The Lac Operon

Genes at Work

In a multicellular organism, every cell contains all the genetic information needed to make every protein that the organism needs.

Many cells become specialised and so do not need to use every protein that is potential available and so it would be a waste of energy to make every protein all the time.

In particular what would be the point of making enzymes in the absence of a substrate to catalyse.......????

Some genes are transcribed and translated all the time....

eg genes for constitutive proteins and enzymes such as those that catalyse the stages in glycolysis and Kreb’s cycle

Other genes are only transcribed and translated only when they are required….

eg genes for inducible proteins and enzymes – β-galactosidase which breaks down lactose in the bacteria E.coli

Therefore genes can be switched on and switched off – this is called GENE REGULATION

JACOB and MONOD’S THEORY of GENE REGULATION

During the 1950’s Jacob and Monod found that the bacterium E.coli would only produce the enzyme β-galactosidase if its substrate lactose was present in the growth medium and at the same time glucose was absent

Before continuing first check out these really good websites!!http://dnaftb.org/33/concept/index.htmlhttp://highered.mcgraw-hill.com/olc/dl/120080/bio27.swfThe LAC OPERON

An operon consists of a group of closely linked genes that act together and code for the enzymes that control a particular metabolic pathway.

The production of the β-galactosidase is controlled by the lac operon along with two other proteins which are required for absorption and metabolism of lactose

The three genes are transcribed together in one mRNA

All three genes are switched OFF when NO LACTOSE PRESENT.

All three genes are switched ON, transcribed and translated when LACTOSE is PRESENT

The operon consists of the following;

A PROMOTOR REGION - RNA polymerase binds to start transcription

An OPERATOR REGION – where an inhibitor/ repressor protein binds

STRUCTURAL lac GENES ( z, y and a ) – for β-galactosidase, a membrane transport protein and another enzyme

There is also the i GENE (REGULATOR/REPRESSOR GENE) on another part of the bacterial chromosome that codes for a REPRESSOR PROTEIN which binds to OPERATOR REGION and INHIBITS transcription of the three lac genes z,y and a.

Simplified diagram....

E.coli prefer to respire glucose as it is more efficient for them, therefore in the presence of glucose and lactose transcription of β-galactosidase is inhibited.........

In the presence of lactose only.....

The operon is both negatively and positively controlled;

Negative control;

Lactose present / glucose absent

lac operon switched on as lactose binds to the repressor protein preventing it binding to the operator and so transcription inhibition is reversed....see above

Positive Control

cAMP is a regulatory substance which can bind to CRP (cyclic AMP receptor protein), which then activates it causing a conformational change and exposing the DNA binding site on the CRP molecule

binding of activated CRP to DNA helps RNA polymerase to bind to the promoter region to start transcription of the genes

levels of cAMP are only high enough to activate CRP in the absence of glucose

Genes in Development

Homeobox Genes

How a single fertilized cell develops into a complex organism like a fly, a mouse, or a human being has long been one of biology's greatest mysteries.

Von Baer in the early 19th century observed that all vertebrates, from salamanders to humans, look very similar in the early stages of their embryonic development.

At about the same time, French zoologist Geoffroy Saint-Hilaire declared that all animals have the same body plan. Because the main nerve cord is in the front part of insects and in the back part of vertebrates, Saint-Hilaire hypothesized that vertebrates are essentially upside-down invertebrates!

Like so many breakthroughs in genetics, this one came from the humble fruit fly, Drosophila melanogaster, a laboratory favourite because it reproduces rapidly, has only 4 chromosomes, and readily exhibits mutations induced by inbreeding and x-rays.

Fruit flies are highly specialized insects with 2 wings and 3 body segments.

Their ancestors had 4 wings and many body segments. The fruit fly embryo starts out with a series of 16 equal-sized segments. Various segments merge to make the 3 segments we recognize as the head, thorax, and abdomen.

Certain genes called homeotic genes are amazingly similar in structure and function in all animals; they serve as molecular architects and direct the building of bodies according to definite detailed plans

In the 1940s, American biologist Edward B. Lewis began studying the homeotic genes that affect segmentation in Drosophila.

He found that mutations in a cluster of genes, caused duplication of a body segment with an extra pair of wings.

These mutations were weird and hard to explain because hundreds of different genes participate in the formation of a body segment and wings. Yet here were single mutations creating new body parts and eliminating others.

These genes were acting as master switches, turning on and off arrays of other genes involved in body shape, and controlling the number, pattern, position, and fusion of segments and appendages.

In the late 1970s, German biologists Christiane Nüsslein-Volhard and Eric F. Wieschaus

sequenced the homeotic genes controlling the development of the fruit fly's body. They observed that:

in each of these genes a particular DNA sequence of 180 bases long is virtually identical.

this DNA sequence, called the homeobox, translates into a protein sequence 60 amino acids in length.

this protein sequence binds to DNA and switches on and off the process of transcription, the expression of genes into proteins.

by controlling the transcription in all cells, homeobox (Hox) genes act as master switches determining cell fates, growth, and development.

For their work on the homeobox genes, Lewis, Nüsslein-Volhard, and Wieschaus received the 1995 Nobel Prize for physiology or medicine.

Hox genes in mice and humans are very similar in number and chromosomal arrangement. It is remarkable that only about 40 genes out of a total of about 100,000 control most of the development, architecture, and appearance of the body plan of complex mammalian species.

Definitions

Hox genes: Hox genes are a subgroup of homeobox genes. In vertebrates these genes are found in gene clusters on the chromosomes. In mammals four such clusters exist, called Hox clusters. The gene name "Hox" has been restricted to name Hox cluster genes in vertebrates.

NB: homeobox genes are NOT Hox genes, Hox genes are a subset of homeobox genes.

HOX cluster: The term Hox cluster refers to a group of clustered homeobox genes, named Hox genes in vertebrates, that play important roles in pattern formation along the anterior-posterior body axis.

homeodomain: a DNA-binding domain, usually about 60 amino acids in length, encoded by the homeobox.

homeobox: a fragment of DNA of about 180 basepairs (not counting introns), found in homeobox genes.

Apoptosis

For every cell, there is a time to live and a time to die. There are two ways in which cells die: They are killed by injury, infection etc They are induced to commit suicide.

Death by injury - Necrosis

Cells that are damaged by injury, such as by mechanical damage exposure to toxic chemicals

undergo a characteristic series of changes:

The cell and its organelles swell (because the ability of the plasma membrane to control the passage of ions and water is disrupted).

The cell contents leak out, leading to Inflammation of surrounding tissues.

Death by suicide – Apoptosis

The pattern of events in death by suicide is so orderly that the process is often called programmed cell death or PCD or APOPTOSIS

Cells that are induced to commit suicide: shrink; develop bubble-like blebs on their surface; the chromatin (DNA and protein) in their nucleus degrades; mitochondria break down with the release of cytochrome c whole cell breaks down into small, membrane-wrapped, vesicles release ATP binds to receptors on phagocytic cells like macrophages and attract them

to the dying cells (a "find-me" signal"). the phospholipid phosphatidylserine, which is normally hidden within the plasma

membrane, is exposed on the surface. this "eat me" signal is bound by other receptors on the phagocytes which then engulf

the cell fragments. the phagocytic cells secrete cytokines that inhibit inflammation (e.g., IL-10 and TGF-β)

Necrosis VS apoptosis.mpg - YouTube

Why should a cell commit suicide?

There are two different reasons.

1. Programmed cell death is needed for proper development

The reabsorption of the tadpole tail at the time of its metamorphosis into a frog occurs by apoptosis.

The formation of the fingers and toes of the feotus requires the removal, by apoptosis, of the tissue between them.

The sloughing off of the inner lining of the uterus (the endometrium) at the start of menstruation occurs by apoptosis.

The formation of the proper connections (synapses) between neurones in the brain requires that surplus cells be eliminated by apoptosis

2. Programmed cell death is needed to destroy cells that represent a threat to the integrity of the organism.

Cells infected with viruses One of the methods by which cytotoxic T lymphocytes (CTLs) kill virus-infected cells

is by inducing apoptosis http://www.sbs.utexas.edu/sanders/bio309/Lectures/2006/Lecture%2019%202006.htm

Cells of the immune system As cell-mediated immune responses wane, the CTLs must be removed to prevent them

from attacking body constituents. CTLs induce apoptosis in each other and even in themselves. Defects in the apoptotic machinery is associated with autoimmune diseases such as

rheumatoid arthritis.

Cells with DNA damage Damage to its genome can cause a cell

to disrupt proper embryonic development leading to birth defects to become cancerous.

Cancer cells Radiation and chemicals used in cancer therapy induce apoptosis in some types of

cancer cells.