mRNA processing
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mRNA processing involves several key steps after transcription. These include 5' capping, which protects mRNA from degradation and aids in translation. Polyadenylation adds a poly-A tail to the 3' end, increasing stability and translation efficiency. Splicing removes non-coding introns and joins exons, which can produce different mRNAs and proteins from the same initial transcript. These processing steps are carried out by cellular enzymes for most eukaryotic mRNAs, but some viruses encode their own enzymes or acquire cellular mRNA caps to modify their own transcripts.
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Translation in eukaryotes
Eukaryotic translation is the process by which messenger RNA is translated into proteins. It involves three main phases: initiation, elongation, and termination. Initiation requires several eukaryotic initiation factors to form a pre-initiation complex and recruit the small ribosomal subunit to the 5' end of mRNA. Elongation then adds amino acids to the growing polypeptide chain via three elongation factors. Termination occurs when release factors recognize a stop codon and allow dissociation of the ribosome and release of the completed protein. The process is more complex in eukaryotes compared to prokaryotes due to the larger ribosome size and additional initiation factors required.
Prokaryotic DNA replication
DNA replication in prokaryotes begins with the unwinding of DNA at the origin of replication by enzymes like DnaA and DnaB helicase. This produces two replication forks that move in opposite directions. The leading strand is replicated continuously while the lagging strand is replicated discontinuously in short segments called Okazaki fragments. DNA polymerase III is the main enzyme that synthesizes new DNA. Replication terminates at the terminus region when the DnaB helicase is stopped by protein Tus bound to Ter sequences.
Rna polymerase & transcription in prokaryotes
RNA polymerase is an essential enzyme that copies DNA to produce different types of RNA in prokaryotes and eukaryotes. In prokaryotes, a single type of RNA polymerase synthesizes mRNA, tRNA, and rRNA. Transcription in prokaryotes involves initiation at promoter sequences, elongation as the RNA polymerase moves along DNA, and termination at specific sequences. Initiation requires the RNA polymerase binding to the promoter, unwinding the DNA, and beginning RNA synthesis. Elongation continues RNA synthesis as the DNA unwinds. Termination occurs at specific sequences like palindromes that allow RNA secondary structure formation and polymerase release.
Trp operon
The trp operon contains a cluster of genes involved in tryptophan biosynthesis that are under the control of a single promoter. It was the first repressible operon discovered in E. coli in 1953. The trp operon contains structural genes that encode enzymes for tryptophan synthesis, as well as a promoter, operator, and regulatory genes. Tryptophan acts as an effector molecule that binds to the repressor protein, increasing its affinity for the operator sequence and repressing transcription when tryptophan is present. The trp operon is also regulated by transcriptional attenuation, where tryptophan levels affect the formation of termination or anti-termination hairpin loops in the mRNA.
Ribozyme
Ribozymes are RNA molecules that act as enzymes and catalyze biochemical reactions. Some key points:
- Ribozymes were first proposed in the 1960s and discovered in the 1980s by Thomas Cech and Sidney Altman, who shared the 1989 Nobel Prize for the discovery.
- Common ribozyme activities include splicing and cleaving RNA and DNA. Ribozymes in the ribosome help link amino acids during protein synthesis.
- Major types of ribozymes include group I and group II introns, hammerhead, hairpin, and RNase P ribozymes. They use mechanisms like metal ion coordination and nucleophilic attacks to catalyze reactions.
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5’ capping
This document discusses RNA processing in eukaryotes. It begins by explaining that in eukaryotes, transcription and translation occur in different cellular compartments, while in prokaryotes they occur simultaneously. It then focuses on 5' capping, which is a key part of eukaryotic pre-mRNA processing. 5' capping involves the addition of a 7-methylguanosine residue to the 5' end of nascent mRNA by capping enzymes. This capping protects the mRNA from degradation and aids in transport from the nucleus to the cytoplasm and binding of ribosomes for translation. The capping can involve one or more methylation steps, producing cap0, cap1 or cap2 structures
Dna replication eukaryotes
DNA replication is the process by which DNA copies itself in living cells. It occurs in three main steps: initiation, elongation, and termination. Initiation begins at origins of replication, where proteins assemble into pre-replication complexes. During elongation, helicase unwinds the DNA strands and DNA polymerase adds complementary nucleotides to each strand. Termination occurs when the replication forks meet, with telomerase ensuring complete replication of chromosome ends.
Nucleosome and histones
The document discusses the nucleosome model of chromosome structure. It describes how DNA wraps around histone proteins to form nucleosomes, which are the basic units of chromatin. Specifically:
- Nucleosomes consist of 146-166 base pairs of DNA wrapped around an octamer of core histone proteins H2A, H2B, H3, and H4.
- Linker histone H1 binds to the DNA as it enters and exits each nucleosome, forming a structure known as a chromatosome.
- Adjacent nucleosomes are joined by 10-80 base pairs of linker DNA. The histone proteins and DNA interact via ionic bonds between negatively charged DNA and positively charged residues on
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Translation in eukaryotes
Eukaryotic translation is the process by which messenger RNA is translated into proteins. It involves three main phases: initiation, elongation, and termination. Initiation requires several eukaryotic initiation factors to form a pre-initiation complex and recruit the small ribosomal subunit to the 5' end of mRNA. Elongation then adds amino acids to the growing polypeptide chain via three elongation factors. Termination occurs when release factors recognize a stop codon and allow dissociation of the ribosome and release of the completed protein. The process is more complex in eukaryotes compared to prokaryotes due to the larger ribosome size and additional initiation factors required.
Prokaryotic DNA replication
DNA replication in prokaryotes begins with the unwinding of DNA at the origin of replication by enzymes like DnaA and DnaB helicase. This produces two replication forks that move in opposite directions. The leading strand is replicated continuously while the lagging strand is replicated discontinuously in short segments called Okazaki fragments. DNA polymerase III is the main enzyme that synthesizes new DNA. Replication terminates at the terminus region when the DnaB helicase is stopped by protein Tus bound to Ter sequences.
Rna polymerase & transcription in prokaryotes
RNA polymerase is an essential enzyme that copies DNA to produce different types of RNA in prokaryotes and eukaryotes. In prokaryotes, a single type of RNA polymerase synthesizes mRNA, tRNA, and rRNA. Transcription in prokaryotes involves initiation at promoter sequences, elongation as the RNA polymerase moves along DNA, and termination at specific sequences. Initiation requires the RNA polymerase binding to the promoter, unwinding the DNA, and beginning RNA synthesis. Elongation continues RNA synthesis as the DNA unwinds. Termination occurs at specific sequences like palindromes that allow RNA secondary structure formation and polymerase release.
Trp operon
The trp operon contains a cluster of genes involved in tryptophan biosynthesis that are under the control of a single promoter. It was the first repressible operon discovered in E. coli in 1953. The trp operon contains structural genes that encode enzymes for tryptophan synthesis, as well as a promoter, operator, and regulatory genes. Tryptophan acts as an effector molecule that binds to the repressor protein, increasing its affinity for the operator sequence and repressing transcription when tryptophan is present. The trp operon is also regulated by transcriptional attenuation, where tryptophan levels affect the formation of termination or anti-termination hairpin loops in the mRNA.
Ribozyme
Ribozymes are RNA molecules that act as enzymes and catalyze biochemical reactions. Some key points:
- Ribozymes were first proposed in the 1960s and discovered in the 1980s by Thomas Cech and Sidney Altman, who shared the 1989 Nobel Prize for the discovery.
- Common ribozyme activities include splicing and cleaving RNA and DNA. Ribozymes in the ribosome help link amino acids during protein synthesis.
- Major types of ribozymes include group I and group II introns, hammerhead, hairpin, and RNase P ribozymes. They use mechanisms like metal ion coordination and nucleophilic attacks to catalyze reactions.
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Dna replication eukaryotes
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Nucleosome and histones
The document discusses the nucleosome model of chromosome structure. It describes how DNA wraps around histone proteins to form nucleosomes, which are the basic units of chromatin. Specifically:
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- Linker histone H1 binds to the DNA as it enters and exits each nucleosome, forming a structure known as a chromatosome.
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chloroplast DNA
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TRANSLATION
Messenger RNA carries genetic code from DNA and is translated by the ribosome into proteins. This involves transfer RNA molecules that associate amino acids with their codons. Translation begins with initiation factors recruiting the small ribosomal subunit to the start codon. Elongation then occurs through peptide bond formation catalyzed by the ribosome and translocation of transfer RNAs. Termination occurs when a stop codon is reached. Translation is highly conserved and essential for protein synthesis in all organisms.
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post transcriptional modifications
Post-transcriptional modifications process the primary transcript RNA in eukaryotic cells through 5' capping, polyadenylation at the 3' end, and splicing of introns. This processing protects the RNA from degradation, aids in export from the nucleus to the cytoplasm, and allows only protein-coding exons to be translated, increasing protein synthesis efficiency. Alternative splicing also expands the proteome by producing multiple mRNA variants from a single primary transcript.
RNA Processing
Messenger RNA (mRNA) undergoes several types of processing in eukaryotes. mRNA contains 5' and 3' untranslated regions and a protein coding region. In eukaryotes, a 5' cap and poly-A tail are added. Introns are removed from pre-mRNA through splicing in the nucleus. Alternative splicing and cleavage sites allow one gene to code for multiple proteins. RNA editing can further modify the mRNA sequence. These processing steps allow for gene regulation and protein diversity from a single DNA sequence.
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Post-transcriptional modifications help process primary transcripts into mRNA in eukaryotes. This involves 5' capping, addition of a poly-A tail, and splicing of introns. 5' capping adds a guanine cap to protect the 5' end from degradation. Poly-A polymerase adds around 200 adenine bases to the 3' end. Splicing removes introns and ligates exons by the spliceosome, producing mature mRNA that can be translated into protein. Alternative splicing allows different mRNA and protein isoforms from a single gene.
Post transcriptional processing
The document discusses various post-transcriptional processes that eukaryotic RNA undergo. It describes:
1) Introns are removed and exons are joined together in mRNA through splicing. Small nuclear RNAs and spliceosomes are involved in splicing.
2) Other processing includes adding a 5' cap and 3' poly-A tail. The tail protects the mRNA and is important for translation.
3) Transfer RNA and ribosomal RNA also undergo processing after transcription to become functional in the cell. Alternative splicing allows different mRNAs and proteins to be produced from the same initial RNA.
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1) Coronaviridae is a family of enveloped, positive-stranded RNA viruses which infect amphibians, birds and mammals. They were recognized as a new virus family in 1968.
2) Coronaviruses have large club-shaped spikes and a genome consisting of the largest known non-segmented RNA. They utilize a distinctive nested set transcription strategy.
3) Viral replication occurs within an extensive membranous network, utilizing a complex strategy of full-length genomic RNA synthesis and a nested set of subgenomic mRNAs. This allows expression of downstream genes and leads to genetic diversity through recombination.
Post transcriptional modification
Post-transcriptional modifications help process primary transcripts into mRNA in three main ways: 1) 5' capping protects the transcript and aids export from the nucleus, 2) Polyadenylation aids stability and transport, and 3) Splicing removes introns and ligates exons to form mature mRNA. In eukaryotes, this occurs in the nucleus and is essential for efficient translation. It can also result in alternative splicing to increase protein diversity from a single gene.
Transcription
1. Eukaryotic pre-mRNA undergoes processing after being exported from the nucleus which involves removal of introns, addition of a 5' cap, and 3' polyadenylated tail to produce mature mRNA.
2. The mature mRNA is translated by ribosomes in the cytoplasm. Some viruses utilize cap-independent translation initiation or other mechanisms like leaky scanning, frameshifting, or suppression of termination to express their genes.
3. Interferons induce antiviral responses including RNase L and PKR activation. Viruses have evolved strategies to antagonize PKR such as inhibiting its dsRNA binding, dimerization, or kinase activity to prevent host translation shutoff.
Transcription sm.pptx
RNA is synthesized from DNA in a process called transcription. There are three main stages: initiation, elongation, and termination. In initiation, RNA polymerase binds to promoter sequences and unwinds the DNA helix. In elongation, RNA polymerase reads the template strand and adds complementary RNA nucleotides. Termination occurs when termination signals are reached. Prokaryotes and eukaryotes differ in their RNA polymerases and transcription control. Eukaryotic pre-RNA undergoes processing including 5' capping, 3' polyadenylation, splicing, and editing to produce mature mRNA.
Transcription
The document discusses the process of transcription in prokaryotes and eukaryotes. It describes the three main stages of transcription - initiation, elongation, and termination - and how they differ between prokaryotes and eukaryotes. In eukaryotes, the mRNA transcript undergoes processing including capping, polyadenylation, and splicing before being exported from the nucleus, while in prokaryotes the mRNA is used directly for translation. The structures of RNA polymerases also differ between the two systems.
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The document discusses molecular information flow in microorganisms. It describes the central dogma of DNA to RNA to protein, with the three main steps being replication, transcription, and translation. Replication copies DNA, transcription transfers genetic information from DNA to RNA, and translation uses the information in mRNA to form a polypeptide on the ribosome. The genetic elements in bacteria are usually a single circular chromosome and sometimes additional plasmids.
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mRNA processing
- 3. 1 Introduction Many of the RNA molecules in bacteria and virtually all RNA molecules in eukaryotes are processed to some degree after synthesis
- 4. 1 Introduction • mRNAs are synthesized by either cellular or viral enzymes • RNA processing- a series of covalent modifications • Facilitate recognition of mRNAs by the protein synthesizing machinery • Most RNA-processing reactions were discovered in viral systems
- 5. Principles of Virology, ASM Press 1 Introduction
- 7. 2 5’Capping Principles of Virology, ASM Press
- 8. 2 5’Capping • Protects mRNAs from 5’ exonucleolytic attack • Essential for the efficient translation of most mRNAs • de novo synthesis by cellular enzymes, synthesis by viral enzymes and acquisition of preformed 5’ cap
- 9. 2 5’Capping Synthesis by cellular enzymes • Pre mRNA are substrates for cellular capping enzymes • RNA pol II • Cotranscriptional reaction that takes place when nascent RNA is only 20 to 30 nucleotides in length • Phosphorylation of paused RNA pol II at specific serines in the C-terminal domain of the largest subunit is the signal for binding of the capping enzyme and capping of nascent RNA
- 10. 2 5’Capping Principles of Virology, ASM Press
- 11. 2 5’Capping Acquisition of viral 5’Cap from cellular mRNA • 5’ caps of orthomyxoviral and bunyaviral mRNAs are produced by cellular capping enzymes • Viral cap dependent endonucleases cleave cellular transcripts to produce the primers needed for viral mRNA synthesis- cap snatching • The 5’ terminal segments and caps of influenza virus mRNAs are obtained from cellular pre-mRNA in nucleus • Bunyaviral mRNA synthesis is primed with 5’ terminal fragments cleaved from mature cellular mRNAs in cytoplasm
- 12. 3 Polyadenylation Addition of poly(A) tail to 3’ end
- 13. 3 Polyadenylation History Principles of Virology, ASM Press
- 14. 3 Polyadenylation • Stability and increases the efficiency of translation • 3’ terminal stemloop structures instead poly(A) tail in mRNAs that encode histones and also in case of reoviral and arenaviral mRNAs • Carried out by either cellular or viral enzymes
- 15. 3 Polyadenylation By cellular Enzymes • Transcription of a gene proceeds beyond the site at which poly(A) tail is added • The 3’ end of the mRNA is determined by endonucleolytic cleavage of its pre-mRNA • Poly(A) is then added to the new 3’ terminus • Termination of transcription • Degradation of RNA downstream to the cleavage site
- 16. 3 Polyadenylation Principles of Virology, ASM PressLehninger principles of Biochemistry, WH Freeman
- 17. 3 Polyadenylation By Viral Enzymes • Either posttranscriptional or during viral mRNA synthesis • Vaccinia virus- 2 processes with one enzyme • Other viruses- during mRNA synthesis • Poly(U) sequence present at the 5’ end of the (-) strand RNA template Principles of Virology, ASM Press
- 18. 4 Splicing Non-coding regions are removed and coding regions are joined together
- 19. 4 Splicing Discovery • Nuclear precursors of mRNAs are larger than mRNAs transported in cytoplasm and heterogenous in size • Both ends of hnRNA retained • 1993- Phillip Sharp and Richard Roberts • Adenoviral late mRNAs
- 20. 4 Splicing Principles of Virology, ASM Press
- 21. 4 Splicing 4 classes of introns • Group I: Found in some nuclear, mitochondrial and chloroplast genes that code for rRNAs, mRNAs and tRNAs • Group II: Found in the primary transcripts of mitochondrial or chloroplast mRNAs in fungi,algae and plants • Rare examples of introns found in bacteria
- 22. 4 Splicing Lehninger principles of Biochemistry, WH Freeman
- 23. 4 Splicing Group III Introns • Largest class • Found in nuclear mRNA primary transcripts • Spliceosomal introns • Splicing by lariat mechanism • Spliceosome is made up of specialized RNA-protein complexes: snRNPs
- 24. 4 Splicing Lehninger principles of Biochemistry, WH Freeman
- 25. 4 Splicing Lehninger principles of Biochemistry, WH Freeman
- 26. 4 Splicing Differential RNA processing Lehninger principles of Biochemistry, WH Freeman
- 27. 4 Splicing Principles of Virology, ASM Press
- 28. 5 Conclusion
- 29. 5 Conclusion • Eukaryotic mRNAs are modified by addition of a 7- methylguanosine residue at the 5’ end and by cleavage and polyadenylation at 3’ end • Introns are removed by splicing • Group I introns require guanosine cofactor • Some group I and group II introns are capable of self splicing • Group III introns are spliced with the aid of RNA-protein complexes called snRNPs • Complex transcripts can have either more than one site for cleavage and polyadenylation or both
- 30. 6 Reference •Flint, Racaniello, 2015, Principles of virology, 4th edition •David L Nelson, M Cox, 2013, Lehninger Principles of Biochemistry, sixth edition, W H Freeman and Company
- 31. THANK YOU!