  • In testes continuously after puberty

  • In Ovaries before birth →Ova held in stasis in meiosis I → Meiosis completed after fertilization

• Produces haploid gametes-1 chromosome of each pair

• DNA replication produces 92 sister chromatids

• TWO DIVISONS

Describe Meiosis I in Cell Division

• DNA replication

  • 92 sister chromatids

• Chromosome condensation

• Chromosomes associate in homologous pairs (tetrads)

• Crossing over between arms of homologous chromosomes

• One homologous chromosome (2 chromatids) goes into EACH daughter cell

• Cell now haploid

  • 2 sister chromatids of one chromosome

Describe Meiosis II in Cell Division

• Daughter cell now has 46 chromatids, 23 chromosomes

• NO further DNA replication

• Meiosis II is like mitosis

  • Sister chromatids divide at centromere

  • One chromatid into each daughter cell

• Daughter cells have 23 chromosomes, one of each homologous pair

Define Apoptosis and the type of pathways

• Programmed cell death

• Important process in embryogenesis

• Culls “sick” or stressed cells

• Produces apoptotic bodies → Cleared by phagocytic cells

• Intrinsic pathway

• Extrinsic pathway

Intrinsic pathway

• More like cell suicide

• Internal signals-SMACs and MACs

Extrinsic pathway

• External activation

• TNF

Describe the relationship between apoptosis and cancer

• Acts as a natural defense mechanism to eliminate potentially malignant cells.

• Its evasion is a hallmark of cancer development and progression.

• Normally, apoptosis removes damaged or dysfunctional cells to maintain tissue health →Cancer cells subvert this process to survive, proliferate, and resist treatment.

• Dysregulated BCL-2 proteins

• TP53 (Tumor Protein 53) Tumor suppressor gene inactivation

• BCL- 2 inhibitors and P53 reactivators are new drugs used in treatment of cancers

What are Dysregulated BCL-2 proteins and how do they affect apoptosis?

• Cancer cells often overexpress anti-apoptotic proteins like BCL-2 and BCL-xL, which INHIBIT a critical step in the intrinsic apoptotic pathway.

• Over 50% of cancers exhibit elevated BCL-2 levels, rendering cells resistant to DNA damage or chemotherapy.

What are TP53 (Tumor Protein 53) Tumor Suppressor Gene Inactivation and how do they affect apoptosis?

• Mutations in the p53 gene DISABLE its ability to trigger apoptosis in response to DNA damage. This allows cells with genetic errors to survive and accumulate oncogenic mutations. Over 50% of tumor cells have p53 mutations.

What is the function of the Mitochondria

• Main generator of ATP

• Kerbs cycle

• Urea cycle

• Outer membrane = permeable to small molecules

• Inner membrane = impermeable except by specific transporters, inner contains bound enzymes

• Matrix= soluble enzymes

Describe the Mitochondrial genome

• Mitochondria contain DNA

• Mitochondrial chromosome is circular, bacterial-like →No introns→ Replication start site

• Encodes some proteins →DNA replication enzymes→ Some respiratory chain components →Set of t-RNAs for protein synthesis

• Several copies in each mitochondrion

• Come from ovum (maternal)

• Most mitochondrial proteins and enzymes are encoded by nuclear genes and are imported into the mitochondria.

Describe the Mitochondrial disorders/ dysfunction

• Defects in respiratory chain activity

• DECREASED ATP generation from oxidation (O2)

• Wide range of symptoms: Weakness, myopathy, Neurologic symptoms, Diabetes, Blindness, Hearing loss

• Some due to mutations in mitochondrial genome

  • Variable due to heteroplasmy (number of mutant copies in each mitochondrion)

  • Show maternal pattern of inheritance

• Many due to nuclear genome mutations

What are the components of DNA?

• Purine bases

• Pyrimidine bases

• Nucleotides

Purine bases

• Adenine

• Guanine

• Aminopurines

Pyrimidine bases

• Uracil

• Cytosine

• Thymine

Nucleotides

• Ribose

• Deoxyribose

Amino groups of_____ form hydrogen bonds with keto groups of ________

• Purines

• Pyrimidines

  • Adenine-Thymine

  • Guanine-Cytosine

Describe the structure of DNA

• Bases are connected through ribose links

• 5’ carbon of one nucleotide attached to 3’ carbon of the next by phosphate bridge

• Gives DNA strand directionality – 5’ to 3’

Describe the process of DNA Replication

• Semiconservative replication

  • One strand is the template, unchanged

  • Second strand is newly synthesized

• DNA unwinds

• A’s in DNA strand bind with free dTTP

• T’s in DNA strand bind with free dATP

• G’s in DNA strand bind with free dCTP

• C’s in DNA strand bind with free dGTP

• DNA polymerase creates new strand

What enzymes are involved in DNA replication

• DNA polymerase

• Helicase

• Primase

• Topoisomerase

• Telomerase

• Ligase

Describe the function of Helicase in DNA replication

Unwinds the double strand

Describe the function of DNA polymerase in DNA replication

Creates the new DNA strand in the 5’ to 3’ direction

Describe the function of Primase in DNA replication

Synthesizes the RNA Primer needed to start sythesis

Describe the function of DNA Ligase in DNA replication

Connects Okazaki fragments on the lagging strand

Describe the function of Topoisomerase in DNA replication

Prevents tangling as the DNA unwinds

Describe the function of Telomerase in DNA replication

Extends telomeres at the end of chromosomes

A gene is a……

linear stretch of DNA that codes for something

Describe the process of transcription

RNA polymerase binds to a promoter sequence (like the TATA box) and synthesizes RNA in a 5’ to 3’ direction using uracil instead of thymine. This binding is regulated by enhancers and repressors.

• 5-methylguanin = beginning of mRNA strand (capping)

• PolyA tail= end of mRNA

Describe the purpose of transcription

To copy a DNA sequence into mRNA for PROTEIN SYTHESIS

What is the Function of mRNA?

Provides templates for protein synthesis

• synthesized by RNA polymerase II

What is the Function of tRNA?

Binds to specific amino acids and has an anticodon to align them correctly during protein synthesis

• Synthesized by RNA polymerase III

What is the Function of rRNA

Provides RNA for ribosomes. Particles containing mRNA binding and protein synthesis machinery

• synthesized by RNA polymerase I

Describe post-transcriptional modifications

Primary mRNA transcripts undergo capping (adding 5-methylguanine), addition of a PolyA tail, and splicing, where introns are removed by the spliceosome

• Alternative splicing can produce multiple proteins from one gene

Describe the purpose of translation

To synthesize a polypeptide chain based on the mRNA template

Describe the process of translation

mRNA binds to a ribosome (two subunits, rRNA and Proteins-elongation factors)→ translation starts at an AUG (methionine) codon→ tRNAs bring amino acids to the ribosome’s A site → they are added to the chain at the P site → empty tRNA leaves via the E site

Polypeptides fold based on _____and _______ interactions, often aided by ______proteins (protein folding)

• Ionic

• Hydrophobic

• Chaperone

Post-translational modifications include cleavage of _______, association of _______, adding cofactors (heme), or _______ in the Golgi apparatus

• pro-proteins

• Multiple subunits

• glycosylation

Describe the mechanisms of gene regulation

• Regulation occurs via DNA methylation (inhibits unwinding), RNA polymerase binding

• Histone acetylation (activates chromatin)

• Use of repressors or enhancers

• mRNA splicing

• miRNA & RISC binding

• mRNA Stability

• Protein folding, Post translational modifications

Explain the various causes of mutations

• Include single base changes, nucleotide deletions/insertions (causing frameshifts)

• Trinucleotide repeat expansions (leading to anticipation)

• Genome/chromosome level (Copy number variants, Recurrent deletions )

• Cell division: Frequency 10^-8 bases/division, Most are not in coding sequences, Most are benign

Describe Nucleotide level mutations

Single base changes

• May change a single amino acid

• May not change anything (synonymous mutation)

• Most common is C-T change at a GC doublet

• The C in a CG doublet may be methylated

• Deamination changes the C to a T

• During DNA replication the T would pair with an A instead of a G

Describe Nucleotide deletions or insertions

Single nucleotide additions or losses

• Within coding sequence will cause a frameshift

• Codons after the mutation will all be changed

• Leads to severely defective protein after mutation

• Early stop codon

Small deletions or insertions

• In multiples of 3 can be in frame

  • Delete single or a few amino acids

  • Protein may still function

  • e.g. delF508 in cystic fibrosis deletes a single phenylalanine

Other changes will cause a frameshift

Describe trinucleotide repeat expansion

• Some genes contain repetitive sequences of trinucleotides

  • Huntington disease - CAG – glutamine repeat in coding region

  • Fragile-X – CGG near promotor

  • Myotonic dystrophy – CTG in 3’ untranslated region

• In meiosis, the repeat region may expand

• If expansion is greater than some threshold, the gene will not function properly causing a genetic disorder

  • Huntington disease: <26 CAG nl, >36 causes disorder

  • Fragile-X: 5-44 CGG nl, >200 causes disorder

  • Myotonic dystrophy: 5-27 CTG nl, >50 causes disorder

• Expansion occurs preferentially in maternal meiosis (FraX, MD) or paternal meiosis (HD)

• Causes “anticipation” in pedigrees

  • Earlier onset in later generations

Describe copy number variant mutations

• Genome contains many duplicated genes

  • Beta-globin

  • Alpha-globin

  • Color vision locus

• Genes may misalign during meiosis

• Crossover will delete gene on one chromosome, duplicate gene on the other

Describe X-linked color blindness mutation

• X-chromosome contains one red-sensitive pigment gene and several green sensitive ones

  • Only first green sensitive gene is expressed

• Color pigment genes have very similar DNA sequences and may misalign during Meiosis I

• Crossover between genes causes deletion of green sensitive visual pigment

• Crossover within a gene causes fusion gene with differing color sensitivity

Describe recurrent deletion mutation

• Several genetic disorders are caused by deletions which occur in the same place in affected individuals

  • Severe Hemophilia A – X-chromosome, Factor VIII gene

  • DiGeorge syndrome – chromosome 22q11.2

  • Prader-Willi syndrome – chromosome 15q11.2-13

  • Williams syndrome – chromosome 7q11.23

• Regions flanked by repetitive or inverted sequences

• Misalignment and crossover causes deletion in the same places

Describe severe hemophilia A mutation

• Duplicated, inverted small gene (A) in Factor VIII gene

• Pairs with homologous gene telomeric to Factor VIII

• Crossover causes inverted deletion of Factor VIII

• Responsible for 40% of severe Hemophilia A

• Mutation occurs only in male meiosis

What are the types of DNA repair?

• Nucleotide excision repair

• Base excision repair

• Mismatch repair

Nucleotide excision repair

• Damaged bases cause DNA helix distortion, e.g. thymidine dimers caused by UV light

• Damaged segment cut out by endonucleases

• Segment removed by helicase

• New segment synthesized by DNA polymerase

• Reconnected by ligase

Base excision repair

• Single chemically damaged base cut out by glycosylase

• New base inserted by ligase

Mismatch repair

• Mismatched bases introduced during errors in DNA replication

• Mismatched segment recognized in newly replicated strand by repair system

• Segment cut out by endonuclease

• New strand made by DNA polymerase

• Reconnected by ligase