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CRAZY ABOUT BIOMECIDINE Having a look into frog oocytes TRANSLATIONAL CONTROL OF mRNAs BY CYTOPLASMIC POLYADENYLATION DURING MEIOSIS

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Gene expression Gene regulation Translational control of mRNAs CPEBs & cytoplasmic polyadenylation Meiosis and translational control of mRNAs mediated by CPEBs CPEBs beyond meiosis

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GENE EXPRESSION Genetic information Effectors Cell signaling Cell adhesion Enzymes Define the structure Transport

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GENE EXPRESSION Genetic information Effectors Cell signaling Cell adhesion Enzymes Define the structure Transport Key steps in gene expression RNA transcription: from DNA to RNA RNA processing: from RNA to mRNA mRNA transport: from the nucleus to the cytoplasm Protein translation: from mRNA to protein

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GENE EXPRESSION Genetic information Effectors Cell signaling Cell adhesion Enzymes Define the structure Transport RNA processing DNA Pre-mRNA mRNA

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GENE EXPRESSION Genetic information Effectors Cell signaling Cell adhesion Enzymes Define the structure Transport mRNA features CDS 5UTR3UTR 5 cap AAAAAAAAAA AUG UGA

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GENE EXPRESSION Genetic information Effectors Cell signaling Cell adhesion Enzymes Define the structure Transport Protein translation

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TRANSLATIONAL CONTROL OF mRNAs BY CYTOPLASMIC POLYADENYLATION DURING MEIOSIS Gene expression Gene regulation Translational control of mRNAs CPEBs & cytoplasmic polyadenylation Meiosis and translational control of mRNAs mediated by CPEBs CPEBs beyond meiosis

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GENE REGULATION EVERY CELL IN A GIVEN ORGANISM CONTAINS THE SAME GENETIC INFORMATION DIFFERENCES IN GENE EXPRESSION

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GENE REGULATION Every step in gene expression is regulated TRANSCRIPTION Accessibility of DNA Transcriptional regulation

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GENE REGULATION Every step in gene expression is regulated RNA PROCESSING Alternative splicing Alternative 3UTR formation: different 3UTR AAA CDS 5UTR 3UTR 5 cap AAA AUG CDS AUG TRANSCRIPTION Accessibility of DNA Transcriptional regulation

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GENE REGULATION Every step in gene expression is regulated RNA PROCESSING Alternative splicing Alternative 3UTR formation: different 3UTR PROTEIN TRANSLATION Global vs. mRNA specific regulation Translational repression precluding ribosome recruitment or progression through the mRNA TRANSCRIPTION Accessibility of DNA Transcriptional regulation

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TRANSLATIONAL CONTROL OF mRNAs BY CYTOPLASMIC POLYADENYLATION DURING MEIOSIS Gene expression Gene regulation Translational control of mRNAs CPEBs & cytoplasmic polyadenylation Meiosis and translational control of mRNAs mediated by CPEBs CPEBs beyond meiosis

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TRANSLATIONAL CONTROL OF mRNAs WHY mRNAs should be stored in the cytoplasm instead of being immediately translated? AAAAAA nucleus cytoplasm AAAAAA A Once transcribed, mRNAs can have different fates.

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TRANSLATIONAL CONTROL OF mRNAs WHY mRNAs should be stored in the cytoplasm instead of being immediately translated? Rapid response Absolutely required in situations with absence of transcription To solve long distances (axons can extend up to 1 meter!)

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TRANSLATIONAL CONTROL OF mRNAs mRNA translation can be divided into 3 phases: initiation, elongation and termination All mRNAs exit the nucleus with a long poly(A) tail and should be efficiently translated INITIATION is the phase more tightly controlled

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TRANSLATIONAL CONTROL OF mRNAs HOW TO CONTROL TRANSLATION AT INITITATION? sequestering initiation factors blocking the 5 cap shortening the poly(A) tail AAAAAAAAAAAAA 4G 4E 4E- BP Sequester initiation factors

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TRANSLATIONAL CONTROL OF mRNAs HOW TO CONTROL TRANSLATION AT INITITATION? sequestering initiation factors blocking the 5 cap shortening the poly(A) tail AAAAAAAAAAAAA 4G 4E AAAAAAAAAAAAA 4E Block the 5 cap no recruitment of the ribosome

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TRANSLATIONAL CONTROL OF mRNAs HOW TO CONTROL TRANSLATION AT INITITATION? sequestering initiation factors blocking the 5 cap shortening the poly(A) tail AAAAAAAAAAAAA 4G 4E A Shorten the poly(A) tail no close loop

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TRANSLATIONAL CONTROL OF mRNAs HOW TO CONTROL TRANSLATION AT INITITATION? sequestering initiation factors blocking the 5 cap shortening the poly(A) tail AAAAAAAAAAAAA 4G 4E TRANSLATIONAL REPRESSION doesnt last forever!!! Activation of mRNA translation Cell stimuli releasing initiation factors allowing the 5 cap to interact with initiation factors elongating the poly(A) tail How to activate translation?

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TRANSLATIONAL CONTROL OF mRNAs BY CYTOPLASMIC POLYADENYLATION DURING MEIOSIS Gene expression Gene regulation Translational control of mRNAs CPEBs & cytoplasmic polyadenylation Meiosis and translational control of mRNAs mediated by CPEBs CPEBs beyond meiosis

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CPEBs and CYTOPLASMIC POLYADENYLATION A 4E NO TRANSLATION Polyadenylation can happen in the cytoplasm!!!! AAAAAAAAAAAAA 4G 4E TRANSLATION

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CPEBs and CYTOPLASMIC POLYADENYLATION CPEBs play an essential role RNA binding proteins Bind to specific sequences in the 3UTR of target mRNAs Have a dual function!!!! CPEB in repressionCPEB in activation CPEB recruits other proteins that shorten the poly(A) tail and block the cap CPEB recruits other proteins that elongate the poly(A) tail WHAT MAKES THE DIFFERENCE???? stimulus Kinase activation

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TRANSLATIONAL CONTROL OF mRNAs BY CYTOPLASMIC POLYADENYLATION DURING MEIOSIS Gene expression Gene regulation Translational control of mRNAs CPEBs & cytoplasmic polyadenylation Meiosis and translational control of mRNAs mediated by CPEBs CPEBs beyond meiosis

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MEIOSIS and CPEBs

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Model system: Xenopus laevis oocytes Oocytes arrested at the prophase of meiosis I (progesterone makes them progress to metaphase of meiosis II) Eggs arrested at the metaphase of meiosis II (fertilization makes them finish meiosis and start embryonic divisions) Oocytes Eggs progesterone fertilization

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MEIOSIS and CPEBs Model system: Xenopus laevis oocytes Prophase I arrested oocytes grow until they are fully competent for development: 6 growth stages >1.2mm! (human: 0.12mm) Accumulation of molecular reserves Like maternal mRNAs! MEIOSIS AND FIRST EMBRYIONIC DIVISIONS HAPPEN IN THE ABSENCE OF TRANSCRIPTION! PROTEIN EXPRESSION DUE TO TRANSLATIONAL ACTIVATION OF mRNAs!

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MEIOSIS and CPEBs sVI. Oocytes progesterone Requires activation of translation of stored mRNAs Model system: Xenopus laevis oocytes Meiotic progression requires protein translation!!!!

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MEIOSIS and CPEBs Model system: Xenopus laevis oocytes Meiotic progression requires protein translation!!!! Stage VI oocytes store A LOT of maternal mRNAs repressed by CPEB1 + progesterone MEIOSIS RESUMPTION CPEB1 is phosphorylated Activation of translation of mRNAs

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MEIOSIS and CPEBs CPEB1 is not the only one! CPEB4 takes the job in late meiotic phases. Oocytes arrested prog. CPEB1CPEB4 CPEB1 degraded CPEB4 synthetized repression activation

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MEIOSIS and CPEBs CPEB1 is not the only one! CPEB4 takes the job in late meiotic phases. They both recognize the same elements in the 3UTR of target mRNAs. CPEB1 degraded CPEB4 synthetized progesterone Aurora A kinase activity CPEB1 repression activation Oocytes arrested

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CURIOSITY THE POWER OF THE CYTOPLASM! Early development happens in the absence of transcription Everything you need for early development is contained in the egg cytoplasm

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TRANSLATIONAL CONTROL OF mRNAs BY CYTOPLASMIC POLYADENYLATION DURING MEIOSIS Gene expression Gene regulation Translational control of mRNAs CPEBs & cytoplasmic polyadenylation Meiosis and translational control of mRNAs mediated by CPEBs CPEBs beyond meiosis

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CPEBs BEYOND MEIOSIS CPEBs are involved in local translation in dendrites CPEBs are required for long term memory CPEBs are involved in cell proliferation, angiogenesis CPEB4 is involved in pancreatic adenocarcinoma

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Ral Mndez Lab