Final Exam Notes Biology PDF

The One-Gene, One-enzyme hypothesis ○ Srb and Horowitz tested hypothesis by studying three-step metabolic pathway that produces arginine: Performed genetic screen by growing mold cells on medium that lacked arginine Cells that die...

The One-Gene, One-enzyme hypothesis ○ Srb and Horowitz tested hypothesis by studying three-step metabolic pathway that produces arginine: Performed genetic screen by growing mold cells on medium that lacked arginine Cells that died came from a colony that was missing an enzyme from that pathway Each gene contains information to make an enzyme The central dogma of molecular biology ○ Francis Crick proposed sequence of bases in DNA acted as a code: DNA is an information storage molecule Different combinations of bases specify the 20 amino acids ○ Particular stretch of DNA (a gene) specifies amino acid sequence of one protein ○ Information in sequence of DNA is not directly translated into amino acid sequence of proteins RNA as the Intermediary between between Genes and proteins ○ Messenger RNA (mRNA): Found to carry information from DNA to site of protein synthesis ○ Enzyme RNA polymerase synthesize RNA: Uses DNA strand as template Copies code by matching complementary nucleotides The roles of transcription and translation ○ Transcription - process of using DNA template to make complementary RNA Making a copy of information ○ Translation - process of using information in mRNA to synthesize proteins: Interprets nucleotide “language” to amino acids The central dogma explains the relationship between genotype and phenotype The genetic code ○ Genetic code specifies how a sequence of nucleotides codes for a sequence of amino acids ○ Three-base code is the least that could specify the 20 amino acids ○ Could code for 4 x 4 x 4 = 64 different amino acids ○ Three-base code is known as a triplet code The genetic code consists of three-letter “words” ○  Analyzing the code ○ The genetic code is: Redundant - all but two amino acids are encoded by more than one codon Unambiguous - one codon never codes for more than one amino acid Non-overlapping - codons are read one at a time Nearly universal - All codons specify the same amino acids in all organisms (with a few minor exceptions) Conservative - if several codons specify the same amino acid, the first two bases are usually identical The genetic code can predict amino acid sequences ○ What are the types and consequences of mutation? ○ Mutation - any permanent change in an organism's DNA Modification in cell’s information archive Change in its genotype New alleles ○ There are different types of mutations: Point mutations result from one or a small number of base changes Chromosome-level mutations are larger in scale Unrepaired mistakes in DNA synthesis lead to point mutations ○ Types of point mutations  ○ Missense mutations change an amino acid in protein ○ Silent mutations do not change amino acid sequence due to redundancy in the code ○ Frameshift mutations shift reading frame, altering meaning of all subsequent codons ○ Nonsense mutations change codon that specifies an amino acid into stop codon Central Dogma ○ Understand the flow of genetic information ○ Define transcription and translation and location these processes occur in the cell ○ Transcription DNA to RNA = Transcription The synthesis of RNA under the direction of DNA Produces messenger RNA (mRNA) and other RNAs (ribosomal RNAs and transfer RNA) ○ Translation RNA to Protein = Translation Occurs on the ribosomes in the cytosol The synthesis of a polypeptide which occurs under the direction of mRNA Transcription ○ Difference between the coding and noncoding strand ○ Understand and describe the general role of RNA polymerase during transcription ○ Distinguish the process of transcription in bacteria and eukaryotes ○ Only one strand of DNA is the template strand Sense (Non-Template/coding) DNA strand (5’-3’) The DNA strand that does not serve as the template from which mRNA is made (not transcribed into mRNA) Antisense (Template/non-coding) DNA strand (3’-5’) The DNA strand that serves as the template from which mRNA is made (transcribed into mRNA) ○ Transcription is the synthesis of RNA from a DNA template Bacteria have one RNA polymerase Eukaryotes have at least three distinct types: RNA polymerase I, II, III Unlike DNA polymerase, RNA polymerase do not require a primer to begin transcription ○ RNA polymerase synthesize an RNA version of the instructions stored in DNA Promoter: Region of DNA that includes the site where RNA polymerase binds, and transcription starts Regulatory protein (SIgma): DNA-binding proteins that will bind to specific DNA sequence at the promoter Helps RNA polymerase bind to promoter to start transcription ○ Bacterial Promoter  The -10 box = ~10 bases upstream of the transcription start site Downstream means in the same direction that RNA polymerase moves Upstream is the opposite direction The -35 box = about 35 bases upstream of the transcription start site ○ Sigma orients the RNA polymerase on DNA during the initiation of transcription ○ Transcription in bacteria terminates when an RNA Hairpin forms ○ Transcription in eukaryotes Three RNA polymerases -TATA box = -30 (upstream of start site) General transcription factors recognize promoters, rather than sigma proteins At termination, a poly(A) signal is transcribed rather than a hairpin and the RNA downstream is cut Transcription occurs in the nucleus, and translation occurs in the cytoplasm RNA processing ○ Understanding the difference between introns and exons ○ Describe the role of spliceosomes in the formation of mRNA ○ Explain how mRNA is processed in eukaryotes (5’ cap, and 3’ poly A- tail) ○ RNA processing in eukaryotes In bacteria, transcription produces fully functional RNAs In eukaryotes, the initial product of transcription is an immature primary transcript or pre-mRNA. Primary transcripts must undergo RNA processing before they can be translated RNA processing = RNA splicing and adding 5’ cap and 3’ tail ○ Introns are not present in the final mRNA = does not code for protein (introns are removed, the spacers in between) ○ Exons are present in the final mRNA = codes for protein (exons are expressed) ○ During RNA splicing, introns are cut out of the primary transcript ○ Formation of spliceosome Small nuclear ribonucleoproteins (snRNPs) form a complex called spliceosome ○ During splicing, introns are cut out of the primary transcript ○ In eukaryotes, a cap and a tail are added to mRNAs Modified guanine nucleotide are attached to the 5’ end of the mRNA strand, called a “5’ cap” Protects the mRNA strand from degradation due to cytoplasmic enzymes Functions as an attachment indicator for ribosomes A long sequence of 100-250 adenine nucleotides is added to the 3’ end, called a “poly (A) tail” protects the mRNA strand from degradation due to cytoplasmic enzymes Functions as an attachment indicator for ribosomes  Facilitates export of the mRNA strand to the cytoplasm Introduction to translation ○ Recognize the key players in translation ○ Describe the structure and role of tRNA ○ Understand the wobble pairing hypothesis of how one tRNA can pair with more than one codon ○ Key players of translation The sequence of mRNA bases is converted to an amino acid sequence Key players of translation mRNA tRNA Amino acid aminoacyl-tRNA synthetase ribosomes ○ Transcription and translation can be coupled, or occur simultaneously, in bacteria Ribosomes are the site of protein synthesis ○ Adapter molecules hold amino acids and interact with mRNA codons The adapter molecule used in translation is called transfer RNA (tRNA) An aminoacyl tRNA is a tRNA linked to its amino acid Amino acids are transferred from tRNAs to a growing polypeptide ○ Structure of tRNAs tRNA form secondary structures by folding into a stem-and-loop The loop at the opposite end contains the anticodon Has a sequence of three nucleotides Can base-pair with the mRNA codon CCA sequence at the 3’ end is the amino acid binding site ○ How are amino acids attached to tRNAs? ATP is required to attach tRNA to an amino acid aminoacyl-tRNA synthetases “charge” the tRNA: Catalyze the addition of amino acids to tRNAs There are 20 amino acids: Each has a different aminoacyl tRNA synthetase For each amino acid, there is one or more tRNAs ○ How many tRNAs are there? There are 61 different codons but only about 40 tRNAs in most cells Wobble pairing: the anticodon’s third position can form a nonstandard base pair One tRNA is able to read more than one codon Ribosome ○ Understand the structure of ribosomes ○ The three tRNA binding sites on the ribosome ○ Ribosome structure Ribosomes contain many proteins and ribosomal RNA (rRNA) Ribosomes can be separated into two subunits:  The small subunit holds the mRNA in place The large subunit is where peptide bonds form Ribosomes contains three tRNA binding sites ○ The tRNAs fit into three sites in the ribosome, bound to corresponding mRNA codons: A site (acceptor or aminoacyl) - tRNA carries an amino acid P site (peptidyl) - holds growing peptide chain E site (exit) - tRNAs without amino acids exit the ribosome Translation ○ Describe the three steps in translation ○ What happens in each of these steps and who are the key players ○ Step 1 - Initiation The initiation phase of translation begins near the AUG start codon ○ Step 2 - Elongation Translocation occurs when the ribosome slides one codon toward the 3’ end of the mRNA. Elongation factors help move the ribosome ○ Step 3 - Termination Gene expression ○ Most proteins go through an extensive series of processing steps before they are functional: called post-translational modification ○ Molecular chaperones guide and speed up protein folding ○ Sugars or lipid groups may be added to proteins ○ Enzymes may add a phosphate group to a protein to alter its activity Gene regulation mechanism ○ Gene expression is a multistep process when information encoded on genes is converted into an active product (protein) ○ Cells are extremely selective about which genes are expressed and in what amount. ○ A cell does not express all of its genes all of the time DNA→mRNA→protein→activated protein ○ Bacterial operons Operon - a functioning unit of DNA containing a cluster of genes under the control of a single promoter 2 types of operons Inducible - operon is turned ON by substrate: catabolic-operons - enzymes needed to metabolize a nutrient are produced when needed. Repressible - genes in a series are turned OFF by the product synthesized; anabolic operon - enzymes used to synthesize an amino acid stop being produced when they are not needed. ○ Arginine operon ○ 1) Transcriptional control - mRNA only made for proteins needed DNA→mRNA→protein→activated protein ○ 2) Translational control - not all mRNAs are translated  DNA→mRNA→protein→activated protein ○ 3) Post-translational control - proteins must be activated by chemical modification DNA→mRNA→protein→activated protein Gene expression in bacteria can be regulated at three levels ○ Slow response; conserves resources ○ Fast response; uses most resources Lactose metabolism in E. Coli. requires two proteins ○ Lactose is a disaccharide (glucose + galactose) ○ beta-glycosidic linkage ○ Galactoside permease transports lactose into the cell ○ beta-galactosidase is the enzyme that breaks the beta-glycosidic linkage Metabolizing lactose overview ○ Transcription of a gene regulated by a negative control ○ Ex. Turning light switch off Transcription of a gene regulated by a positive control  ○ ex. Turning light switch on A gene needed to regulate lactose metabolism ○ Research objectives: To find genes that code for beta-galactosidase or galactoside permease Or to find genes that code for regulators of genes that code for these proteins ○ Monod and Jacob isolated and analyzed E. Coli. mutants that could not metabolize Three classes of lactose metabolism mutants ○ lacZ-mutants = could not cleave lactose because they lack functional beta- galactosidase ○ lacY-mutants = do not accumulate lactose in their cells because they lack galactoside permease ○ lacI-MUTANTS = mutants produce beta-galactosidase and galactoside permease even when lactose is absent Called constitutive mutants Have a defeat in gene regulation Transcription regulation in presence of a repressor ○ The lacZ and lacY genes are under negative control ○ The lacI gene codes for a repressor ○ Binds on or near the lacZ and lacY promoter Transcription regulation in presence of lactose ○ Lactose induces transcription by removing the repressor Transcription regulation in presence of mutant lacI- gene ○ lacI- mutant have no repressor ○ lacZ and lacY genes are constitutively expressed with or without lactose The Lac operon ○ Group of genes (lacZ, lacY, lacA) involved in lactose metabolism is termed lac operon Operon = set of coordinately regulated bacterial genes that are transcribed together into one mRNA (polycistronic mRNA) ○ A fourth gene, lacA, is also apart of the lac operon The lacA gene codes for the enzyme transacetylase Exports excess sugar from the cell ○ Lac I is a repressor of lac operon Expressed constitutively  Binds to the operator and blocks RNA polymerase Glucose prevents the expression of lac operon: CAP regulation ○ CAP (catabolite activator protein) exerts positive control of lac operon ○ CAP is transcribed and translated constitutively ○ CAP binds to regulatory sequence upstream of promoter (CAP binding site) ○ CAP must be bound to cyclic AMP (cAMP) in order to bind to DNA Glucose prevents the expression of lac operon: Inducer exclusion ○ Transport of lactose into cell is inhibited when glucose is high ○ Lactose does not remove repressor from operator ○ When glucose is low, more lactose enters, and repressor is removed The trp operon: in presence of tryptophan ○ Trp operon involved in synthesizing amino acid tryptophan ○ Trp repressor binds to operator only when bound by its regulator - tryptophan (co-repressor) The trp operon: in absence of tryptophan ○ When tryptophan levels are low, repressor no longer binds operator ○ Trp operon genes are now transcribed; mRNA is translated and tryptophan is restored. Differential Gene expression ○ In multicellular organisms, regulation of gene expression is essential for cell specialization ○ Differences between cell types result from the expression of different genes by cells with the same genome (differential gene expression) ○ Abnormalities in gene expression can lead to diseases including cancer Gene expression is regulated at many stages Chromatin structure ○ Chromatin - DNA is wrapped around histones proteins ○ Chromatin contains nucleosomes - repeating bead like structures ○ Nucleosomes consist of negatively charged DNA wrapped twice around eight positively charged histone proteins ○ A histone protein called H1 functions to maintain the structure of each nucleosome. Chromosomes have several levels of organization ○ Nucleosomes interact with each other to form 30-nm chromatin fibers ○ Fibers are attached to scaffold proteins to hold entire chromosome in place ○ When chromosomes condense before cell division, they are even more tightly packed ○ Chromatins elaborate structure: Allows the DNA to fit in the nucleus Plays a key role in regulating gene expression Decondensed chromatin promotes gene expression ○ Chromatin must be decondenses to expose promoter for RNA polymerase to bind ○ DNAse I cannot cut tightly condensed DNA  Chromatin remodeling: DNA methylation ○ Methylation = addition of methyl (-CH3) (gene inactivation) by DNA methyltransferases ○ Methylation is important Recognized by proteins that will trigger chromatin condensation Actively transcribed genes usually have few methylated CG sequences near promoter Chromatin remodeling: Histone modification ○ Histone acetyltransferases (HATs): Add acetyl groups to histones, decondensing the chromatin (gene activation) ○ Histone deacetylases (HDACs): Remove them, leading to chromatin condensation (gene deactivation) Epigenetic inheritance ○ Changes in gene expression caused by factors other than alterations in a cell’s DNA ○ Epigenetic mechanisms can record life events that influence phenotypes of offspring Does poor nutrition in a mother produce epigenetic effects in offspring? ○ ○ Experimental setup ○ Hnf4a = hepatocyte ○ Nuclear factor 4 alpha  ○ Results: 2) Transcriptional Control ○ Eukaryotic promoters have the common conserved sequence - TATA box and associated TATA binding proteins ○ Regulatory sequences are sections of DNA that are involved in controlling the activity of genes ○ Enhances: DNA regulatory regions located away from the promoter to which specific proteins bind; increasing transcription  ○ Silencers: DNA regulatory regions to which repressors can bind; inhibiting transcription ○ Promoter-proximal elements: located close to promoter to which regulatory proteins bind Transcription initiation in Eukaryotes is a multistep process ○ 3) Post-Transcriptional Control ○ Mechanisms for post-transcriptional control: Alternative splicing Alternative splicing: leads to production of different mature mRNAs from the same primary transcript ○ Mechanisms for post-transcriptional control: mRNA stability  4) Post translational control ○ Post-translational control: proteins targeted for activation Proteins may be activated when a protein kinase adds a phosphate group. cyclin-CdK complex by phosphorylation 5) linking cancer to defects in gene regulation  ○ P53 is a tumor suppressor gene 6) comparing gene expression in bacteria and eukaryotes ○ Recombinant DNA technology: DNA cloning  ○ ○ Polymerase chain reaction (PCR) ○ The polymerase chain reaction (PCR): Quicker way to clone DNAs millions of copies obtained in hours using plasmid takes days or weeks ○ PCR made the following techniques possible: DNA fingerprinting Paternity tests Testing for bacterial and viral pathogens DNA-based genealogies PCR application in forensic science and genealogy ○ DNA fingerprinting - also known as DNA profiling or DNA typing: Identifying individuals based on their unique genome  PCR in Action: DNA fingerprinting ○ DNA fingerprinting process: Obtain DNA sample → PCR → gel electrophoresis Gel will determine number of repeats: the fewer the repeats, the shorter the PCR product ○ Bioinformatics is used to find genes ○ Gene annotation: identifying genes or other functionally important sequences ○ Computer programs scan sequences in both directions to identify reading frames: Three reading frames are possible on each strand Total of six possible reading frames  Look for long stretches of codons with no stop codon: Called open reading frames (ORFs) Good indication of a protein-coding sequence Open reading frame can identify genes ○ Chapter 21: Genes, Development, and Evolution Define differentiation, commitment, determination, genomic equivalence, differential gene expression, stem cells Understand the cloning process to create Dolly Five cellular processes of developmental biology Define morphogen (Bicoid, and what happens if bicoid is removed) Recollect segmentation genes (gap genes, pair-rule genes, segment polarity genes) Analyze the Hox genes (conserved, mutation) Introduction to development Development allows a multicellular individual to form from one cell ○ A zygote is a fertilized egg: Divides and forms a ball of cells, called an embryo Eventually forms an organism with many different types Differentiation Cells acquire specialized properties during differentiation: ○ Distinct cell types have different structures and functions ○ This is because they contain different molecules Different cells in an individual contain same genes: ○ Differential gene expression accounts for cell differentiation; only a subset of genes are expressed  Genomic Equivalence All cells are genetically equivalent - they contain the same genes If cells from a cut branch or stem can be-differentiate to form roots, these differentiated cells must contain the genes required by root cells. Plant stem cells de-differentiate into roots Genomic Equivalence in animal via cloning How does differential gene expression occurs? Commitment Commitment is when an embryonic animal cell becomes dedicated to a cell specialization path ○ This occurs gradually, initially weak and reversible ○ Cell → committed Determination Eventually, a cell is locked into a certain path - determined: ○ Cells are moved to a new location within a developing embryo If the cell is determined, it will continue on its normal path If it is not determined, it can take on a new fate Cell → committed → determine → differentiated Experimental Setup Results A master regulator is a gene product that can unleash a series of events that produce a specialized, cell type, tissue, or body structure MyoD is a master regulator of muscle differentiation Stem Cells Stem cells remain undifferentiated  Divide to produce a cell that remains a stem cell and one that differentiates Animals have stem cells in a variety of locations: ○ May replace skin, blood, and gut cells that die ○ Repair wounds ○ Create a supply of disease-fighting cells ○ Types of Stem Cells Embryonic stem cells 1. Pluripotent: cells can develop into ALL cell types 2. Plentiful 3. “Immortal”: can self-renew indefinitely 4. Embryos ARE destroyed in order to obtain them Adult stem cells 1. Multipotent: cell can develop into FEW cell types 2. Located in few organs (bone marrow, intestine) 3. Can’t renew indefinitely 4. Embryos ARE NOT destroyed in order to obtain them Stem cells in plants are called meristems: ○ Present in embryos and adults ○ Produce plant structures throughout a plant’s life ○ Shared developmental processes Five essential cellular processes lead to an individual organism’s development  1. Cells divide 2. Signal to one another what they are 3. Begin to express certain genes rather than others 4. Move, expand, or contract in specific directions 5. Some cells die 1) Cell division The location, timing, and extent of cell division must be tightly controlled Cells initiate mitosis in response to M-phase-promoting factor Cells cycle checkpoints regulate progression through the cell cycle Cells respond to signals from other cells to control division 2) Cell-Cell interactions Cells interact constantly during development through signaling molecules: ○ May diffuse through watery environment around cell ○ May be present on surface of other cells ○ May be bound to the extracellular matrix Receptors can be inside the cell or on its surface In response, gene expression changes and cells may divide, differentiate, move, change shape, or die 3) Cell differentiation Two mechanisms for specifying cell fate: ○ Cytoplasmic determinants ○ Induction  ○ ○ Cytoplasmic determinants – regulatory molecules that are unequally distributed to daughter cells ○ Induction – one daughter cell receives a signal that the other does not 4) Cell movement and Changes in shape  In animals, cells rearrange into three layers through a process called gastrulation. Plant cells have cell walls and do not move Gastrulation leads to the formation of three germ layers (Ectoderm, mesoderm, endoderm) Ectoderm derivatives = skin, neurons, pigment cells, teeth nails Mesoderm derivatives = bone, blood cells, facial muscle, kidney Endoderm derivatives = stomach, thyroid, lung 5) Programmed Cell Death Programmed cell death is a highly regulated and essential part of development: ○ Occurs in both plants and animals ○ Happens as tissues and organs take shape  ○ Programmed Cell Death is a Normal Part of Development Apoptosis is the most common type in animals: ○ Cells that form webbing between toes die ○ About half of neurons die as nervous system is wired ○ Harmful immune cells are eliminated ○ Body Axes Fate of a cell depends on its position along the three body axes: ○ One axis runs anterior to posterior ○ One axis runs dorsal to ventral ○ One axis runs left to right  ○ Morphogens Morphogens is a secreted molecule that induces cell fate decisions in recipient cells in a concentration-dependent manner Morphogens activate a network of genes that sends signals with more specific information about the spatial location of cells Bicoid Bicoid protein is a regulatory transcription factor shown to act as a master regulator with Drosophila embryos The concentration gradient formed by Bicoid protein provides cells with then information about their position along the anterior-posterior axis Segmentation genes Three types of segmentation genes 1) Gap genes: map out subdivisions along anterior-posterior (A-P) 2) Pair rules genes: define smaller segments 3) Segment polarity: define the A-P axis of each individual segment Selector genes (Hox genes) Selector genes specify the identity or fate of each segment Homeotic (Hox) genes determine the segment on each appendage or other structure will form. Hox genes are evolutionary conserved. Hox genes are evolutionary conserved Hox gene mutation Mutations in the Hox genes result in the transformation of one body segment into another FINAL REVIEW (OPTIONAL) Transcription & Translation: DNA → RNA → Protein (DNA is library, RNA is blueprint, Protein is actual thing) ○ DNA to RNA is transcription ○ RNA to protein is translation Transcription is when you have a strand of DNA one with 5’ 3’ end and the other with 3’ 5’ end with ATCG Every 3 nucleotides make an amino acid Mechanisms of gene regulation: 3 amino acids go in (for every amino acid its 100 ATP) = 300 ATP Genes cost a lot of “money” or energy