Central Dogma: Information Flow from DNA to RNA to Protein (Ch.6)- The Big Pictureo Genetic Information stored in chromosomes must be read and converted into proteins (usually) in the cytosol in order to be usefulo DNA-encoded information is first transcribed into an RNA intermediary by RNA polymeraseo After transcription, the RNA molecule must undergo several processing steps to become a mature messenger RNA (mRNA)o mRNA molecules are exported from the nucleus to the cytosol, where they can betranslated into proteins by ribosomeso Some important RNA molecules do not code for proteins, and are processed differently than mRNA moleculeso Both transcription and translation are highly regulated, and use many energy-dependent steps to ensure high fidelity- The Central Dogmao Information flow in all prokaryotes and eukaryotes proceeds from DNARNAproteino Although the mechanisms are very similar between prokaryotes and eukaryotes, there are several differences that address specific difficulties in eukaryotes: Eukaryotic chromosomes are highly packaged into chromatin DNA is in the nucleus, but protein synthesis machinery is in the cytosol Eukaryotic genes are interrupted by large intervening sequences (“introns”), which must be removedo Information content: DNA>mRNA>proteins DNA has most because it must have the information and the genes that regulate them. A lot of information not present in protein itself mRNA has intermediate amount because it must have the information to regulate and transcribe the protein protein has least because it doesn’t have regulatory information (used to generate protein) but does have some folding information- RNA Transcriptiono Transcription generates a single-stranded RNA molecules that is complementary to the DNA template strando RNA is synthesized 5’3’ (and so DNA is read 3’5’) by RNA polymerase, a complex multi-subunit enzymeo Eukaryotes have 3 different RNA polymerases, which synthesize different types of RNA’s (prokaryotes have only one RNA pol) RNA pol has a problem because Eukaryotic DNA is packaged into Chromatin needs promoterso Transcription initiates at special DNA sequences called promoters Binding sites for accessory proteins called general transcription factors, which help position RNA polymerase and start the processo Additional proteins are required to modify chromatin structure and fully active transcriptiono Extension of RNA chain requires elongation factors, which use ATP hydrolysis to assist RNA polymerase to move along chromatin Help to move it along Unwind DNA and needs energyo RNA transcription stops when RNA polymerase encounters a special DNA sequence called a terminatoro Most RNA post-transcriptional processing before it can be functionalo Unlike DNA, single-stranded nature of RNA allows it to fold into complex 3-D structures, comparable to tertiary structure of proteins- mRNA Processingo For RNA’s that are destined to encode proteins, substantial processing is required before they are considered mRNAo First modification occurs immediately after 5’ end of RNA exits polymerase: addition of 7-methylguanosine “cap” to 5’ end of RNA marks RNA as an mRNA-to-be Protects 5’ end of mRNA to indicate it will become messenger RNA 5’ end of mRNA to 5’ of guanosine by 3 phosphate So it is different from other RNAs by 3 5’-phosphates- Pre-mRNA Processingo Most protein-coding genes contain intervening sequences (introns) that interrupt the actual coding sequences (expressed sequences, aka exons)o Introns must be removed by the process of RNA splicing Carried out by complex machinery called spliceosome Made up of small nuclear ribonucleoproteins (snRNPs) – small nuclear RNAs (snRNAs) + multiple proteins Directed by RNA sequences found at intron-exon boundarieso Spliceosome assembles on pre-mRNA while it is still being transcribed, but splicing process may be delayedo Splicing process is extremely flexible—a given transcript may have many possible splicing patterns Different functionality comes from same gene by using alternative splicingo Once transcription is complete and RNA is released from RNA polymerase, the 3’end received a poly-A tailo First, the 3’ end of the original RNA is cleaved off, and then a series of ~200 A’s are added by a poly-A polymeraseo Poly-A binding proteins bind to the tail—important for export from the nucleus and later protein synthesis After = fully mature mRNA- mRNA Exporto RNA synthesis and processing all occurs in the nucleus, but protein synthesis occurs in the cytosolo Only fully processed, mature mRNA is exported from the nucleus—depends on removal of some proteins (e.g. snRNPs) and addition or retention of others (exon junction complex at splice sites (to know it has been spliced), cap-binding proteins (to know 5’ cap has been added), poly-A binding proteins (to know poly-A tail has been added), etc.)o Mature mRNA binds to nuclear export receptor, which guides it through the nuclear pore complex into the cytosol- “Other” RNAs?o mRNA represents only ~5% of cellular RNAo Up to 80% of cellular RNA is ribosomal RNA (rRNA)—makes up the structural and catalytic core of ribosomeso rRNA is synthesized by RNA pol III (18S, 5.8S and 26S) and RNA pol I (5S) rRNA is heavily processed and assembled with ribosomal proteins in the nucleolus, a non-membranous organelle within the nucleus Other “non-coding” RNAs have functions in pre-mRNA splicing (snRNAs), ribosome assembly (snoRNAs), protein synthesis (tRNAs), regulation of gene expression (siRNAs and miRNAs), telomere synthesis, and more- Splicing: mRNAs, rRNAs, tRNAs, snRNAs- Protein Translationo Once mature mRNA has been exported to the cytosol, it can be translated into protein by the ribosomeo Transcription = DNA RNA One-to-one correspondence of subunits Essentially the same language, with minor changes (U for T, ribose for deoxyribose)o Translation = RNA protein No one-to-one correspondence: 20 amino acids, but only 4 bases Totally different chemical languageo in order to accommodate 20 different amino acids, genetic code must use combination of at least 3 nucleotides double-nucleotide code: 4x4= 16 combinations (need 20) Triple-nucleotide code: 4x4x4 = 64 different combinations (more than enough so minimum it could be)o Each set of 3 nucleotides is called a codon. Since there are more codons
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