Question

1. Describe in detail how both positive and negative regulation control expression of the lac operon in bacteria. In doing this, also convey understanding of how and why expression of this operon is controlled two ways (positive and negative). Also answer, what are the energetic benefits to the cell of operon regulation? 2. What is horizontal gene transfer? Describe in detail the three mechanisms of horizontal gene transfer. What are the evolutionary advantages and costs of horizontal gene transfer. I encourage you to frame your answer with respect to the paper by Dantas and Sommer. Reference them appropriately.

Answer 1

1. Positive regulation of lac operon

In case of lac operon, the presence of lactose causes positive regulation of the operon. During this condition, the repressor protein cannot bind to the operator region. This is due to the present of allolactose. Allolactose is a form of lactose that is made by the help of the beta-galactosidase enzyme that the cell already has a bit before the beginning of the lac operon. This beta galactosidase enzyme is coded by lacZ gene and this breaks up the lactose to form allolactose. This allolactose binds to the repressor protein that is synthesized by lacI and removes the repressor protein from the operator region. In presence of lactose, cAMP levels are high and so the CAP levels are activated. CAP binding protein helps in the binding of the RNA polymerase to the promoter region and proceed in transcription. With the removal of the repressor protein, the RNA polymerase can now start the transcription of the lacZ, lacY and lacA genes. The lacZ increases production of beta-galactosidase hence more of allolactose is being produced, lacY encodes for the permease that allows the cell membrane to let more of lactose enter the cell and keep the lac operon on and the lacA gene encodes for the transacetylase that attaches chemical groups to the lactose molecules to direct to the operon.

Negative regulation of lac operon:

This occurs when the cell grows in a medium that has no or very less amount of lactose but more of glucose. In presence of glucose the cell does not need to transcribe the lac operon genes at a higher rate and so the cAMP levels are very low. The CAP is inactive and cannot bind DNA so RNA polymerase has no guide to direct it for transcription. No allolactose is produced hence the constitutively expressed repressor protein produced by the lacI gene remains bound to the operator regio, inhibiting the RNA polymerase to transcribe any of the genes.

2. Horizontal gene transfer:-

Horizontal gene transfer is the phenomenon of transfer of genetic materials between unicellular organisms. There are three different methods of gene transfer-

• Transformation

Some bacteria can take up fragments of DNA from the external medium.Source of the DNA can be other cells of the same species or other species.In some cases the DNA has been released from dead cells.In other cases the DNA has been secreted from live bacterial cells.If this DNA is of a different genotype from the recipient, the genotype of the recipient can become permanently changed, a process aptly termed transformation.In gram negative bacteria, the basic steps of transformation are:

Binding of double stranded DNA to the outer cell surface of the bacterium,Movement of the double stranded DNA across the outer membrane and cell wall, Degradation of one of the DNA strands, Translocation of the remaining single strand of DNA into the cytoplasm of the cell, the incoming donor allele replaces the allele of the recipient; that is, DNA substitution occurs.The DNA fragment containing the donor allele is added to the genome of the recipient, by insertion adjacent to the recipient allele, thereby forming a gene duplication (partial diploid)

• Conjugation

This represents transfer of DNA from one cell to another by direct cell-to-cell contact. The transfer occurs from a donor cell to a recombinant recipient.The first step is contact between the cells via the conjugation tube that is formed with the help of sex pilus between the two cells. After the formation of the conjugation tube, the DNA from the donor cell is transferred to the recipient cells along with the replication machinery containing the oriT region. The DNA undergoes full replication in the recipient cell and after the completion, the conjugation tube breaks and the transfer is complete.

• Transduction

In this process , the bacterial DNA is transferred from one cell to another by the help of a virus or bacteriophage. The donor cell replicates it's own DNA along with the bacteriophage DNA and assembles the phage with the packaged DNA of it's own plus the phage DNA. This phage now infects the recipient cell and released the DNA content within the cell that has both the phage DNA as well as the DNA from the recipient cell.

The evolutionary advantages of the horizontal gene transfer are:

1. It helps in denoting the last common ancestor of the organism in question. The transfer of the same gene, say for example an antibiotic resistant gene, will help to trace back the origin of the gene from the early member to the newest organism.
2. It helps in phylogenetic analysis
3. The adaptation of the bacteria can be learnt by the help of the study of the gene transfer. The idea of the bacterial chromosomal organization helps to study the origin of a particular gene that is being transferred from one cell to another.