REGULATION OF GENE EXPRESSION

By:

Khaled El MasryAssistant Lecturer of Human Anatomy & Embryology

Mansoura Faculty of Medicine

What is the GENE????

A gene is the sequence of nucleotides in DNA encoding one polypeptide chain or one mRNA molecule.

Gene expression is carried out in 2 steps:

1 .transcription.2 .translation.

Factors inducing gene expression:

1 .Environmental factors as heat and light.2 .Signaling molecules as hormones and

growth factors.

HEAT:Exposure to high temperature

Transcription of heat shock genes

Synthesis of heat shock proteins

Stabilize the internal cellular environment

LIGHT:In plants exposure to light

Activate transcription of the gene for

Ribulose Carboxylase The enzyme that plays a critical

role in

PHOTOSYNTHESIS

HORMONES

”STEROID HORMONES”

Hormone receptor complex

Act as a transcription factor

“PEPTIDE HORMONES”

Activate a signaling system

Regulation of eukaryotic gene expression is dependent on 2 factors:

1 .Regulatory elements:Promoters & Enhancers

2 .Transcription factors:Basal & Special

Regulatory elements

PromotersThe region necessary to initiate transcription.

Consists of short nucleotide sequence that serve as the recognition point for binding of RNA polymerase.

Located immediately adjacent to the genes they regulate, upstream from the transcription startpoint.

There are significant differences in number , orientation and distance between promoters in different genes.

Promoters for RNA polymerase II include:

TATA box,CAAT box,GC box,

&Octamer box.

Promoters for RNA polymerase I & III have a different sequence and bind different transcription factors.

Importance Structure Site

Mutations in this sequence greatly reduce transcription

(Loosing the ability to bind to transcription factors)

8 bp sequences composed only of T=A pairs.

25-30 bp upstream

(from the initial point of transcription

TATA box

Mutations in this sequence greatly reduce transcription

CAAT or CCAAT sequence.

70-80 bp upstream

(from the initial point of transcription

CAAT box

Documented by mutational analysis

GGGCGG sequence, often present in multiple copies.

110 bp upstream(from the initial

point of transcription

GC box

Affects the efficiency of promoter in initiating transcription.

ATTTGCAT sequence.

120-130 bp upstream

(from the initial point of transcription

Octamer box

2 .Enhancers

DNA sequences interact with regulatory proteins

increase the efficiency of initiation of transcription

increase its rate .

Enhancers:

up to several hundred bp long).) Large >>>>

Tissue- specific >>>> ( stimulate transcription only in certain tissues).

Mechanism of action of Enhancers???

1 .The proteins that bind to enhancers affect the activity of proteins that bind to promoters???.

2 .Enhancers may allow RNA polymerase to bind to DNA and move along the chromosome till it reaches a promoter site.

3 .May respond to molecules outside the cell ( e.g : steroid hormones).

4 .May respond to molecules inside the cell ( e.g : during development thus the gene participates in cell differentiation).

How enhancers can control transcription although they

are located away from the transcription site???

Enhancers bind to transcription factors by at

Least 20 different proteins

Form a complex

change the configuration of the chromatin

folding, bending or looping of DNA.

DNA looping will bring the distal enhancers close to the promoter site to form activated transcription complexes, then the transcription is activated, increasing the overall rate of RNA synthesis.

Differentiation ( ) Enhancers and Promoters

1 .The enhancers affect the gene expression independent of their position or orientation.

2 .The enhancers operate from a distance away from their target genes.

Transcription factors

Def: . “they are proteins essential for initiation of

the transcription, but they are not part of RNA polymerase molecule that carry out

the transcription process.”

Function:

Each RNA polymerase requires a number of transcription factors which help in:

1 .Binding of the enzyme to DNA template.

2 .Initiation and maintenance of transcription.

3 .Control the rate of gene expression.

Structure & Mechanism of action

These proteins contain 2 functional domains (a.as that perform specific function).

1 .DNA binding domain: binds to DNA sequences present in regulatory regions (e.g : TATA binding protein).

2 .Transcriptional activating domain: activate transcription via protein-protein interaction

Types of transcription factors:

1 .Basal transcription factors:

The initiation of transcription by RNA polymerase II requires the assistance of several basal transcription factors.

Each of these proteins binds to a sequence within the promoter to facilitate the proper alignment of RNA polymerase on the template strand of DNA.

The basal TFs must interact with the promoters in the correct sequence to initiate transcription effectively.

TFIID is the 1st basal TF that interact with the promoter ; it contains TATA- Binding Protein.

Followed by TFII B, F, E, H & J.

2 .Special TFs:Involved in regulation of heat, light, and hormone inducible genes.They bind to:

a. enhancers.b. Basal TFs.c. RNA polymerase that bind to the gene promoter.

Thus, special TFs can regulate the transcriptional activity of the gene.

Chromatin Conformation and Gene Expression

The normal structure of the chromatin suppresses the gene activity, making the DNA relatively inaccessible to transcription factors, and thus active transcription complex can’t occur.Thus chromatin remodeling is needed

(it is a change in chromatin conformation in which proteins of nucleosomes are released from DNA , allowing DNA to be

accessible for TFs and RNA polymerase .)

Ways of Chromatin Remodeling Inactive chromatin remodeled into active

chromatin by 2 biochemical modifications:

1 .Acetylation of histone proteins by histone acetyl transferases which loosen the association between DNA and histone.

2 .Specialised protein complexes disrupt

the nucleosome structure near the gene’s promoter site.

(this protein complex slides histone along DNA

transfer the histone to other location on DNA molecule.

Active chromatin can be deactivated by 3 biochemical reactions:

1 .Histone deacetylation ( catalysed by histone deacetylase).

2 .Histone methylation ( catalysed by histone methyl transferases).

3 .Methylation of some DNA nucleotides by DNA methyl transferases.

(Chromatin subjected to these modifications tends to be transcriptionaly silent)

Post-transcriptional regulation of gene expression by RNA INTERFERENCE

siRNA & miRNA

These molecules 21-28 bp long.

Able to interact with specific mRNA molecules.

This interaction occurs by base pairing ( ) a single strand of iRNA and a complementary sequence in the mRNA molecule.

siRNA

Base pair perfectly with the target

sequence in mRNA

mRNA cleavage and degradation

miRNA

Base pair imperfectly with the target

sequence in mRNA

mRNA inhibition of translation