【正文】 English Review: Control Of Expression In Eukaryotes ? Eukaryotic genes are controlled individually and each gene has specific control sequences preceding the transcription start site Eukaryotes Have Large Complex Geneomes ? The human genome is about 3 x 109 base pairs or ≈ 1 m of DNA ? Because humans are diploid, each nucleus contains 6 x 109 base pairs or ≈ 2 m of DNA ? That is a lot to pack into a little nucleus! Eukaryotic DNA Must be Packaged ? Eukaryotic DNA exhibits many levels of packaging ? The fundamental unit is the nucleosome, DNA wound around histone proteins ? Nucleosomes arrange themselves together to form higher and higher levels of packaging. Highly Packaged DNA Cannot be Expressed ? The most highly packaged form of DNA is “heterochromatin” ? Heterochromatin cannot be transcribed, therefore expression of genes is prevented ? Chromosome puffs on some insect chomosomes illustrate where active gene expression is going on Only a Subset of Genes is Expressed at any Given Time ? It takes lots of energy to express genes ? Thus it would be wasteful to express all genes all the time ? By differential expression of genes, cells can respond to changes in the environment ? Differential expression, allows cells to specialize in multicelled anisms. ? Differential expression also allows anisms to develop over time. DNA Cytoplasm Nucleus G AAAAAA Export Degradation etc. Control of Gene Expression G AAAAAA RNA Processing mRNA RNA Transcription Translation Packaging Modification Transportation Degradation Logical Expression Control Points ? DNA packaging ? Transcription ? RNA processing ? mRNA Export ? mRNA masking/unmasking and/or modification ? mRNA degradation ? Translation ? Protein modification ? Protein transport ? Protein degradation Increasing cost The logical place to control expression is before the gene is transcribed A “Simple” Eukaryotic Gene Terminator Sequence Promoter/ Control Region Transcription Start Site 3’ 5’ RNA Transcript Introns Exon 2 Exon 3 Int. 2 Exon 1 Int. 1 3’ Untranslated Region 5’ Untranslated Region Exons 5’ DNA 3’ Enhancers Enhancer Transcribed Region 3’ 5’ TF TF 3’ 5’ TF TF 5’ RNA RNA Pol. RNA Pol. Many bases Promoter Eukaryotic mRNA Protein Coding Region 3’ Untranslated Region 5’ Untranslated Region Exon 2 Exon 3 Exon 1 AAAAA G 3’ 5’ 3’ Poly A Tail 5’ Cap ? RNA processing achieves three things: ? Removal of introns ? Addition of a 5’ cap ? Addition of a 3’ tail Regulation of Gene Expression Six steps at which eucaryotic gene expression can be regulated. DNA RNA transcript mRNA mRNA inactive mRNA protein inactive protein NUCLEUS CYTOSOL transcriptional control RNA processing control RNA transport and localization control translation control mRNA degradation control protein activity control Translational control In principle, every step required for the process of gene expression could be controlled. Predominant form: Control of initiation of transcription. ?1. Transcriptional Initiation: ?This is the most important mode for control of eukaryotic gene expression. ?promoter elements : ?enhancer sequences can enhance the activity of RNA polymerase at a given promoter by binding specific transcription factors. 2. Transcript Processing and Modification: Eukaryotic mRNAs must be capped and polyadenylated, and the introns must be accurately removed. Several genes have been identified that undergo tissuespecific patterns of alternative splicing, which generate biologically different proteins from the same gene. ?3. RNA Transport: A fully processed mRNA must leave the nucleus in order to be translated into protein. ?4. Transcript Stability: Unlike prokaryotic mRNAs, whose halflives are all in the range of 15 minutes, eukaryotic mRNAs can vary greatly in their stability. Certain unstable transcripts have sequences that are signals for rapid degradation. ?5. Translational Initiation: Since many mRNAs have multiple methionine codons, the ability of ribosomes to recognize and initiate synthesis from the correct AUG codon can affect the expression of a gene product. ?Several examples have emerged demonstrating that some eukaryotic proteins initiate at nonAUG codons. This phenomenon has been known to occur in E. coli for quite some time, but only recently has it been observed in eukaryotic mRNAs. ?6. PostTranslational Modification: Common modifications include glycosylation, acetylation, fatty acylation, disulfide bond formations, etc. ?7. Protein Transport: proteins must be biologically active following translation and processing, they must be transported to their site of action. ?8. Control of Protein Stability: Many proteins are rapidly degraded, whereas others are highly stable. Specific amino acid sequences in some proteins have been shown to bring about rapid degradation Control of Eukaryotic Transcription Initiation ?Transcription of the different classes of RNAs in eukaryotes is carried out by three different polymerases. RNA pol I synthesizes the rRNAs, except for the 5S species. RNA pol II synthesizes the mRNAs and some small nuclear RNAs (snRNAs) involved in RNA splicing. RNA pol III synthesizes the 5S rRNA and the tRNAs. The vast majority of eukaryotic RNAs are subjected to posttranscriptional processing. ?The most plex controls observed in eukaryotic genes are those that regulate the expression of RNA pol IItranscribed genes, the mRNA genes. ?Almost all eukaryotic mRNA genes contain a basic structure consisting of coding exons and non