Good morning everyone!

This baby girl was born happy and innocent. She just wantedto play and have fun all the time. But, destiny has decided

otherwise.

40 years from now she started feeling different. Something is going wrong in her body. She decided to go see a doctor.

After extensive tests, the doctors found abnormal cells in her body. They looked liked this

She is diagnosed with BREAST CANCER!

What is cancer? How is it caused?These are some of the questions that plague us.

A normal cell looks like this

Three important features ofa normal cell:

1. It has a regular shape and size2. It has a normal DNA3. It divides normally

When a cell undergoes mutation, the above mentioned features are altered, leading to a cancerous cell.

A crash course in cancer!

Cancerous cells divide abnormally causing tumors.

After a mutation, the sequence of base-pairs in DNA changes.

Now u know what is cancer!!

Coming to breast cancer……What is that thing? How is it caused?

Normal female breasts are made up of1) Lobules – milk-producing glands 2) Ducts – milk transporting tubes3) Stroma – fatty tissue surrounding the ducts and lobules4) Blood vessels and Lymph vessels

Breast cancer is a malignant tumor thathas developed from cells of breast.

Some facts about the breast cancer:•Breast cancer is the most common cancer among women.•It also occurs in men.•It is estimated that in 2003 about 211,300 new cases will be diagnosed among women. •It is hereditary!

Doctors cannot explain why one gets cancer and why one doesn’t.They can help you by diagnosing the cancer at its early stages and help prevent its development.

But we are not doctors. We are BIO-INFORMATICISTS. We have the opportunity to understand the disease and informmany others, including the doctors about it.

Nobody has so far discovered the true reason behind the cause of this disease. But, we know that its hereditary.

We also know the risk factors involved in the disease. Some of them are:1. Prolonged exposures to estrogen, female hormone.2. Late childbearing (having first child after about age 30)3. Breasts that have a high proportion of lobular and ductal tissue density4. Exposure to radiation 5. High alcohol consumption

Getting bored????

Genetics of breast cancer

So…no cure?? What next?? Are pain and suffering the only answers??

In 1994 scientists have discovered two tumor suppressor genes:1. BRCA12. BRCA2

Any mutations in the above genes causes breast cancer. After 1994 many othergenes were discovered that were linked to the cause of breast cancer.

But these genes don’t directly involve in tumor formation.

BRCA1 gene identified on the long arm of chromosome 17BRCA2 gene identified on the short arm of chromosome 13

45% of familial cancers and more than 80%of ovarian cancers are alsocaused due to mutations in BRCA1.

A quick fact

The function of these genes was not clear until studies on a related protein in yeast revealed their normal role: they participate in repairing radiation-inducedbreaks in double-stranded DNA.

This means that mutations might disable this mechanism leading to more errors in DNA replication.

These defective genes are transferable from parents to offspring.

I have concentrated on BRCA1 to make it easy for myselfas well as for you all.

From our very own NCBI web site, I have secured a lotof info on BRCA1

A brief explanation

NCBI is the mother of all genomic web sites. Every new nucleotideor protein or genomic material will be readily available in the database.

Search over 18 vast databases for related sequences.

Translate sequences

Know lot of other stuff

The latest discovery is the SYNTHETIC LETHAL SCREENING

When cell death is caused by the combination of two or more mutations,the genes involved are said to be synthetically lethal.

This indicates a functional relationship between the corresponding gene products.

So what’s my objective??

I chose to work on BRCA1 gene

I’ve decided to find relative genes in other organisms that belong to different classes.

I want to learn the evolution of this genein different organisms. May be this infomight help me better understand the gene, and its function.

NCBI was helpful in finding the related genes in other organisms.

Related sequences??

BRCA1 is a gene, and most genes make their products. Gene products aremostly proteins.

In related sequences the gene product is almost the same.

How will this help us?

We will try to compare the gene products in all the selected organisms.This will lead us to a point where we can understand the evolution of thespecific gene we are looking for.

Isn’t that simple?? At least sounds like it.

Chosen sequences

I chose sequences from the following organisms (organisms listed withscientific name, common name and class they belong).

Homo sapiens Man MammaliaPan troglodytes Chimp MammaliaGallus gallus Hen AvesXenopus laevis Frog AmphibiaDrosophila Fruit fly InsectaCaenorhabditis Nematode NematodaPlasmodium Sickle parasite ProkaryoteArabidopsis Thale cress Plant

Scientific name Common name Class/Kingdom

Did you find anything strange in the list of the sequences?

Yes…how come I included Plasmodium (prokaryote), Arabidopsis(plant), nematode, and an insect?

Scientists have reported related proteins in these organisms.Isn’t it surprising??

Lets go ahead and analyze these sequences using the programs that Dr.Joshi taught us. Its fun, and very interesting.

I am using only those programs that are required for my analysis.

NOTE:

Important analyses

%Similarity and identity:

Using doublegap.csh script I identified percentage similarity and identity.

ResultsHuman and Chimp proteins showed 99% similarity.Arabidopsis, Drosophila, Nematode are no way similar to Human and Chimp.Frog and Hen come closer in terms of similarity to Human and Chimp.

Multiple sequence alignment:

Using pileup function I aligned all the sequences.I encountered a problem.Plasmodium had many amino acid sequences. This led to the incorporation ofGaps beyond the limit (actually we can set the limit).

So I ignored Plasmodium sequence.

I used the following script to run the pileup program:

“pileup -MAXSeg=2000 –MAXGAP=5000 –OUTfile1=listgen.msf @listgen”

This script will allow me to extend the gap sequences to 5000 and maximumSegment alignment to 2000. Observe that the sum is 7000. It should always be 7000 (Well I don’t know why? Its in the program.)

After aligning the protein sequences, I used PRETTY to find the consensus of the aligned sequences.

Do u wanna take a look at the pretty file?

Br1: DrosophilaBr2: Chimp

Br3: HenBr4: Frog

Br6: HumansBr7: Arabidopsis

Br8: Nematode

Consensus sequences

Some of the stuff is confusing. Some of it makes sense.

To better understand it, lets go ahead and get a phylogenetic picture.

Phylogenetic tree:

I went to clustalw web site to get a beautiful phylogenetic treeby feeding the pileup file.

The Result???

An amazing phylogenetic tree that supports the theory of evolution.

Phylogenetic tree

Analysis of the phylogenetic tree

According to the theory of evolution, at the bottom ofthe phylogenetic tree are the prokaryotes.

Prokaryotes are followed by nematodes.

Nematodes are followed by insects, amphibians, aves and mammals.

So we are at the top of the phylogenetic tree.Humans and Chimps are mammals. So there is a close relation between theproteins.

It looks great!

But how does this help??

HOPE!!

These techniques will help scientists work on a differentliving system to understand the functioning of the gene.

Yeast (we did not discuss about it) is one of the living systems that is extensively used for research.

May be in the future, we will find out the actual causeof this dreadful disease, and find a cure for it.

We should never lose hope. It is the only thing that will inspire us to reach our goals!

THANKS!