PCB3063 EXAM THREE

Cancer

uncontrolled cell growth and spread of abnormal cells

Cyclin-dependent kinases & Ras-signaling pathway

two mechanisms involved in the regulation of the cell cycle

Protooncogenes

stimulate cell division

• mutations cause increased cell growth (gain-of-function)

Tumor Suppessor Genes

stop the cell cycle, DNA repair, or induce apoptosis

• mutations cause damaged cells to continue growing and dividing

Basal Lamina and Attachment Proteins

mutations allow cells to break away from their original tissue

DNA repair systems

cells accumulate more mutations due to lack of repairs

Cell identity

cancer cells can have an abnormal appearance and invade other tissue

Benign Tumors

form noncancerous, single multicellular mass

• due to mutations in TS genes and proto-oncogenes

Malignant Tumors

benign cells break away and starts multiplying

• invade other tissues and can cause life-threatening problems

Hyperplasia

increase in cell number

Dysplasia

abnormal cells

Metastasis

cell migration, breaking free from tissue

Malignancy

cell invasiveness

Driver Mutations

gives a growth advantage to a tumor cell

Passenger Mutations

no direct contribution to the cancer phenotype and is acquired over time during cell division

XP

involved in nucleotide excision repair and mutations lead to skin cancer

FAP

genetic predisposition to colon cancer due to:

mutation in APC gene

• tumor-suppressor controlling cell-cell contact and growth inhibition

mutation in MUTYH gene

• gene involved in DNA repair and mutations can cause loss of heterozygosity

pRB

phosphorylated to obtain transcription factor E2F

• E2F activated transcription of genes required for the cell cycle like cyclins

Hardy-Weinberg Assumptions

1. no natural selection

2. random mating

3. no new alleles from mutations

4. infinitely large population

5. no migration

Hardy-Weinberg for two alleles

p = dominant allele & q = recessive allele

• p + q = 1

• p² + 2pq + q² = 1

Hardy-Weinberg for multiple alleles

p + q + r = 1

• p² + q² + r² +2pq + 2pr + 2qr = 1

Hardy-Weinberg for X-linked alleles

frequency of allele = frequency of males expressing the X-linked gene

• female carriers = 2pq

• females affected = q²

Hardy-Weinberg with Fitness

1. find total fitness

2. find new values

Directional Selection

selection favors one genotype related to a phenotypic extreme

• shift in population mean

Stabilizing Selection

both phenotypic extremes are selected against and intermediate types are favored

• mean stays the same but variance decreases

Disruptive selection

both phenotypic extremes are selected for and intermediates are selected against

• increasingly bimodal population

Migration effect on allele frequency

p_i’ = (1 - m)p_i + mp_m

Positive Assortive Mating

similar genotypes are more likely to mate than dissimilar ones

Negative Assortive Mating

dissimilar genotypes are more likely to mate than similar ones

Inbreeding

mating individuals are related, removing heterozygosity

Change in Allele Frequency

s = 1 - W_aa

for lethal recessive alleles

Meristic Traits

polygenic traits in which the phenotype is counted in integers

• ex. number of appendages

Threshold Traits

polygenic but have a small number of discrete phenotypic classes, often influenced by environmental factors

Estimating number of polygenes

n = number of polygenes

• any amount of polygenes: (1/4)ⁿ = 1/(individuals expressing one extreme)

• low number of polygenes: 2n + 1

Variance

Covariance

Correlation Coefficient

r = cov_xy/s_xs_y

Heritability

proportion of total phenotypic variation in a population due to genetic factors

Broad-sense Heritability

H² = V_g/V_p

Phenotypic Variance

V_P = V_G + V_E + V_GxE

Narrow-sense Heritability

• h² = V_A/V_P

• h² = (M₂ - M)/(M₁ - M)

• h² = R/S

Genetic Variance

genetic variance = additive variance + dominance variance + interactive variance

• V_G = V_A + V_D + V_I

Quantitative Trait Loci

multiple genes contributing to a quantitative trait

Log Of Odds (LOD)

odds = [prob(data: QTL)] / [prob(data: no QTL)]

• LOD ≥ 3 are considered significant

DNA methylation

reversible addition of methyl groups to DNA, blocking transcription factors by blocking the major groove

CpG islands

• clusters of CpG dinucleotide sites in promoter and upstream sequences

• genes with adjacent methylated regions are transcriptionally silenced

heterochromatin methylation

dinucleotides adjacent to genes but are in repetitive DNA sequences like the centromere, maintaining chromosome stability by preventing translocations and related abnormalities

demethylation

• passive - failure to methylate new DNA strands

• active - removal of methyl groups independent of DNA replication

histone modifications

covalent modification of amino acids of the N-terminal through acetylation, methylation, and phosphorylation

histone acetylation

relaxes the grip of histones of DNA, allowing transcription to occur

short ncRNAs

act as repressors of gene expression

• piRNA interact with proteins to form RNA-protein complexes that participate in epigenetic gene-silencing in germ cells

Antisense lncRNA

lncRNA genes partially overlap protein-coding genes and transcribe in opposite directions, reducing transcription

Intronic lncRNAs

lncRNA genes are located in introns and transcription does not overlap adjacent exons

Bidirectional lncRNAs

lncRNA genes use protein-coding genes’ promoter but is transcribed in opposite directions

Intergenic lncRNAs

lncRNAs are discrete transcription units located outside protein-coding genes

Monoallelic expression

only the parental or maternal allele is transcribed and the other is transcriptionally inactive

• occurs in genomic imprinting, autosomal genes, and random inactivation of an X chromosome

Genomic Imprinting

expressed in a parent-of-origin pattern

• genomic region escapes demethylation and remethylation during gamete formation making the alleles stay transcriptionally silent

• usually found in clusters on the same chromosome

Epimutations

mutations in imprinted genes that affect adjacent genes due to DNA sequence changes or coordinately controlled imprinted genes

random inactivation of an X chromosome

happens in cells of female mammals

• Xist lncRNA turns inactivated chromosome into a Barr body

• Tsix lncRNA prevents the active chromosome from being silenced and represses the Xist lncRNA

Monoallelic expression (MAE) of autosomal genes

• scattered throughout the genome

• causes expression of both alleles (biallelic), only the maternal or paternal allele, or expression of neither allele

genomic hypomethylation

occurs in all cancer cells allowing for unrestricted transcription of many gene sets

selective hypermethylation

occurs in certain regions of the genome in cancer cells to silence certain genes like tumor-suppressors

alternative splicing

creates different spliceforms/variation which different protein variant

isoforms

different variants of mRNA caused by alternative splicing

cassette exons

• most common type of alternative splicing in mammals

• exons can be excluded by joining the 3’ end of an upstream exon to the 5’ end of a downstream exon

alternative splice site

type of alternative splicing where an exon near an intron gets spliced as well

alternative promoters

type of alternative splicing where there is more than one site where transcription is initiated

CT/CGRP gene

• in thyroid cells, mature mRNA contains only the first four exons which is processed into a calcitonin (CT) peptide that regulated blood calcium levels

• in neurons, exon 4 is alternatively spliced out, processing a peptide hormone, CGRP, that stimulates the dilation of blood vessels

proteome

number of proteins in an organism

Dscam gene

• gene in fruit flies that codes for a protein that guides axon growth

• alternative splicing leads to different tags on neurons, allowing for correct wiring

splicing enhancers and silencers

cis-acting sequences which promote or inhibit the splicing of nearby splice sites

SR proteins

bind to enhancers and activate splicing by recruiting spliceosome components

hnRNPs (heterogeneous nuclear ribonucleoproteins)

class of proteins that bind to splicing silencers and inhibit splicing

RBPs (RNA-binding proteins)

class of proteins that bind to RNA sequences or structures and can bind or hide splice sites to promote the use of alternative sites

Sex lethal (Sxl) gene

regulatory gene that encodes an RNA-binding protein

sex determinism in fruit flies

1. transcription factors encoded by genes on the X chromosome allow for the transcription of the Sxl (sex lethal) gene

   • lower concentration of transcription factors in males does not activate transcription

2. SXL protein binds to tra (transformer) gene and uses alternative splicing to create a fully functional SR protein

   • translation of male mRNA leads to a nonfunctional protein

3. male and female spliceoforms create different DSX (double sex) isoforms

   • DSX-F proteins repress transcription of genes that control male sexual development

   • DSX-M proteins activate transcription of genes that control male sexual development and repress the transcription of genes that control female sexual development

exoribonucleases

enzymes that degrade RNA via the removal of terminal nucleotides

deadenylation-dependent decay

1. process initiated by deadenylases, enzymes that shorten the poly-A tail

   • if it shortens thedeadeny poly-A tail to less than 30 nucleotides, mRNA will be degraded

2. Exoribonuclease destroys the mRNA in a 3’ to 5’ manner

OR

3. decapping enzymes remove the 5’ cap, allowing XRN1 to destroy mRNA in the 5’ to 3’ direction

deadenylation-independent decay

endoribonucleases that cleave mRNA internally

P (processing) bodies

mRNAs accumulated in cytoplasmic complexes due to not being actively translated

Adenosine-uridine rich element (ARE)

cis-acting sequences that regulate mRNA stability

• ARE-containing mRNAs encode proteins that promote cellular multiplication

TTP (tristetrapolin)

recruits decay machinery to promote mRNA degradation

RNAi (RNA interference)

mechanism by which ncRNA molecules guide the posttranscriptional silencing of mRNAs in a sequence-specific moments

microRNA (miRNA)

regulatory ncRNA that causes translational downregulation of mRNA due to complementary base pairing

• if the match is perfect, the mRNA is degraded

• if the match is partial, translation is blocked

small interfering RNAs (siRNAs)

RNA derived from double-stranded RNA caused by virus infection

1. Dicer enzyme cleaves and evicts one of the two strands as a siRNA guide to recruit RISC to a complementary mRNA

2. RISC cleaves mRNA in the middle of the siRNA-mRNA complementary region

   • cleaved mRNA lacking a cap or poly-A tail is degraded by exoribonucleases

microRNA formation

1. primary miRNAs are created from miRNA genes

2. Dorsha enzymes removes noncomplementary 5’ and 3’ ends to produce pre-miRNAs and hairpins are exported to the cytoplasm

3. cleaved by Dicer to produce mature double-stranded miRNAs

4. further processing by RISC to become single stranded

long noncoding RNAs (lncRNAs)

• linked to diverse regulatory functions with chromatin modification, altering patterns of gene expressions, and alternative splicing

• can function as competing endogenous RNAs, which act as decoys binding to miRNAs, leading to more cell differentiation

  • circular RNA (circRNA) compete for miRNA binding

CPE (cytoplasmic polyadenylation element) sequence

1. CPEB protein recognizes mRNAs not being translated

2. PARN shortens the poly-A tail

3. shortened tail is less bound to PABP

4. Makin binds to a cap binding protein (eIF4E), blocking its interaction with eIF4G which is important for translation initiation

reversal of CPE-containing mRNAs

1. CPEB is phosphorylated by kinases

2. PARN is released

3. cytoplasmic poly-A polymerase lengthens the poly-A tail

4. poly-A tail is bound by additional PABPs which displace Maskin allowing for translation initiation

ZPB1 (zip code binding protein 1)

blocks transition initiation by preventing the association of the large subunit of the ribosome until mRNA is in the correct location

Phosphatases

enzymes that remove phosphate groups from proteins

Ubiquitin

a protein that targets others for degradation

1. covalently attaches to target protein’s lysine side chain through ubiquination

2. proteasome recognizes tagged proteins, unwinds them, removes ubiquitin tags, breaks proteins into small peptides

Ubiquitin ligases

enzymes that bind to specific proteins and catalyze the processive addition of ubiquitin residues

p53

• activates transcription of genes that encode proteins that stop the cell cycle and promote DNA repair

• bound to Mdm2 and tagged for degradation in normal cells, but Mdm2 is phosphorylated when the cell senses DNA damage