Eukaryotic Gene regulation Flashcards

this is the dumbest class Ive ever taken

Steps in Eukaryotic Gene Expression

1.) DNA transcribed into RNA

2.) RNA processing makes mature mRNA

3.) mRNA is translated to protein

4.) protein modification and action then occurs

Eukaryotic Cis-Acting Elements

1.) Promoters- DNA sequence that is adjacent to transcribed region where RNA pol binds

2.) Enhancers- DNA sequence far away from gene

Eukaryotic Enhancers

DNA sequence that can be far away from gene

bind proteins that augment or repress the basal level of transcription

may be located 5’ or 3’ to transcription start site

function when moved to different positions relative to promoter

Identifying Enhancers in Eukaryotes

construct recombinant DNA with enhancer sequence and GFP reporter gene

• used for testing in cell culture

• generating transgenic organism that has recombinant DNA in its genome

• enhancer increases GFP expression

Eukaryotic Promoters

DNA Sequence that is usually directly adjacent to the gene

site of RNA polymerase binding

often have TATA box: TATA-A/T-A-A/T

allows basal amount of transcription

Important information about Eukaryotic Cis Acting Elements

regulation happens at specific DNA sequences

Access to promoter region of a gene is regulated

enhancer region are part of mechanism for regulating access to promoter region/transcription initiation

reporter gene are used to determine specific DNA sequences and their role in gene regulation

Transcription Factors

sequence specific DNA binding proteins

bind to promoter and enhancer regions

recruit other proteins to influence transcription

Types of transcription Factors in Eukaryotes

1.) Basal Factors

2.) Activators

3.) Repressors

Basal Factors- Type of Transcription Factor in Eukaryotes

bind to promoters of protein-encoding genes and recruit in RNA polymerase

Basal Factor Order of Assembly at the Promoter

1.) TBP binds to TATA box

2.) TAF bind to TBP

3.)RNA pol II binds to TAF’s

Activators- Transcription factor in eukaryotes

transcription factors that bind to enhancers

responsible for much variation in levels of transcription of different genes

increase levels of transcription by interacting directly or indirectly with basal factors at the promoter

Mechanisms of Activator Proteins

1.) Stimulate recruitment of basal factors and RNA pol II to promoters

2.) stimulate RNA pol II initiation

3.) recruit coactivators to open chromatin structure

How does Chromatin Regulate Transcription Levels?

- if chromatin is intact and bound to promoter, there is no space for TATA binding proteins and transcription factors to interact with DNA

-euchromatin need to be removed so RNA pol can have access to promoter

How do Histone Modifying Proteins effect transcription in Eukaryotes

• Histone Acetylation opens chromatin- favors gene expression

  • HATS acytlate histone tails

  • coactivators

• Histone methylasetreansferases can activate or repress transrciption

  • coactivators and coirepressors

• histone methylation and acytlation are dynamic

  • modifications can be taken off rapidly by histone deacetlases or histone demethyalses

How do transcription factor effect transcription

transcription factors are proteins with multiple domains

activator proteins have 2 domains

• DNA binding domain- bind to enhancer

  • helix turn helix

  • zinc fingers

• Activator Domain- bind to other proteins

Dimerization domains also bring domains together so they can work together and with other proteins

Leucine Zipper

allows two proteins to interact and form a dimer that then interacts with DNA sequences

type of dimerization domain

Main Facts about Trans Acting Transcription Factors

proteins bind to promoter and enhancer DNA sequences to initiate transcription

general transcription factors recruite RNA polymerase

transcription factors can act as activators by

• recruit other factors

• initiate RNA pol. II

• remodel chromatin structure

they are multi-domain proteins with activation, DNA-binding, and often dimerization domains

Dimerization Domains

specialized for polypeptide-polypeptide interaction

• Homodimers: multimeric proteins made of identical subunits

• Heterodimers: multimeric proteins made of nonidentical subunits

Main Ideas about Gene Regulation in Eukaryotes

Gene regulation is the link between genotype and phenotype

regulation happens at all levels of eukaryotic gene expression

Transcription Factors in Eukaryotes- Repressors

suppress transcription by recruiting corepressor

this directly prevents RNA pol II complex from binding to promoter

corepressor can also close chromatin

Indirect Repressor

interferes with the activator

competition due to overlapping binding sites

repressor binds to activation domain

binding to activator keeps it in the cytoplasm

binding to activator also prevents homodimerization

Gene Regulation and Transcription Initiation

complex regulatory regions enable fine-tuning of gene expression

Insulators organize genomic DNA into a loop

• enhancers activate promoters located in the same loop

How are transcription factors Identified

identified using reporter genes (GFP)

mutations in a gene encoding an activator = reduce expression of the reporter

mutations in a gene encoding a represor= increase expression of the reporter

Function of trans acting proteins changes by:

allosteric interactions

• hormone receptor binds to enhancer only when bound to steroid hormone

Modification to transcription factors

Transcription factor cascades

Eukaryotic Regulation AFTER transcription

Can occur at the level of RNA

• splicing,m stability, and localization

Can occur at the level of protein

• synthesis,stability, modification, and localization

Different Small RNA’s in Eukaryotes

1.) miRNA’s

2.)siRNA’s

3.)piRNA’s

specialized RNAs that prevent expression of specific genes through comp base pairing

regulate stability and translation

miRNA’s (micro-RNAs)

targets mRNA

block mRNA translation/ destabilizes mRNA

most (blank) are transcribed by RNA pol II

the primary transcripts have double stranded stem loops

siRNA (small interferring RNA)

targets mRNA and nascent transcripts of chromosomal regions destined to become heterochromatin

block translation/destabilize mRNA and recruit histione-modifying enzymes to DNA recruiting in heterochromatin formation

piRNA (Piwi-Interacting RNA)

targets transposable element transcripts and transposable element promoters

effects the degradation of transposable elements in mRNA and facilitate histone modifications that inhibit transposable element transcription

miRNA Processing

Drosha:

• excises stem-loop from primar miRNA and generate pre-miRNA

Dicer:

• processes pre-miRNA to a mature duplex miRNA

regulating mRNA translation in response to Environment

• translation factors bind to mRNA to get it to the ribosomes for translation

• bind to 5’ cap and polyA tail

• factors lead to high mRNA translation

Translational control through Poly-A tail length

Negative Regulation of Translation: Poly-A tail is shorted (less translation)

Positive Regulation of Translation; Poly-A tail is lengthened (more translation)

How miRNA can down regulate expression of target genes

When complementarity is Perfect

• target mRNA is degreaded

• downregulation=+++

When complementary is imperfect

• translation of mRNA target is repressed

• Downregulation=+