Genetics Chapter 23, 24, and 25

Retinoblastoma

- "two-hit" model for retinoblastoma - Retinoblastoma requires 2 mutations to occur - People with the inherited form have already received one mutation from one of their parents, it is likely that a second mutation will occur in one of the retinal cells at an early age, leading to disease - People with the noninherited form, must have 2 mutations in the same retinal cell to cause the disease: 2 rare events are much less likely to occur than a single event, therefore, the noninherited form occurs much later in life, and only rarely - Persons with hereditary retinoblastoma have inherited one defective copy: in non-tumorous cells of the body, they have one normal (functional) copy and one defective copy of rb, in retinal tumor cells, the normal rb gene has also suffered the second hit, rendering it defective

Genetic Variation Can Affect the Proper Dosage of a Specific Drug

- A patient's genotype is important in determining the proper dosage of certain drugs - Pharmacogenetics is the study or clinical testing of genetic variation that causes differing responses to drugs - The amount of a drug needed to treat a patient with a disease depends on: rate of transport from digestive tract, rate of transport into target cells, ability of the drug to affect target protein, ability of drug to be metabolized by liver, rate of excretion of drug from body - These events are affected by genetic variation of each patient - Many drugs are broken down in the liver by a family of related enzymes called cytochrome P450 - Ex. warfarin (coumadin), which is used clinically as an anticoagulant: metabolized by a cytochrome P450 enzyme designated CYP2C9, over 80 variants (SNPs) in the gene that encodes CYP2C9 in human populations - Variation in CYP2C9 activity results in 4 different levels of warfarin metabolism: ultrarapid, extensive, intermediate, and poor, genetic tests can help doctors predict the proper warfarin dosage for their patients - In the future, patients' genomes may be sequenced to improve diagnosis, proactive care, and the proper dosages of medications

Genetic Changes in Proto-Oncogenes Convert Them to Oncogenes

- A proto-oncogene is a normal cellular gene that can incur a mutation to become an oncogene - By studying proto-oncogenes, researchers have found that such cancer-causing mutations occur in 4 main ways: 1. Missense mutations 2. Gene amplifications 3. Chromosomal translocations 4. Viral integration

Inherited Forms of Cancers

- About 5% to 10% of all cancers involve inherited (germ-line) mutations: people who have inherited such mutations have a predisposition to develop cancer - Genetic testing exists for certain types of cancer - Most inherited forms of cancer involve a defect in tumor-suppressor genes - Tumor-suppressor genes: VHL, APC, p53, rb, BRCA-1, BRCA-2, NF1 - Oncogenes: RET - Predisposition for developing cancer is often the result of being heterozygous for one of these genes; one copy is a loss-of-function allele - Cancer results from loss of the functional (normal) copy: this is known as loss of heterozygosity (LOH), tends to be inherited in a dominant fashion, can result from a point mutation in the normal allele, can also occur if chromosome carrying the functional copy is lost

Proteome Much Larger than Genome

- Actual size of proteome difficult to determine - Larger size rooted in a number of cellular processes: alternative splicing, RNA editing, postranslational covalent modification - These processes increase the number of potential proteins in proteome

Autosomal Recessive Inheritance: Tay-Sachs Disease (TSD)

- Affected individuals appear healthy at birth, but then develop neurodegenerative symptoms at 4-6 months - cerebral degeneration, blindness and loss of motor function - TSD patients typically die at 3 or 4 years of age - TSD is 100 times more frequent in Ashkenazi Jewish populations than others - TSD is the result of a mutation in the gene that encodes the enzyme hexosaminidase A (hexA): HexA breaks down a category of lipids called Gm2-gangliosides - An excessive accumulation of this lipid in cells of the CNS causes the neurodegenerative symptoms - TSD is inherited in an autosomal recessive manner

Gene Knockout Collections

- Allow researchers to study gene function at genomic level - To determine function of genes in genome: one approach is to generate collection of organisms from a species each with one gene knocked out - The phenotype could indicate function - Could combine knockouts to study pathways - Many ways to generate collections of knockouts: transposable element jumps, CRISPR-Cas technology

RNA-Seq - Newer Method to Identify Expressed Genes

- Also used to study simultaneous transcription of many genes - Compares transcriptomes - set of all RNA molecules, including mRNAs and non-coding RNAs, that are transcribed in one cell or a population of cells - RNA-Seq used to compare transcription in: different cell types, healthy vs diseased cells, different stages of development and response to different environmental agents such as hormones or toxic chemicals.

Functional Cycle of the Ras Protein

- As a specific example, mutations that convert normal Ras into an oncogenic Ras either, decrease the GTPase activity of the Ras protein or increase the rate of exchange of bound GDP for GTP - this results in greater amounts of the active Ras/GTP complex: signaling pathway stays ON longer than normal

DNA Microarrays Used in Molecular Profiling of Tumors

- As an example, researchers did a microarray analysis of many patients with diffuse large B-cell lymphoma - Highly expressed genes are shown in red and low expression is in green - The cancer cells in some patients had an expression patterns that was similar to germinal center B cells; other patients' cancer cells were like activated B cells - highly expressed: red, low expression: green

Genetic Changes in Cancer

- Both genetic and epigenetic changes can promote cancerous growth - Cancer-causing mutations affect genes in 2 categories: Oncogene - a mutant gene that is over-expressed and contributes to cancerous growth Tumor-suppressor gene - gene that prevents cancer; a loss-of-function mutation in this type of gene allows cancerous growth to occur

Viral Integration

- Can convert proto-oncogenes into oncogenes - Certain viruses integrate into host DNA as part of their life cycle - This can cause activation of a cellular proto-oncogene: direct transcription from viral promoter, activation of cellular promoter by viral enhancer - Ex. in certain lymphomas in birds, avian leukosis virus is integrated next to the c-myc gene, enhancing its level of transcription

Chromatin Immunoprecipitation (ChIP)

- Can determine if proteins can bind to a particular region of DNA in the chromatin of living cells - Proteins in living cells that bind to specific sites in the DNA are covalently cross-linked to those DNA sites using formaldehyde - Cells are lysed and DNA broken into small pieces - Antibody is used to precipitate the protein of interest - DNA is chemically freed from the cross-links - DNA is amplified with PCR - Sequence of DNA is identified directly or by using it as a probe on a microarray (ChIP-chip assay) - To identify the DNA bound by the protein, researchers can use: PCR or DNA microarray

Analysis of DNA Microarrays

- Cells respond to environmental changes via the coordinate regulation of their genes, some genes are turned on while others are turned off - Microarrays make it possible to study the expression of the whole genome under different environmental conditions

Role of Epigenetics in Cancer

- Correlation coefficient: compares 2 variables to see if they are related to each other - Association: suggests that changes in the 2 variables follow a pattern - Many associations between epigenetics and a human disease have been found; these associations can be due to: 1. The epigenetic changes directly contribute to the disease symptoms 2. The disease symptoms may arise first, and then they cause subsequent epigenetic changes to happen 3. The association is indirect because a third factor is involved; a third factor causes both the disease and epigenetic changes even though those epigenetic changes do not contribute to the disease.

DNA Microarrays

- DNA fragments on microarray can be either: amplified by PCR and then spotted onto the microarray, or synthesized directly on the microarray itself - A single slide contains tens of thousands of different spots in an area the size of a postage stamp: the relative location of each spot is known - Making DNA microarrays involves spotting technologies similar to the way an inkjet printer works - Once a DNA microarray has been made, ti is used as a hybridization tool

Abnormalities in Chromatin Modification Are Common in Cancer Cells

- DNA methylation: ex. hypermethylation-an abnormally high level of methylation, typically at CpG islands, is often observed, may promote cancer by inhibiting the expression of tumor-suppressor genes - Covalent modification of histones: depending on the specific type of modification, such changes could increase the expression of oncogenes or inhibit the expression of tumor suppressor genes - Chromatin remodeling: abnormalities in the locations of nucleosomes have been frequently found in cancer cells

Microarray Can Identify Genes that are Transcribed

- DNA microarrays (gene chips): makes it possible to monitor thousands of genes simultaneously - DNA microarray is a small silica, glass or plastic slide that is dotted with many sequences of DNA - Each of these sequences corresponds to a known gene, these fragments are made synthetically, and act as probes to identify genes that are transcribed

Maintenance of Genome Integrity

- DNA repair enzymes are a second class of proteins involved with genome maintenance: are inactivated in some cancers - In these cancers, it is more likely for a cell to accumulate mutations that: create an oncogene, eliminate the function of a tumor-suppressor gene - Ex. defects in the nucleotide excision repair process are responsible for the disease called xeroderma pigmentosum, predisposition to developing skin cancer

Overview of Cancer

- Disease characterized by uncontrolled cell division - More than 100 kinds of human cancers are known: - classified according to the type of cell that has become cancerous - ~1.5 million Americans are diagnosed with cancer each year - Over 500,000 will die from the disease

Cancer

- Disease that occurs in multicellular organisms; characterized by uncontrolled cell division - Second leading cause of death worldwide - Higher predisposition to develop cancer is inherited in around 10% of cancer - 80% of human cancers are related to exposure to carcinogens - environmental agents that increase the likelihood of developing cancer

Genetic Basis For a Human Disease

- Diseases are caused, at least in part, by genes 1. When an individual exhibits a disease, the disorder is more likely to occur in genetic relatives than in the general population 2. Identical twins share the disease more often than fraternal twins: - identical twins are called monozygotic (MZ) twins, formed from the same sperm and egg - fraternal twins are called dizygotic (DZ) twins: formed from separate pairs of sperm and egg - Geneticists evaluate the concordance of the disorder, which is the degree to which it is inherited: concordance refers to the percentage of twin pairs in which both twins exhibit the disorder or trait - Identical twins - concordance is 1 - Actual concordance is often less than theoretical values 3. The disease does not spread to individuals sharing similar environmental situations 4. Different populations tend to have different frequencies of the disease 5. The disease tends to develop at a characteristic age: many genetic disorders exhibit a specific age of onset 6. The human disorder resembles a genetic disorder that has a genetic basis in another mammal 7. A correlation is observed between a disease and a mutant human gene or a chromosomal alteration

Autosomal Recessive Human Disorders

- Disorders that involve defective enzymes typically have an autosomal recessive mode of inheritance - The heterozygote carries has 50% of the functional enzyme: this is sufficient for a healthy (unaffected) phenotype - Hundreds of genetic diseases are inherited this way, in many cases, the mutant genes responsible have been identified and characterized - Examples: Albinism-tyrosinase, cystic fibrosis-CFTR, a chloride transporter, phenylketonuria (PKU)- phenylalanine hydroxylase, sickle cell disease- beta globin

Functional Genomics

- Elucidate the roles of genetic sequences in a given species: in most cases, it aims to understand gene function - Entire collection of proteins that organism can make - proteome - Proteomics: understand functional roles of proteins of a species, aims to understand the interplay among many different proteins - Bioinformatics: analysis of biological information using methematical/computational approach

Creating Computer Data Files

- Entering data into the computer file is done: manually, or usually by instruments - Genetic sequence can be analyzed in many ways: 1. Does sequence contain a gene? 2. Where are functional sequences, like promoters and splice sites? 3. Does the sequence encode a polypeptide? If so, what is the amino acid sequence of that polypeptide 4. Is the sequence homologous to other sequences? 5. What is the evolutionary relationship between 2 or more genetic sequences?

Proteomics

- Examines functional roles of proteins that a species can make: entire collection of proteins in a proteome - RNA-Seq and DNA microarrays provide insight about the transcription of genes but may not provide an accurate measure of protein abundance - Genomic insights are often followed up with research that involves protein analysis directly

Approaches to Identify Genes

- Gene prediction: the process of identifying regions of genomic DNA that encode genes: protein-encoding genes, genes for non-coding RNAs - Search by Signal: program tries to locate an organization of known sequence elements that are normally found within a gene (promoter, start/stop codons, etc.) - Search by Content: program tries to identify sequences that differ significantly from a random distribution due t codon bias within protein-encoding genes

Maintenance of Genome Integrity

- Genome Maintenance: refers to the mechanisms that prevent mutations or prevent mutant cells from surviving or dividing: these proteins can detect abnormalities such as DNA breaks and improperly segregated chromosomes, many of these proteins are called checkpoint proteins (they check the integrity of the genome and prevent cells from progressing past a certain point of the cell cycle if there is damage - Cyclins and cyclin-dependent kinases (Cdks) are responsible for advancing a cell in the cell cycle

Determining Gene Sequence from Peptide Sequence

- Genome sequences of species may have been determined - Computer software allows amino acid sequences obtained by mass spec to be used to search database of protein sequences - Computer program may locate a match between identified sequence and a protein within a species - Mass spectrometry can also be used to identify protein covalent modifications: ex. the mass of a phosphorylated protein increases by the mass of a phosphate

Cell Cycle is Regulated by Growth Factors

- Growth factors bind to cell surface receptors and initiate a cascade of cellular events leading ultimately to cell division - Epidermal growth factor (EGF) is a growth hormone in vertebrates - Oncogenes often encode proteins that function in cell signaling pathways related to cell division: growth factor receptors, intracellular signaling proteins, transcription factors

Autosomal Dominant Disorders

- Haploinsufficiency: heterozygote has 50% of the functional protein, this is not sufficient for a healthy (unaffected) phenotype - Gain-of-function mutations: mutation changes protein so it gains a new function - Dominant negative mutations: the mutant gene product acts antagonistically to the wild-type gene product - Examples: aniridia, achondroplasia, marfan syndrome, familial hypercholesterolemia

Multiple Sequence Alignment

- Homologous genes can be compared to each other in this way - Globin Gene Family: in humans, 9 paralogs are functionally expressed, the 9 paralogs fall into 2 categories: alpha and beta - Each hemoglobin is composed of 2 alpha chains and 2 beta chains - The composition of hemoglobin changes during the course of development - Comparing sequences of hemoglobin chains can give insight into their structure and function: the sequences of the globin polypeptides can be compared using multiple sequence alignment - A conserved site is a site that is identical or similar across multiple homologs: conserved sites tend to be functionally important

Cell Cycle Checkpoints

- If DNA damage is found, these checkpoint proteins can prevent the formation of active cyclin/Cdk complexes - The M checkpoint is monitored by proteins that can sense if a chromosome is not correctly attached to the spindle apparatus - Mutations that eliminate checkpoint proteins make it more likely that mutant cells will be able to divide

Open Reading Frames

- In a DNA sequence, the reading of codons could begin with the first, second or third nucleotide, these are called reading frame 1, 2 and 3, respectively - An open reading frame (ORF) is a nucleotide sequence that does not contain any stop codons: in prokaryotes, long ORFs are contained within the chromosomal gene sequences, in eukaryotes, however, the chromosomal coding sequences may be interrupted by introns - Computer program can translate a genomic DNA sequence in all 3 reading frames, seeking to locate a long ORF - Reading frames can also proceed from right to left. Thus, 6 reading frames are possible in a newly discovered sequence

Searching Databases for Homologous Sequences

- In general, a strong correlation is typically found between homology and function - Homology between genetic sequences can be identified by computer programs and databases, powerful tool for predicting the function of genetic sequences - BLAST: basic local alignment search tool, relationship between the query sequence and each matching sequence is given an E-value (Expect value), represents the number of times that the match or a better one would be expected to occur purely by random chance in the entire database

Inheritance Patterns of Human Diseases

- Inheritance patterns of human diseases may be determined via pedigree analysis - The pattern of inheritance of a human disorder that is caused by a mutation in a single gene can be deduced by analyzing human pedigrees

Gene Amplification

- Many human cancers are associated with the amplification of particular oncogenes - Increase in copy number: expected to increase the amount of protein - Examples: c-myc in a leukemia cell line, N-myc in neuroblastomas, erbB-2 in breast carcinomas

Genetic Disorders Exhibit Locus Heterogeneity

- Many inherited diseases exhibit locus heterogeneity - Locus heterogeneity refers to the phenomenon that a disease can be caused by mutations in 2 or more different genes - Consider hemophilia: blood clotting involves a cellular cascade that involves several different proteins, therefore, a defect in any of these proteins can cause the disease - Hemophilia A is a defect in (clotting) Factor VIII - Hemophilia B is caused by a defect in Factor IX: both are X-linked recessive disorders - Hemophilia C is caused by a defect in Factor XI, found on chromosome 4 (autosomal recessive)

Cancer Involved Multiple Genetic Changes Leading to Malignancy

- Most forms of cancer involve multiple genetic changes leading to malignancy - Many cancers begin with a benign mutation that, with time and more mutations, leads to malignancy: furthermore, a malignancy can continue to accumulate genetic changes that make it even more difficult to treat

Cell Division Pathways

- Oncogenes have gain-of-function mutations that may affect proteins that are directly involved with cell division - Eukaryotic cells advance through the cell cycle: G1, S, G2, M phases - Restriction point in G1 - cell becomes committed to advance through the rest of the cell cycle

Oncogenes

- Oncogenes promote abnormal cell growth - Proto-oncogenes: normal cellular genes that can be mutated into an oncogene - Expression becomes abnormally active: this is a gain-of-function mutation - This typically can occur in 3 ways 1. The amount of encoded protein is greatly increased 2. A change occurs in the structure of the encoded protein that causes it to be overly active 3. The encoded protein is expressed in a cell type where it is not normally expressed

Advantages of RNA-Seq

- RNA-Seq has several advantages over microarrays - More accurate at quantifying the amount of each RNA transcript - Superior at detecting RNA transcripts that are in low abundance - Identifies the exact boundaries between exons and introns; identifies new splice variants - Identifies the 5' and 3' ends of RNA transcripts

Cancer Treatments May be Aimed at Epigenetic Changes

- Researchers are actively investigating drugs that may inhibit cancer cells by affecting either DNA methylation or covalent histone modifications - Ex. inhibitors of DNA methyltransferase are being developed to treat certain forms of cancer including leukemia - In some cases, improvement in patients with leukemia has been associated wit ha decrease in DNA methylation - One possibility is that the lower level of DNA methylation has reversed the inhibition of tumor suppressor genes

Sequence and Pattern Recognition

- Sequence Recognition: program has information that a specific sequence of symbols has a specialized meaning, with information from a dictionary, the first program can identify sequences of letters that make words - Pattern Recognition: does not rely on specialized sequence information, rather certain programs look for a pattern of symbols that can occur within any group of symbol arrangements

Genetic Changes Leading to Cancer

- Some genes affect growth directly; others may enable metastasis which allows expansion to new locations, giving the cells a growth advantage - Estimated that 300 different genes may play a role in the development of human cancer: over 1% of our genes, mutations in ncRNAs are associated with some cancers - Chromosomal abnormalities are often associated with cancer: missing chromosomes carried a tumor suppressor gene, duplicated chromosomes may over-express proto-oncogenes, translocations fuse or disrupt genes

Chromosomal Translocations

- Specific chromosomal translocations occur in certain types of tumors - Chronic myelogenous leukemia was correlated with a shortened chromosome 22: called Philadelphia chromosome after the city where it was discovered, a translocation between chromosomes 9 and 22, this puts the proto-oncogene abI under the control of the bcr promoter, which is active in white blood cells, leading to leukemia - Translocation creates an oncogene that encodes an abnormal fusion protein - The drug imatinib mesylate (Gleevec) fits into the active site of ABL, preventing ATP binding - Almost 90% of CML patients respond to treatment with this drug

BLAST

- Starts with a genetic sequence (the query sequence) and then located homologous sequences in a large database - Homology among protein sequences is easier to identify than is DNA sequence homology - Small E-values indicates that similarity is unlikely to be due to random events; the genes are likely to be homologous, E-values depend on length of query, number of gaps in alignment, and database size

Molecular Function of Rb

- The Rb protein prevents the proliferation of cancer cells - Rb is phosphorylated when the cell is about to divide: dissociates from E2F, a transcription factor that activates genes for cell cycle progression - When bot hcopies of the Rb protein are defective, the E2F protein is always active, this leads to uncontrolled cell division

Bioinformatics

- The computer has become an important toll in genetic studies: the marriage between genetics and biocomputing has yielded an important branch of science: bioinformatics - Computer analysis of genetic sequences usually relies on 3 basic components: a computer, a computer program, some type of data

Hemophilia

- The major symptom is that blood cannot clot properly when a wound occurs - For hemophiliacs, common accidental injuries pose a threat of severe internal or external bleeding - Hemophilia A (also called classical hemophilia) is caused be a defect in an X-linked gene that encodes a clotting protein called Factor VIII - This disease has also been called the "royal disease" because it has affected many members of European royal families - The inheritance pattern of Hemophilia A in the royal families of Europe

Autosomal Dominant Inheritance: Huntington Disease (HD)

- The major symptom of the disease is the degeneration of certain types of neurons in the brain - This leads to personality changes, dementia, and early death (usually in middle age) - HD is the result of the mutation in a gene that encodes a protein named huntingtin: the mutation adds a polyglutamine tract to the protein - This causes an aggregation of the protein in neurons - HD is inherited in an autosomal dominant manner

p53 Is a Master Tumor-Suppressor Gene

- The p53 gene was the second tumor-suppressor gene discovered - About 50% of all human cancers are associated with defects in the p53 gene - A primary role for the p53 protein is to determine if a cell has incurred DNA damage: if so, p53 will promote 3 types of cellular pathways to prevent the division of cells with damaged DNA 1. Activates genes that promote DNA repair - prevents accumulation of mutations 2. Activates genes that arrest cell division and generally repress and generally repress other genes that are required for cell division: stimulates the expression of p21, which inhibits the formation of cyclin/CDK complexes that are needed to advance the cell cycle 3. Activates genes that promote apoptosis, or programmed cell death - involves cell shrinkage, chromatin condensation, and DNA degradation, facilitated by proteases known as caspases, which digest selected cellular proteins, cell is broken down into small vesicles that are eventually phagocytosized by cells of the immune system

Inheritance Patterns of Genetic Diseases

- The study of human genetic diseases provides insights regarding our traits: ex. by analyzing people with hemophilia, researchers have identified genes that participate in blood clotting - Thousands of human diseases have a genetic basis

Personalized Medicine

- The use of a patient's genotype to select a treatment suited for that patient - Expected to become increasingly important - Can be used to choose the best treatment for cancer or to determine best drug dosage

X-linked Recessive Inheritance

- This type of inheritance poses a special problem for males - Males have only a single copy of X-linked genes: hemizygous - A female heterozygous for an X-linked recessive allele will pass this trait to half of her sons

Tumor-Suppressor Genes

- Tumor-suppressor genes can no longer inhibit cancer when their function is lost - During the past 3 decades, researchers have identified many tumor-supressor genes 1. Proteins that negatively regulate cell division: ex. Rb negatively regulates E2F 2. Proteins that maintain genome integrity: prevent damaged cells from dividing (checkpoint proteins), DNA repair

Tumor-Suppressor Genes

- Tumor-suppressor genes play a role in preventing the proliferation of cancer cells - Tumor-suppressor genes prevent the proliferation of cancer cells: if they are inactivated by mutation, it becomes more likely that cancer will occur - The first identification of a human tumor-suppressor gene involved studies of retinoblastoma: a tumor of the retina of the eye, there are 2 types of retinoblastoma 1. Inherited, which usually occurs in childhood 2. Non-inherited, which occurs later in life

Personalized Medicine

- Use of information about a patient's genotype and other clinical data in order to select a medication, therapy, or preventative measure that is specifically suited to that patient - Thousands of genetic diseases afflict people, many of these are the direct result of a mutation in one gene - Genes also play roles in the development of diseases that have a complex pattern of inheritance, involves multiple genes, ex. diabetes, asthma, mental illness.

Mass Spectrometry

- Used to identify proteins - Want to correlate a given spot on 2-D gel with a particular protein - Spot is cut out from gel - Protein is eluted from gel in purified form - Amino acid sequence of protein is revealed via tandem mass spectrometry - 2 spectrometers used: first measures mass of given peptide (generated from protein digestion), second analyzes peptide after is has been digested into smaller fragments, one amino acid at a time

2-D Gel Electrophoresis

- Used to separate a mixture of different proteins - An given cell only produces subset of proteins in its proteome - Subset depends on the: cell type, stage of development, environmental conditions - 2-D gel electrophoresis: separation technique that can distinguish hundreds or thousands of different proteins in a cell extract - Technique involves 2 different gel electrophoresis experiments: first separates by pH/charge interactions (isoelectric focusing), second separates by size (mass) - Specific spots may be of special interest: proteins that are very abundant in cell (may be important for cell's structure or function), spots present in only given circumstances (cells exposed to a hormone vs. not), spots present only in abnormal cells (common in cancer cells)

Protein Microarrays

- Used to study protein expression and function - Technology used to make DNA microarrays is being applied to make protein microarrays: proteins rather than DNA are spotted onto a slide - Harder to develop protein microarrays than DNA microarrays - Proteins are much more easily damaged by the manipulations that occur during microarray formation - Synthesis and purification or proteins tend to be more time-consuming compared to DNA

Molecular Profiling

- Using various methods to understand the molecular changes behind a disease - In cancer, molecular profiling allows - differentiation between cancers which might look similar under the microscope, predicting drugs that will target altered genes (tamoxifen can be used for tumors over-expressing estrogen receptor

Certain Viruses Can Cause Cancer

- Viruses can infect normal laboratory-grown cells and convert them into malignant cells - The Rous sarcoma virus (RSV), was isolated from chicken sarcomas by Peyton Rous in 1911 - This virus carries a gene designated src (sarcoma, the type of cancer it causes) - Up to 15% of human cancers are estimated to be associated with viruses

Epigenetic Changes May Arise Because of Mutations in Genes That Encode Chromatin-Modifying Proteins

- What causes chromatin modifications to become abnormal and promote cancer 1. Mutations may occur in genes that encode chromatin-modifying proteins - Ex. mutation of DNA methyltransferase in acute myeloid leukemia 2. Environmental agents may alter the functions of chromatin modifying proteins. - Ex. polycyclic aromatic hydrocarbons in tobacco smoke, automobile exhaust, charbroiled food in lung, breast, stomach, and skin cancer

Orthologs and Paralogs

- When 2 homologous genes are found in different species, these genes are termed orthologs - When 2 homologous genes are found in a single organism, these genes are termed paralogs - A gene family consists of 2 or more copies of homologous genes within the genome of a single organism - Homologous genes are often analyzed and identified using computer data bases

X-linked Dominant Inheritance

- X-linked dominant inheritance is rare - Characteristics of such disorders: - males are often more severely affected (females may be less affected due to wild-type copy on the other X chromosome), - females are more likely to exhibit the trait when it is lethal to males - affected mothers have a 50% chance of passing the trait to daughters - Ex. Vitamin D-resistant rickets, rett syndrome, aicardi syndrome, incontinentia pigmenti

Homologous Genes

- derived from the same ancestral gene - When comparing genetic sequences, researchers sometimes find 2 or more similar sequences - The 2 sequences may be similar because the genes are homologous to each other: these are orthologs - They are found in different species and have been derived from the same ancestral gene

Missense Mutation

- the human genome contains 4 different but evolutionary related ras genes: H-ras, N-ras, K-4a-ras, and K-4b-ras - Missense mutants in these genes are associated with certain cancers - Ex. Missense mutation in H-ras changes in 12th amino acid from a glycine to a valine, converts H-ras into an oncogene - Experimentally, chemical carcinogens have been shown to cause these missense mutations and thereby lead to cancer

Tumor-Suppressor Genes Can Be Silenced a Variety of Ways

1. A mutation in the tumor-suppressor gene itself: the promoter could be inactivated, an early stop codon could be introduced in the coding sequence 2. Aneuploidy: chromosome loss may contribute to the progression of cancer if the lost chromosome carries one or some tumor-suppressor genes 3. Epigenetic changes: discussed later in the chapter

Alterations that Affect the Proteome

1. Alternative Splicing - Most important alteration that occurs in eukaryotes - A single pre-mRNA is spliced into more than one version - Splicing is often cell specific or related to environmental conditions 2. RNA Editing - Much less common than alternative splicing - Leads to changes in the coding sequence of mRNA after the mRNA is made 3. Postranslational Covalent Modification - Irreversible changes to produce functional protein, proteolytic processing: attachment of prosthetic groups, sugars or lipids - Reversible changes that transiently affect function of protein: phosphorylation; acetylation; methylation

Types of Protein Microarrays

1. Antibody Microarrays: consist of collection of antibodies that recognize short peptide sequences, used to assess level of protein expression 2. Functional Microarrays: consist of many different cellular proteins, used to probe function of proteins: for instance, are certain proteins phosphorylated by a set of kinases

Identification Strategies

1. Locate specialized sequences (sequence elements) within a very long sequence: sequence elements are predefined and are contained explicitly within the computer program, a genetic sequence within a particular function is called a sequence element or sequence motif 2. Locate an organization of sequences or sequence elements 3. Locate a pattern of sequences

Characteristics of Cancer

1. Most cancers originate in a single cell: in this regard, a cancerous growth is considered to be clonal 2. At the cellular and genetic levels, cancer is usually a multistep process: it begins wit ha precancerous genetic change (that is, a benign growth), following additional genetic changes, it progresses to cancerous cell growth 3. Once cellular growth has become malignant, the cells are: invasive - they can invade healthy tissues, metastatic - they can migrate to other parts of the body

Tandem Mass Spectrometry

1. Peptides mixed with an organic acid and dried onto a metal slide 2. Sample is subjected to a laser beam: peptides become ejected as an ionized gas where the peptide contains one or more positive charges 3. The charged peptides are then accelerated via an electric field and fly toward a detector: the time they spend in flight is determined by their mass and net charge and reveals the mass of the peptide, each amino acid has its own characteristic mass

Features of X-linked Recessive Inheritance

3 Common Features of X-linked Recessive Inheritance are as Follows: 1. Males are much more likely to exhibit the trait 2. The mothers of affected males often have brothers or fathers who are affected with the same trait 3. The daughters of affected males will produce (on average) 50% affected sons - Ex. duchenne muscular dystrophy, hemophilia A, hemophilia B, androgen insensitivity syndrome

Features of Autosomal Dominant Inheritance

Five common features of autosomal dominant inheritance are as follows: 1. An affected offspring usually has one or both affected parents: can be altered by reduced penetrance 2. An affected individual with only one affected parent is expected to produce (on average) 50% affected offspring 3. 2 affected, heterozygous individuals will have (on average) 25% unaffected offspring 4. The trait occurs with the same frequency in both sexes 5. For most dominant disease-causing alleles, the homozygote is more severely affected with the disorder

Features of Autosomal Recessive Inheritance

Four common features of autosomal recessive inheritance are: 1. Frequently, an affected offspring will have 2 unaffected parents 2. When 2 unaffected heterozygotes have children, the percentage of affected children is (on average) 25% 3. 2 affected individuals will have 100% affected children 4. The trait occurs with the same frequency in both sexes