Lesson 7 - From Plan to Protein - BIOL 1441

Lesson 7: From Plan to Protein BIOL 1441 Cell & Molecular Biology  Learning Objectives - Study Guide By the end of this lesson, students will be able to: 1. Explain the main differences between DNA, RNA, & proteins. 2. Identify the locations of DNA & the various RNAs in the cell....

Lesson 7: From Plan to Protein BIOL 1441 Cell & Molecular Biology  Learning Objectives - Study Guide By the end of this lesson, students will be able to: 1. Explain the main differences between DNA, RNA, & proteins. 2. Identify the locations of DNA & the various RNAs in the cell. 3. Explain the relationship between DNA, a gene, nucleotides, amino acids, and proteins. 4. List the parts of a gene & describe the functions of each. 5. Describe how the process of transcription works. 6. Transcribe a DNA sequence into an mRNA sequence. 7. Describe the structures that are added to & removed from an mRNA transcript before it leaves the nucleus. 8. Explain what alternative splicing is & why it is important for cells. 9. Identify the organelles that perform the process of translation.  Learning Objectives - Study Guide By the end of this lesson, students will be able to: 10.Describe how the process of translation works. 11.Explain what a codon is. 12.Use the Genetic Code Table to translate an mRNA sequence. 13.Explain what differential gene expression is & why it is used in cells. 14.Describe the role of transcription factors in differential gene expression. 15.Explain what occurs in a point mutation & a frameshift mutation. 16.Describe the effects of silent mutations, missense mutations, and nonsense mutations on translation. 17.Identify the major sources of mutations in a cell. 18.Describe how a cell responds to mutations.  Remember, You are Your own Instruction Manual! In the last lecture we discussed how DNA makes more DNA (DNA replication). In this lecture we will learn about how cells assemble proteins, using the instructions in the DNA.  Gene Expression Gene 2 Gene expression is the process of DNA molecule using the information stored in a gene Gene 1 to build a functional protein or RNA Gene 3 molecule A gene is a region of DNA that carries the instructions for making specific DNA proteins or RNA molecules. template strand Gene expression involves 2 steps: Step #1 – Transcription TRANSCRIPTION DNA for a specific gene is used as a mRNA template for making mRNA (messenger RNA) Codon TRANSLATION Step #2 – Translation Protein mRNA is used as a template for Amino acid assembling a protein  Gene Expression and RNA There are multiple kinds of RNA involved in protein synthesis: Messenger RNA (mRNA) carries protein-building information from DNA to the ribosome Ribosomal RNA (rRNA) and proteins make up ribosomes Transfer RNA (tRNA) brings amino acids to the ribosomes  Transcription During transcription, RNA is made using the information in the DNA template. RNA contains uracil in place of thymine. DNA-RNA base pairing follows these rules: A-U C-G mRNA (RNA that specifies the amino acid sequence of a protein) then carries the instructions out of the nucleus to the ribosome  Reminder: DNA vs. RNA Transcription uses the DNA template to build RNA. DNA bases RNA bases RNA is single- DNA is double- stranded stranded DNA is made RNA is made with with ribose sugar deoxyribose sugar RNA includes A, DNA includes A, U, G, & C T, G, & C nucleotides nucleotides RNA leaves the DNA is found in nucleus & enters the cell’s the cytoplasm nucleus  Nucleotid es DNA and RNA consist of monomers known as nucleotides Nucleotides consist of three parts 1. Nitrogenous base 2. Pentose sugar 3. One phosphate group (when the nucleotide is part of a DNA or RNA strand) Types of nitrogenous bases Pyrimidines – cytosine, thymine, uracil (uracil in RNA replaces thymine in DNA) Purines – Adenine, guanine Types of pentose sugars Deoxyribose (found in DNA) Ribose (found in RNA)  Let’s Practice! Use the DNA strand below to transcribe the complementary mRNA strand: 3’ T A C G G T A G C G A T T T C 5’  Stop & Think it Through! DNA mRNA Where is it found? What sugar does it contain? List its 4 nucleotides Is it typically single- stranded or double- stranded? What is the function of this molecule?  The Central Dogma: An Overview Transcription occurs in the nucleus of a eukaryotic cell RNA polymerase opens the DNA double helix & creates a complementary pre-mRNA strand using the sequence of nucleotides in a gene That pre-mRNA is processed, then leaves the nucleus Translation occurs in the cytoplasm of a eukaryotic cell The mRNA molecule attaches to a ribosome The ribosome “reads” the mRNA & creates a complementary amino acid chain  Parts of a Gene The promoter region of a The terminator region (stop gene is the location where RNA sequence) signals to RNA polymerase polymerase attaches to the that the whole gene has been DNA molecule. transcribed, so it can release the DNA. The specific directions for building the protein are stored in this part of  Transcription: An Overview Step 1 – Initiation RNA polymerase attaches to the DNA strand at the promoter Transcription factors are proteins that bind to the DNA and regulate transcription Step 2 – Elongation RNA polymerase creates a complementary pre-mRNA molecule Step 3 – Termination  Introns vs. Exons Pre-mRNA molecules in eukaryotes include introns & exons Introns are “intervening sequences” between the exons in a pre-mRNA molecule Introns do NOT have information involved in protein synthesis Exons are the protein- coding regions of a pre- mRNA molecule After the introns are spliced  Alternative Splicing Alternative splicing allows a cell to make more than one protein using a single gene Exons are selectively removed from a pre-mRNA transcript This can generate multiple different mRNA molecules from the same pre-mRNA transcript  Translation mRNA is used by the ribosome as the instructions for assembling the correct amino acids in the correct order tRNA (transfer RNA) carries amino acids to Each tRNA molecule has an: the ribosome so they amino acid attachment site that enables can be assembled into a tRNA to bring amino acids to the protein ribosome anticodon, the 3 nucleotides it will use to “read” the mRNA codons  Ribosome Structure Ribosomes are made of ribosomal RNA (rRNA) & proteins Ribosomes have 2 subunits Small subunit is the part of a ribosome that mRNA molecules attach to Large subunit joins the amino acids together as tRNA delivers them Each ribosome has 3 binding sites for tRNA molecules: the A, P, & E sites  “Reading” mRNA tRNA “reads” mRNA in short 3- nucleotide groups called codons Each codon is complementary to the anticodon on a specific tRNA molecule To correctly read an mRNA message, translation must start at the correct location on the mRNA strand This establishes the reading frame, ensuring that the right groups of 3 letters are “read” as a codon The start codon, which marks the beginning of every protein, is AUG  Translation: An Overview Step 1: Initiation Step 2: Elongation The Start codon tRNA “reads” the is located on the mRNA and puts mRNA the correct amino Step 3: Termination The two ribosomal acids in the The ribosome subunits come correct order reaches a stop together (sequence) codon A chain of amino It releases its acids is created amino acid chain Initiation Elongation Termination Translation Animation  The Genetic Code Table The Genetic Code table can be used to determine which amino acid correlates to each codon in an mRNA molecule The Genetic Code is degenerate This means more than one codon leads to the same amino acid This creates some “wiggle room” for errors in transcription BUT – only the last letter in a codon can be changed without major consequences This is called the codon’s wobble base  Let’s Practice! Translate the following mRNA molecule into an amino acid sequence (remember the importance of reading frame): 5’ A U G C C A U C G C U A U G A 3’ From … To Plan… Protein A “copy” of the blueprint is redrawn with The workers specific use that measurements version to build that the a new building. construction workers can follow.  Transcription Translation The nucleic acid it starts with: What it ends with: Where this happens: DNA Amino acids DNA Amino acids Circle all the things it mRNA Ribosomes mRNA Ribosomes uses: Nucleotides tRNA RNA Nucleotides tRNA RNA Polymerase Polymerase  Consider This… Every cell in your entire body has the exact same genome This means that all of your cells have the potential to make ANY of the proteins your genome encodes! But… not every cell uses (expresses) all the genes Neurons don’t make myosin (a muscle contraction protein), and… Muscle cells don’t make insulin (a protein that enables glucose entry into your cells), and… Pancreatic cells don’t make acetylcholine (a protein neurons use to communicate with one another)  Differential Gene Expression Differential gene expression is what cells use to selectively express only the genes that encode proteins they actually need Differential gene expression enables non-specialized stem cells to Stem differentiate into cell specific types of functional cells.  Transcription factors are proteins that assist with differential gene expression. Repressor proteins block RNA polymerase, leading to decreased protein production. Activator proteins increase the rate of transcription, leading to increased protein production.  Differential Gene Expression & Insulin Resistance Insulin resistance is a common symptom of Type 2 diabetes. In this condition, pancreatic β cells still make insulin, but the body’s cells don’t respond to it This means the cells are less able to import glucose One cause of insulin resistance is a decrease in the expression of the insulin receptor gene As gene expression decreases, fewer insulin receptors are made The lack of these receptors makes cells unable to take up glucose  Mutations Mutations are changes to the DNA or mRNA of a gene that is being used to build a protein If mutations are not repaired, the proteins may be misshapen and potentially non-functional Types of mutations In a point mutation, a single nucleotide is incorrect In a frameshift mutation, Point mutations occur when an Types of Point incorrect nucleotide is used to Mutations build a DNA or mRNA strand. Silent mutation - the changed nucleotide does NOT impact the amino acid added to the protein Missense mutation - the changed nucleotide leads to the WRONG amino acid being added to the protein Nonsense mutation - the changed nucleotide leads to the generation of a STOP codon  Frameshift mutations are caused by the insertion / deletion of 1 - 2 nucleotides (occasionally more). Frameshift mutations disrupt the ribosome’s reading frame. This means the mRNA nucleotides are “read” in incorrect groups. Frameshift mutations can have Frameshift mutations can missense effects  the wrong have nonsense effects  amino acid is added to the a new STOP codon to is protein. generated (ending  If 3 nucleotides are inserted or deleted at the same time, the reading frame does not change. BUT : The protein that is generated will still have an incorrect amino acid sequence. “Mutations” in a Sentence: An Example Point mutatio ns Frameshift mutations  Mutations Faulty proteins can arise due to Mutations can arise due to errors in normal cellular exposure to mutagens in the processes. environment. If the error is in the DNA, it is called a mutation.  Mutation If the mutation Detected! cannot be Response repaired… Response #1: #2: Attempt DNA Repair Apoptosis + Halt the Cell Cycle This prevents mutated cells from undergoing mitosis This programmed cell death & generating ensures that cells with mutated offspring. mutated DNA are eliminated.  Diabetes & Mutations Diabetes has been linked to multiple mutations in in the insulin-encoding gene (INS gene) These mutations affect non- protein-coding regions of the gene. These mutations affect protein- coding regions of the gene.  Mutations in the INS gene ultimately lead to changes the structure of the insulin protein. What does this picture show? The central (colored) circles represent the normal amino acid sequence of the insulin protein. The external circles represent incorrect amino acids present in the insulin made by patients with diabetes.  To Prepare for Next Class…  Review your class notes Use the eTextbook & Other Helpful Resources to supplement your lecture notes  Complete the homework assignment and use it to direct your studying  Print the slides for Lesson #8: The Cellular Circle of Life