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prokaryotic gene expression differs in many ways from eukaryotic differences in polymerases transcription factors do not exist in bacteria initiation of translation can differ
eukaryotes small ribosomal subunit binds the cap prokaryotes binds a specific sequence at the 5 end initiation factors differ
What is a gene?
Mendel genetic information that gives rise to a specific phenotype Morgan assigned genes to specific chromosomes (loci) sequence of DNA nucleotides within the chromosome codes for a specific polypeptide sequence through transcription into mRNA and translation
but can also be transcribed into rRNA, tRNA, snRNA (non translated RNAs)
promoter upstream of the coding region not transcribed enhancer regions repressor regions
three downstream genes lacZ, lacY and lacA in the absence of lactose
Chromatin Structure
packing of DNA into histones can regulate its ability to be transcribed
closer the promoter is to the nucleosome less likely it will be transcribed attachment of the chromatin to the nuclear matrix (nuclear lamina) or the scaffolding of a chromosome can affects its transcription
genes within heterochromatin are usually not expressed chemical modifications of histones can modify the packing of the DNA
tails of the histones project outward modifications:
acetylation acetyl groups (COCH3) attached to the lysines of the tails
deacetylation is their removal- e.g. HDAC (histonedeacetylase) acetylation neutralizes the histones positive charge no longer bind to the neighbouringhistone loosens the packing and allows access to the transcription machinery acetylation = gene expression
methylation methyl groups (CH3) attached to the tails promotes condensation of chromatin
direct methylation of the DNA bases can also occur usually cytosine can inactivate the gene i.e. not transcribed longer-term methylation/shut-down is important for development of the embryo methylation retained over mitosis daughter DNA is methylated just like the parent so the pattern of methylation can be passed on methylation patterns can also develop de novo DNA traits produced by things other than the DNA sequence = epigenetic inheritence can be reversed in succeeding generations = ???? mechanism????
Transcriptional initiation
after the chromatin structure is changed to allow transcription regulate the initiation of transcription 1st level promote the binding of the RNA polymerase to the promoter
upstream of the 1stexon site for RNA polymerase II binding assisted through the binding of other proteins = transcription factors some transcription factors regulate transcription of ALL genes = general transcription factors
some binding the DNA directly sequence called the TATA box others bind RNA polymerase II others bind other transcription factors e.g. CBFA-1 TF for osteogenic gene expression
general TFs, specific TFs, RNA pol II and other transcriptional mediators come together to form a large protein complex near the promoter = transcriptional initiator complex activator TFs have two things in common
1. DNA binding domain 2. Activation domain(s) bind other TFs or mediator proteins can bind directly to the control elements blocking the binding of activator TFs turn off transcription even when the activator TF is bound block the formation of the initiator complex
repressor TFs
so it isn t the sequence of these elements that is important it is the combination of the control elements in the gene that is it is also the TFs they bind
many cell types can express the same TFs not all will be active
Post-transcriptional regulation
occurs after transcription 1. RNA processing
transcription of DNA within the gene -> primary RNA transcript (Figure 8.8) RNA is processed into mRNA with its coding region, its 5 and 3 UTRs
5 cap and poly A tail added on introns are spliced out how the splicing occurs can give multiple mRNA transcripts spliced regions determined by regulatory proteins cell-type specific
2. mRNA degradation
life span of mRNA within the cell determines its availability for translation
varies from mRNA to mRNA e.g. Hb mRNA = 120 days!!
initiation of breakdown starts with the removal of the polyA tail then triggers the removal of the 5 cap nucleases then chew up the remaining RNA degradation can be sped up by the binding of microRNA sequences (miRNA) to their complement in the mRNA
blocks the translation of the targeted mRNA and promotes its breakdown
3. translation initiation
binding of regulatory proteins to the untranslated regions of the mRNA prevents binding of ribosome lack of a 5 cap lack of a poly A tail can block translation also important for ribosome binding
Post-translation regulation
protein processing & degradation degradation rate = half-life of the protein
attachment of a small protein called ubiquitin these proteins are targeted for degradation by the proteosome(giant protein complexes within the cell)
modifications essential for function occur in the ER and Golgi apparatus ER modifications:
1. formation of disulfide bonds
help stabilize the tertiary and quaternary structure of proteins misfolded proteins remain in the cytoplasm and are degraded only properly folded proteins get transported to the Golgi for additional processing and transport
2. folding of the peptide chain 3. addition and processing of carbohydrates (glycosylation) 4. breakage of specific peptide bonds proteolytic cleavage 5. assembly into multimeric proteins (more than one chain)
Golgi modifications:
1. glycosylation addition of chains of carbohydrates to specific amino acids 2. trimming of the protein
pattern formation studied by genetically manipulating the fruit fly and observing the resulting progeny
pioneered by Ed Lewis in the 1940s mapped mutations to the genome and altered the genome to make more mutants discovered what are called homeobox genes (or homeotic genes) the genes of pattern determination many genes named after their mutations e.g. wingless, hairy some have very bizarre names e.g. sonic hedgehog, patched, jagged, bicoid
first determined by cytoplasmic factors encoded by maternal effect genes = a gene that when mutated in the mother results in a mutated offspring despite the offspring s genome these gene products are placed in the egg prior to fertilization in the ovary the maternal effect genes controlling orientation of the egg = egg-polarity genes e.g. bicoid= two tailed mutation in this gene results in a fly that lacks the front half of its body posterior structures at both ends!!! found that the product of bicoid concentrates at the site that will become the anterior half of the body = morphogen gradient morphogen gradient = chemicals that help determine the form of the embryo figure 18.19 decreasing amounts of bicoid protein from anterior to posterior end of embryo