Quorum Sensing and Biofilms

Bacteria constantly release signal molecules If cell density is not big enough nothing

happens Threshold density all bacteria

simultaneously carry out gene expression “Quorum Sensing” from counting as to take

a vote before acting

Quorum Sensing Basics

Acyl homoserine lactones (AHL molecules) Easy transport of molecules outside of cells

Gram Negative Bacteria

Precursor Oligopeptides Must be exported via active transport

proteins

Gram Positive Bacteria

Al-2 molecule is universal Produced by LuxS enzyme from dihydroxy

pentanedione substrate

Inter Species Communication

Microorganisms form layer with proteins, DNA and long chain sugars

Often pathogenic Concern for health and food industry

◦ Spoiled food◦ Biofilms in cystis fibrosis

Quorum sensing plays big role in the high cell density communication and virulence

Biofilms

Scientists are investigating varied bio-film formations and mutations that could help decrease formation

Looking at mutant luxS enzymes and other quorum sensing systems

Disrupt quorum sensingdisrupt biofilm formation and virulence

Biofilms Today

Ep-CAM Gene Silencing in

Craniopharyngiomas

Benign epithelial brain tumor common in children

Cannot spread but presses on vital structures

Can cause stunted growth, obesity, diabetes insipidus, and other morbidity

Hard to remove due to strong adherence to important structures

Often reoccurs

Background

Ep-CAM gene expression is highly over expressed

Produces Ep-CAM glycoprotein adhesion molecule

Is linked to tumor genesis and metastasis Makes tumor strongly adherent

Ep-CAM Glycoprotein

Craniopharyngioma

Radiotherapy◦ Kills healthy cells◦ High risk of second cancer in children and adolescents

Competing Technologies

Silencing Ep-CAM gene by inhibiting transcription through methylation of the promoter region

Design

Inducing Bacteria

Aerotolerant E.Coli bacteria engineered to produce invasin protein and produce m5c DNA methyltransferase enzymes on signal

Calcification material

Ep-CAM glycoproteinReceptor

Receptor

Invasin and m5c Production

m5c methyltransferase enzymes

Invasin proteins

Bacteria Produces

B-1 Integrin receptor

Invading Tumor Cells

Tumor cell

Invasin Proteins

Inside Cell

Ep-CAM promoter

M5c DNA methyltrasnferase enzymes

Inhibition of RNA Polymerase Binding

RNA Polymerase

Ep-CAM Transcription

Ep-CAM glycoprotein

Calcium deposits m5c methyltransferase production

0 0 00 1 01 0 01 1 1

AND Gate Truth Table

Methylated CpG Islands on Promoter

Ep-CAM Expression

0 11 0

Inverter Logic Gate Truth Table

No damage to healthy cells Easy to administer No radioactive material Helps surgeons remove tumor Decrease reoccurrence

Advantages

Ep-CAM promoter region specific methylation

Laboratory precaution while working with cells with invasin

Precautions to keep bacteria from evolving and reproducing out of control

Possible Problems

Testing on tissue culture and many other steps before reaching human testing

Similar system could be tweaked and developed for other cancers

Testing