Control of Gene Expression

E. coli lac Operon

lacY DNA

Lac repressor

Regulatory gene

lacI

Binds Lac repressor

Binds RNA polymerase

Transcription initiation site

Promoter Operator

Sequences that control the expression of the operon

β-Galactosidase

lacZ

lac operon

Permease Transacetylase

Transcription termination site

lacA

Regulation of Inducible lac Operon

lacA DNA

mRNA Lac

repressor (active)

lacI

a. Lactose absent from medium

RNA polymerase cannot bind to promoter

Promoter Operator

Transcription blocked

When lactose is absent from the medium, the active Lac repressor binds to the operator of the lac operon, blocking transcription.

lac operon lacZ lacY

Inducible lac Operon

Transcription occurs

b. Lactose present in medium

DNA

Lac repressor

(active)

mRNA

Binding site for inducer

Allolactose (inducer)

RNA polymerase binds and transcribes operon

lacI Promoter Operator

When lactose is present in the medium, some of it is converted to the inducer allolactose. Allolactose binds to the Lac repressor, inactivating it so that it cannot bind to the operator. This allows RNA polymerase to bind to the promoter, and transcription of the lac operon occurs. Translation of the mRNA produces the three lactose metabolism enzymes.

Lactose metabolism enzymes

lacA

mRNA Translation

lacY

lac operon

lacZ

Inactive repressor

Repressable trp Operon

mRNA

DNA

a. Tryptophan absent from medium

Trp repressor (inactive)

mRNA

Regulatory gene

trpR

RNA polymerase binds and transcribes operon

Promoter Operator trpE

Transcription occurs

When tryptophan is absent from the medium, the Trp repressor is inactive in binding to the operator and transcription proceeds.

trp operon

trpD trpC trpB trpA

Tryptophan biosynthesis

enzymes

Translation

Regulation of the repressable trp Operon

trpE trpD trpC trpB trpA trpR Promoter Operator

trp operon

b. Tryptophan present in medium

Transcription blocked

DNA

mRNA

Trp repressor (inactive)

Tryptophan (corepressor)

RNA polymerase cannot bind to promoter

Trp repressor (active)

Tryptophan-binding site

When tryptophan is present in the medium, the amino acid binds to, and activates, the Trp repressor. The active repressor binds to the operator and blocks transcription.

Positive Regulation of lac Operon

DNA lacI

CAP site Promoter Operator

lacZ

a. Lactose present; glucose low or absent

mRNA

Lac repressor (active)

Allolactose (inducer)

Inactive repressor

cAMP

CAP

Active CAP

Transcription occurs

Translation

Lactose metabolism

enzymes

mRNA RNA polymerase

When lactose is present and glucose is low or absent, cAMP levels are high. cAMP binds to CAP, activating it. Active CAP binds to the CAP site and recruits RNA polymerase to the promoter. Transcription then occurs.

Positive Regulation of lac Operon

RNA polymerase binding site

b. Lactose present; glucose present

DNA

mRNA

Lac repressor (active)

Allolactose (inducer)

Inactive repressor

Inactive CAP

CAP site Promoter Operator

No transcription

RNA polymerase cannot bind

lacI lacZ

When lactose is present and glucose is present, cAMP levels are low. As a result, CAP is inactive and cannot bind to the CAP site. RNA polymerase then is unable to bind to the promoter, and no transcription occurs.

Gene Regulation Cytoplasm

Nucleus Chromatin

Pre-mRNA

DNA Determines which genes are translated

Transcriptional regulation • Chromatin remodeling to make genes accessible for transcription • Regulation of transcription initiation

Mature RNAs

Mature RNAs

Determines types and availability of mRNAs to ribosomes

Posttranscriptional regulation • Variations in pre-mRNA processing • Removal of masking proteins • Variations in rate of mRNA breakdown •RNA interference

Initiation of protein synthesis

Ribosome

Determines rate at which proteins are made

Translational regulation • Variations in rate of initiation of protein synthesis

New poly-peptide chains

Finished proteins

Protein breakdown

Determines availability of finished proteins

Posttranslational regulation • variations in rate of protein processing • Removal of masking segments • Varieties in rate of protein breakdown

Chromatin Remodeling Promoter

Promoter not accessible to proteins for transcription initiation

Promoter now accessible to proteins for transcription initiation Gene

Promoter

Remodeling complex

Chromatin remodeling exposes promoter

Activator

Gene

Regulatory sequence Nucleosomes

Organization of Eukaryotic Gene

DNA

Regulatory sequences

Enhancer

Promoter proximal elements

(regulatory sequences)

Promoter proximal

region

TATA box

Promoter

5' UTR

Transcription unit of gene

Exon Exon Exon Intron Intron

3' UTR

Transcription Complex on the Promoter

Transcription begins

Initial general transcription factor

Additional general transcription factors

Transcription complex

TATA box

Promoter

RNA polymerase

Site where transcription starts

DNA

DNA

The first general transcription factor recognizes and binds to the TATA box of a protein-coding gene’s promoter.

1

2 Additional general transcription factors and then RNA polymerase add to the complex, and then transcription begins.

Interactions Between Activators

Enhancer

Activators

Coactivator (multiprotein complex)

Interaction between activators at the enhancer, coactivator, and proteins at the promoter and promoter proximal region

Promoter Promoter proximal

region

Transcription initiation site

Gene

Maximal transcription

Gene

DNA loop

Activators

Combinatorial Gene Regulation

A unique combination of activators controls gene A.

DNA

Gene A, controlled by activators 2, 5, 7, and 8 binding to regulatory sequences in its enhancer

Gene A

Transcription Activators 2 5 7 8

2 5 7 8

Enhancer

Enhancer regulatory sequences

Combinatorial Gene Regulation

Gene B

Transcription Activators 1 5 8 11

Enhancer

Enhancer regulatory sequences

DNA 1 5 8 11

A different combination of activators controls gene B.

Gene B, controlled by activators 1, 5, 8, and 11 binding to regulatory sequences in its enhancer

Hormonal Gene Regulation

Hormone bound to receptor

Steroid hormone

Steroid hormone receptor

Protein

DNA

RNA polymerase

Transcription

Gene controlled by the steroid

hormone

Steroid hormone response element

Pre-mRNA

mRNA

mRNA + ribosomes

Gene Silencing via DNA Methylation

! The hemoglobin genes for instance are highly methylated and thus silenced in most vertebrate body cells except red blood cells.

! DNA methylation sometimes silences large blocks of genes or even whole chromosomes like one of the X chromosome in female mammals (Barr bodies)

! DNA Methylation underlies genomic imprinting in which either the paternal or maternal allele of a particular gene is silenced.

Genomic Imprinting: Prader-Willi and Angelman Syndrome

•  Specific partial deletion of chromosome 15 results in:

Angelman syndrome if it’s from the mother

Prader-Willi syndrome if the chromosome is from the father

From chapter 13

Histone acetylation activates genes

The DNA around acetylated histones is less tightly wrapped around the histones in the nucleosome and thus more accessible to DNA binding proteins including transcription factors and RNA polymerase.

Gene Regulation Cytoplasm

Nucleus Chromatin

Pre-mRNA

DNA Determines which genes are translated

Transcriptional regulation • Chromatin remodeling to make genes accessible for transcription • Regulation of transcription initiation

Mature RNAs

Mature RNAs

Determines types and availability of mRNAs to ribosomes

Posttranscriptional regulation • Variations in pre-mRNA processing • Removal of masking proteins • Variations in rate of mRNA breakdown •RNA interference

Initiation of protein synthesis

Ribosome

Determines rate at which proteins are made

Translational regulation • Variations in rate of initiation of protein synthesis

New poly-peptide chains

Finished proteins

Protein breakdown

Determines availability of finished proteins

Posttranslational regulation • variations in rate of protein processing • Removal of masking segments • Varieties in rate of protein breakdown

RNAi

Nucleus

Cytoplasm

Other proteins bind and degrade one RNA strand

Dicer

Dicer Precursor RNA folds into stem-loop

miRNA gene

miRNA precursor

miRNA binds to complementary target mRNA

Degradation of mRNA

Inhibition of translation

Stepped Art

Double-stranded RNA

Protein Degradation

Development of Colorectal Cancer

Colon

Colon wall

Loss of tumor- suppressor gene APC (or other)

Normal colon epithelial cells

Small benign growth (polyp)

Larger benign growth (adenoma)

Activation of ras oncogene

Loss of tumor- suppressor gene DCC

Loss of tumor- suppressor gene p53

Additional mutations

Malignant tumor (carcinoma)

Add Cytoplasmic Determinants, Induction and Development