BIO230 Lecture 7: Regulation of the Proteome PDF

Reminders 1. Check Quercus for announcements. 2. Read the textbook. 3. Come to the lectures. Add extra notes. 4. Re-read the textbook and add to your notes. 5. Use the recordings (if available) as a backup. 6. Tuesday 1:1 question period / discussion board...

Reminders 1. Check Quercus for announcements. 2. Read the textbook. 3. Come to the lectures. Add extra notes. 4. Re-read the textbook and add to your notes. 5. Use the recordings (if available) as a backup. 6. Tuesday 1:1 question period / discussion board Try ChatBIO230.com Use at your own risk Interact with the material critically to improve your learning No lab material BIO230 Section 1: Regulation of Genome Expression Lecture 7: Regulation of the Proteome BIO230H1F: From Genes to Organisms Prof. Kenneth W. Yip, Ph.D. Assistant Professor, Teaching Stream Cell & Systems Biology, University of Toronto Fall 2024 Hematopoiesis BIO230 Section 1 Lecture 7: Regulation of Proteome 1. Translational Regulation 2. Post-translational Regulation Pg. 159-161 Many Changes in Proteins Are Driven by Protein Phosphorylation A Eukaryotic Cell Contains a Large Collection of Protein Kinases and Protein Phosphatases Pg. 162 Regulatory GTP-binding Proteins Are Switched On and Off by the Gain and Loss of a Phosphate Group Pg. 456-457 Untranslated Regions of mRNAs Control Their Translation Readings (Alberts et al.): The Phosphorylation of an Initiator Factor Regulates All listed on Quercus Protein Synthesis Globally Pg. 459-461 Changes in mRNA Stability Can Control Gene Expression Pg. 380-382 The Ribosome Coordinates the Folding, Enzymatic Modification, and Assembly of Newly Synthesized Proteins Molecular Chaperons Help Guide the Folding of Most Proteins BIO230 Lecture 1-7 3 Regulation of Genome Expression Genome Transcriptome Proteome Post-Transcriptional Post-Translational DNA Transcription RNA Translation Protein Sorting Organization Splicing Localization Metabolome Interactome BIO230 Lecture 1-7 4 Regulation of the Proteome Differences in the proteins expressed by two human tissues Red: common to both Blue: Tissue specific BIO230 Lecture 1-7 5 Translational Regulation Both prokaryotes and eukaryotes use translational control mechanisms to regulate protein expression, often in response to stressful situations such as low nutrients, infection, or environmental stresses (e.g., temperature) Prokaryotes mRNAs have a six nucleotide Shine-Dalgarno (SD) sequence upstream of the AUG start codon correctly positions AUG in the ribosome and provides translational control mechanisms BIO230 Lecture 1-7 6 Translational Regulation: Prokaryotes Mechanism 1: A specific RNA binding protein blocks access to the SD sequence BIO230 Lecture 1-7 7 Translational Regulation: Prokaryotes Mechanism 2: Temperature regulated RNA structures e.g., virulence genes of human pathogen Listeria monocytogenes BIO230 Lecture 1-7 8 Translational Regulation: Prokaryotes BIO230 Lecture 1-7 9 Translational Regulation: Prokaryotes Mechanism 3: Riboswitch e.g., S-adenosyl methionine BIO230 Lecture 1-7 10 Translational Regulation: Prokaryotes Mechanism 4: Antisense RNA e.g., iron storage proteins Antisense RNA produced elsewhere in the genome base-pairs with mRNA and blocks SD BIO230 Lecture 1-7 11  Translational Regulation: Eukaryotes No Shine-Dalgarno sequences, but there are similar mechanisms translational repressors can bind near initiator AUG and inhibit translation (e.g., aconitase) Ferritin binds iron and releases it in a controlled manner not needed when iron is low aconitase binds to the ferritin RNA near the start site and blocks translation translated when iron is in excess aconitase binds iron conformational change ferritin RNA released (recall aconitase also regulates transferrin receptor) BIO230 Lecture 1-7 12 Translational Regulation: Eukaryotes Repressor proteins can also interfere with 5’ cap and 3’ poly-A tail interactions required for efficient translation Small RNA molecules can also regulate eukaryotic translation (miRNAs) different mechanism than in prokaryotes There are also other eukaryotic specific mechanisms (e.g., eIF2)… BIO230 Lecture 1-7 13 Translational Regulation: Eukaryotic Translation Initiation Regulation of eukaryotic initiation factors (eIFs): eIF2 plays a crucial role in translation initiation eIF2 forms a complex with GTP and recruits the initiator tRNA (methionyl) to the small ribosomal subunit The small ribosomal subunit binds the 5’ end of mRNA and scans for the first AUG BIO230 Lecture 1-7 14 Translational Regulation: Eukaryotic Translation Initiation Regulation of eukaryotic initiation factors (eIFs): when AUG is recognized, eIF2 hydrolyzes GTP to GDP GTP hydrolysis causes a conformational change in eIF2 eIF2 bound to GDP is released eIF2 bound to GDP is inactive… BIO230 Lecture 1-7 15 Translational Regulation: Eukaryotic Translation Initiation Regulation of eukaryotic initiation factors (eIFs): reactivation of eIF2 requires eIF2B which is a guanine nucleotide exchange factor (GEF), meaning it causes the exchange of GDP for GTP BUT eIF2 reactivation is regulated by phosphorylation… BIO230 Lecture 1-7 16 Translational Regulation: Eukaryotic Translation Initiation Regulation of eukaryotic initiation factors (eIFs): phosphorylated eIF2 sequesters eIF2B as an inactive complex since there is more eIF2 than eIF2B in cells, all eIF2B is sequestered and translation is dramatically reduced not all mRNAs are equally affected by eIF2 phosphorylation BIO230 Lecture 1-7 17 Regulation of Genome Expression Genome Transcriptome Proteome Post-Transcriptional Post-Translational DNA Transcription RNA Translation Protein Sorting Organization Splicing Localization Metabolome Interactome BIO230 Lecture 1-7 18 Post-Translational Regulation: Proteins Proteins undergo a number of steps in order to become functional: 1. proteins must fold properly to adopt their 3D structure 2. proteins are covalently modified with chemical groups (eg. sugars, phosphate) 3. Proteins interact with other proteins and small molecules (cofactors) BIO230 Lecture 1-7 19 Post-Translational Regulation: Protein Folding hydrophobic amino acids are buried in the interior core (ie. not surface exposed) For some proteins, folding begins as they emerge from ribosomes; some are completely folded after synthesis BIO230 Lecture 1-7 20 Post-Translational Regulation: Protein Folding most proteins require a special class of proteins called chaperones for proper folding many chaperones are called heat-shock proteins (Hsp) since they are synthesized to high amounts by cells at elevated temperatures Hsp70 and Hsp60 (chaperonin) assisted protein folding: Both interact with exposed hydrophobic residues of misfolded proteins Both use energy from ATP hydrolysis to promote proper folding BIO230 Lecture 1-7 21  Post-Translational Regulation: Protein Folding Improperly folded proteins can aggregate and become toxic to cells misfolded proteins are the cause of many inherited human diseases the process is closely monitored by a protein degrading apparatus called the proteasome exposed hydrophobic residues mark protein for degradation by the proteasome; competes with chaperones for misfolded proteins longer time to fold, more chance of being degraded BIO230 Lecture 1-7 22 Review Question How many of the following statements are correct? i. Treatment of cells with HSP inhibitors should result in a decrease in protein degradation. ii. Treatment of cells with an iron chelator (which removes excess cellular iron) should result in a decrease in ferritin transcription. iii. Treatment of cells with a miRNA complementary to eIF2B should result in an increase in transcription. a. 0 b. 1 c. 2 d. 3 BIO230 Lecture 1-7 23 Review the textbook and add to your notes. If that was too quick for you, or if you have additional questions, please review the textbook, review the recording (if available), post on the Discussion Board, stay for the Q&A sessions, and try ChatBIO230. This is your responsibility.