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143 Cards in this Set
- Front
- Back
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genomic imprinting
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patterns of cytosine methylation in DNA modulate gene expression
inherited through cell division and through gametes |
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RNA binding proteins determine patterns of...
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alternative splicing
allows primary transcripts to be processed into different mature mRNAs |
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promoter
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sequence near transcription start site
attracts RNA polymerase to gene |
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function and structure of eukaryotic polymerase II promoters
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bind RNA polymerase indirectly
contain a TATA box |
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TATA box
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~7 nucleotide sequence
all T's and A's |
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basal factors
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transcription factors - bind the promoter
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example of basal factor
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TATA-box binding protein
and TBP (^) associated factors these proteins form basal factor complex |
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basal factor complex function
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attracts RNA polymerase to promoter
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enhancer
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cis acting DNA element
regulates transcription of 1 or more genes (may be located thousands of base pairs from promoter) contain binding sites for transcription factors responsible for spatiotemporal specificity of transcription |
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activators
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eukaryotic transcription factor
stabilizes basal factor complex at promoter or recruits coactivators |
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coactivators
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proteins that displaces nucleosomes from promoter
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repressors
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eukaryotic transcription factor
recruit corepressors |
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corepressors
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proteins that either disrupt basal factor complex at the promoter
or close chromatin at the promoter |
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indirect repressors
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prevent transcription of gene by inhibiting the function of the activator
do not bind to DNA |
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GFP (green fluorescent protein)
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jellyfish protein
used as reporter protein by researchers glows in live cells (w/UV light) |
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insulators
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DNA sites
bind proteins - organize chromosome into loops |
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which promoters can enhancers interact with?
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only those in the same loop
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DNA methylation
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in vertebrates
addition of methyl group to cytosine base in CpG sequence to form 5-methylcytosine |
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CpG sequence
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5' CG 3'
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CpG island
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cis acting transcriptional regulatory element
found upstream of some eukaryotic genes rich in CpG sequences |
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nonmethylated CpG islands
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bind activators
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methylated CpG islands
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do not bind activators
may bind repressors |
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epigenetic phenomenon
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heritable change in gene expression not caused by mutation in gene's base pair sequence
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genomic imprinting
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epigenetic phenomena
due to methylation of transcriptional regulatory elements maternal or paternal origin of an allele affects its expression in the progeny |
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effect of imprinted alleles
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transcriptionally silenced
(not transcribed by RNA poly) |
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miRNAs (micro-RNAs) and siRNAs (small interfering RNAs) structures and functions
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small, trans acting RNAs
regulate stability or translation of specific mRNA targets through complementary base pairing |
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how miRNAs are produced
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by processing of long primary transcripts (pre-miRNAs)
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how siRNAs are produced
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by processing of double stranded RNAs
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RISC (RNA induced silencing complex)
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substrate which miRNAs or siRNAs bind to
guides it to target mRNA |
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piRNAs (Piwi-interacting RNAs)
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small, trans acting RNAs
guide complexes containing Piwi proteins to transposable elements (TEs) or to TE transcripts limit mobilization and transposition of TEs |
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RNA interference
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research technique
reduces/turns off gene expression that exploits the siRNA pathway |
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RNA interference procedure
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double stranded RNA (dsRNA) corresponding to base pair sequence of gene is introduced into cell or organism
dsRNA processed into siRNA by enzymes guides RISC to target gene's mRNA leading to degradation of that mRNA |
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enhancers normally found where?
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upstream from promoter (before it)
can be downstream of entire transcriptional unit or even introns of the gene |
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insulators normally found where?
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flank the gene (on outsides)
*if the gene's enhancer(s) do not activate expression of other gene's also |
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start and stop codons normally found where?
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can be in any exons
*several exons can be devoted to 5'UTR and/or 3'UTR |
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Eukaryotic promoter characteristic
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bind basal factor complex
^attracts RNA polymerase |
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Prokaryotic promoter
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binds RNA polymerase directly
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Euk enhancers
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determine temporal and spatial specificty of transcription
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prok operators and sigma factors
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control transcriptional response to environment
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euk insulators
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organize genomic DNA to control enhancer/promoter interactions
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prok enhancers/insulators
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do not exist
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euk activators
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bind enhancers
stabilize the interaction of basal complex with the promoter or recruit coactivators that clear the promoter of nucleosomes |
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prok activators
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bind DNA near promoters
interact directly with RNA poly to stabilize interaction between RNA poly and promoter |
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euk repressors
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bind enhancers
and recruit corepressors that either destabilize interaction of RNA poly with basal complex or recruit nucleosomes to promoter |
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prok repressors
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bind operator and either inhibit RNA poly from binding to promoter
or prevent RNA poly from initiating transcription |
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prok corepressors
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effectors that bind repressors and alter their conformation so that they can bind to DNA
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euk alternate RNA splicing
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can generate different proteins from single gene
and be regulated by cell type specific splicing factors |
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prok RNA splicing
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does not occur
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euk small RNAs
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miRNAs, siRNA, piRNAs
control stability or translation of mRNA targets through complementary base pairing |
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prok small RNAs
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sRNAs regulate translation of mRNA targets through complementary base pairing
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euk transcription and translation (where do they occur)
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in different cellular compartments
mechanisms such as attenuation cannot occur |
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prok transcription and translation occur...
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simultaneously
allowing mechanisms such as attenuation to exist |
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in fusion gene diagrams
p = ? o = ? colored boxes = ? arrows = ? |
p = promoter
o = operator colored boxes = DNA sequences from different genes arrows = transcription start sites |
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lacZ encodes for what
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B-galactosidase
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GFP encodes for what
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jellyfish green fluorescent protein
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lacZ and GFP are both examples of what
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reporter genes
used in both prokaryotes and eukaryotes |
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basal factors
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bind to promoters
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repressors
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bind to enhancers
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CpG
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site of DNA methylation
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imprinting
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pattern of expression depends on which parent transmitted the allele
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miRNA
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prevents or reduces gene expression posttranscriptionally
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coactivators
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bind to activators
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epigenetic effect
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change in gene expression caused by DNA methylation
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insulator
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organizes enhancer/promoter interactions
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enhancer
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activates gene transcription temporal and tissue specifically
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ChIP-Seq
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identifies DNA binding sites of transcription factors
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RNA poly II
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transcribes all protein coding genes (mRNAs) and micro-RNAs
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promoters
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DNA sequence usually directly adjacent to the gene
cis acting regulatory element binds RNA poly often has TATA box allows basal level of transcription |
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enhancers
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DNA sequence (can be far away from gene)
cis acting regulatory element can increase of decrease transcription can be at 5' or 3' can function when moved to different positions relative to the promoter |
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transcription factors
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sequence specific DNA binding proteins
bind promoters and enhancers recruit other proteins to influence transcription trans acting |
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TFIIs
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RNA general transcription factors
work with RNA poly II |
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TFIID components
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TBP + TAFs
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RNA poly II holoenzyme
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core RNA poly II CTD (C terminal domain)
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Mediator
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multiprotein complex
enables RNA poly II to respond to transcriptional activators present in all eukaryotes |
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activators
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transcription factor
bind to enhancers increase levels of transcription by interacting directly or indirectly with basal factors at the promoter recruit RNA pol II to basal promoter |
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mechanisms of activator effects on transcription
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stimulate recruitment of basal factors and RNA poly II to promoters
recruit coactivators to open chromatin structure (displace nucleosomes) |
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dimerization domains
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specialized for polypeptide-polypeptide interactions
located on activators |
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homodimers
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multimeric proteins made of identical subunits
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heterodimers
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multimeric proteins made of nonidentical subunits
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can Myc/Myc homodimers form?
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no
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can Max/Max homodimers form?
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yes
bind DNA but do not act as activators |
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can Myc/Max heterodimers form?
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yes
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when Myc is present, which heterodimer forms?
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Myc/Max will form
and activate transcription |
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more affinity- Myc/Max or Max/Max
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Myc/Max
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repressor proteins
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recruit corepressors
and suppress transcription |
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corepressor functions
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prevent RNA poly II from binding to promoter
and modify histones to close chromatin strucutre |
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function of trans acting proteins changes by...
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allosteric change in steroid hormone enables transport into the nucleus
and modification of transcription factors (ex. phosphorylation) |
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indirect repressor
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interferes with function of activator
-competition -repressor binds to activation domain -binding to activator + keeping it in cytoplasm -binding to activator + preventing homodimerization |
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reporter genes
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identify enhancers in euk
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enhancers can be identified by:
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making recombinant DNA with enhancer sequence fused to reporter gene such as GFP
make transgenic organism that has recombinant DNA in genome |
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mutations in gene encoding an activator does what to expression of reporter
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decreases reporter expression
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mutations in gene encoding an repressor does what to expression of reporter
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increases reporter expression
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chromatin immunoprecipitation-sequencing (ChIP-Seq)
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1.crosslink DNA and chromatin
2. fragment DNA 3. specific antibody binds to protein 4. purify these complexes 5. sequence DNA |
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insulators |
sequences between enhancer and promoter block access to promoter |
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enhancer regulates what genes |
those that do not get blocked by insulator |
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how do insulators organize genomic DNA |
into loops |
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enhancers activate what promoters? |
only those on same loop |
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genomic imprinting |
expression of allele depends on the parent that transmits it |
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methylation at imprinting control regions causes what... |
silences gene expression |
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Paternally imprinted |
paternal allele is silenced not passed down on pedigree |
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dna methylation |
methyl group added to cytosine in 5'CpG 3' dinucleotide by DNA methyl transferases (DNMTs) |
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CpG islands |
regions with high concentration of CpG dinucleotides near genes - are unmethylated (activator binds and blocks access by DNMTs) chromatin is open and transcription is activated |
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unmethylated |
transcription occurs |
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CpG vs absence of activators |
becomes methylated |
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Methyl-CpG-binding proteins (MeCPs) |
bind and close chromatin structure no activator to shield from DNA methyl transferase |
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epigenetic phenomenon |
heritably alter gene expression without changing DNA sequence DNA methylation is example ^cytosine methylation pattern is copied during DNA replication |
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epigenetic imprints vs mammal lifespan |
remain throughout |
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epigenetic imprints vs germ cells |
reset as each generation during meiosis, imprints erased and new ones are set |
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imprinting mechanisms |
insulator mechanism ncRNA mechanism |
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insulator mechanism of imprinting |
insulator controls transcription of gene nonmethylated maternal insulator is functional and transcription is silenced (insulator blocks transcription) methylated paternal insulator is nonfunctional and transcription is active |
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insulator mechanism general |
methylated insulator = nonfunctional = transcription is active and vice versa |
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ncRNA mechanism of imprinting |
nonmethylated paternal CpG island allows production of antisense RNA - silences transcription methylated maternal CpG prevent production of antisense RNA - activates transcription |
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ncRNA mechanism general |
nonmethylated CpG = antisense RNA produced = transcription silenced |
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Praeder Willi syndrome |
deletion inherited from father chromosome 15 |
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Angelman syndrome |
deletion inherited from mother chromosome 15 |
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(in general) small RNAs are responsible for... |
RNA interference act post transcriptionally prevent expression of specific genes through complementary base pairing |
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miRNAs
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transcribed by RNA poly II from non coding regions (introns) of protein coding genes --> primary transcripts have double stranded stem loops |
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Drosha |
removes stem loop from primary miRNA to generate pre-miRNA |
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Dicer |
processes pre-miRNA (removed by primary-miRNA) to mature duplex miRNA |
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function of one strand of the mature duplex miRNA made by dicer |
incorporated into miRNA induced silencing complex (RISC) |
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two ways miRNAs can decrease expression of target genes |
1. when complementarity is perfect 2. when complementarity is imperfect |
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when complementarity is perfect |
mRNA cleavage target mRNA is degraded (decreased expression) |
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when complementarity is imperfect |
translocational repression translational of mRNA target is repressed (decreased expression) |
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dsRNAs |
precursors of siRNAs transcription of both strands of endogenous genomic sequence or arise from exogenous virus
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dsRNAs are processed by... |
Dicer |
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what do processed dsRNAs form |
ribonucleoprotein complexes with Argonaute proteins interfere with gene expression or may destroy viral mRNAs |
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siRNAs vs experimental tools |
selectively knock down expression of target genes |
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posttranscriptional regulation vs what levels it can occur on |
RNA level - splicing stability, localization protein level - synthesis, stability, localization |
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piRNAs |
make piwi proteins that form complexes with RNAs |
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piwi - RNA complex function |
modify histones to interfere with transposable element transcription or degrade transposable element RNAs |
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(in general) piRNAs function |
minimize transposable element mobilization |
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what determines sex, fertility and viability in Drosophila |
ratio of X chromosomes to autosomal chromosomes |
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X:A ratio influences sex through 3 different pathways |
1. male vs female appearance and behavior 2. development of germ cells as eggs or sperm 3. dosage compensation - males have increased transcription of x linked genes |
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what is weird about males in drosophila |
have increased dosage of x chromosome |
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fruitless mutation in males |
male with aberrant courtship behavior |
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sex lethal Sxl gene encodes what |
protein required for female development early embryos - only transcribed in females |
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sex lethal promoter |
"counts" x chromosomes xx cells express Sxl from establishment promoter (Pe) (female development) genes encoding activators of Pe are on X chromosome |
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Pm promoter |
transcribe SxI in males - transcript has stop codon females - alternative splicing removes exon and protein is produced |
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SxI protein triggers what splicing cascades (2) only in females (stop codon occurs in males) |
1. regulates splicing of transformer (tra) mRNA 2. ^ Tra regulates splicing of double sex mRNA |
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Dsx-F and Dsx-M relationship |
have opposite effects one is activated and transcribed and other is not transcribed |
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how are Fru-M and Fru-F produced |
by alternative splicing of same primary transcripts |
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expression of Tra and Tra2 in females vs males |
expressed in females - block splice acceptor site causes different exons to be translated into different proteins for males and females |
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ef you |
16 |
