Transcript Analyzing the Increased and Decreased Expression

Analyzing the Increased and Decreased
Expression of Microarray Data for Vibrio
cholerae
J’aime Moehlman
Amanda Wavrin
April 27th, 2010
BIOL 398/S10: Bioinformatics Lab
Outline
• Vibrio cholerae is a waterborne disease that is
infectious to humans
• Statistical analysis of the microarray data
• Microarray data from the Merrell et al. (2002) study
• Increased expression
in microarray
• Decreased expression
in microarray
Vibrio cholerae is a waterborne disease
that is infectious to humans
• Organisms in the genus Vibrio are one of the most
common surface water organisms in the world.
• They can be found in both fresh and marine waters and
live in association with other animals.
• V. cholerae is noninvasive and affects the small
intestine.
• It produces a cholera toxin, which is responsible for the
characteristic diarrhea.
• This leads to dehydration, anuria, acidosis, shock,
cardiac complications, and circulatory failure.
Microarray Analysis in the Merrel et al.
study
• The study was conducted in Dhaka, Bangladesh
• Samples were collected from three patients.
• The collected strain, O1 Inaba El Tor, was combined
with an in vitro strain and used to inoculate mice.
• The output ratios were corrected to represent the
competitive indices (CI) of the V. cholerae.
• A CI above 1 indicates increased infectivity.
• A CI below 1 indicates decreased infectivity.
The human-shed V. cholerae had a CI
above 1 indicating an enhanced infectivity.
• V. cholerae that was cultured and purified in vitro did
not show enhanced infectivity
• RNA from each sample was used for DNA synthesis
• This was labeled with Cy5 and hybridized to the
microarray with a Cy3- labeled common reference
strain (exponential growing).
• A two-fold difference indicated differentially
expressed genes
GenMAPP was used to download the
correct V. cholerae gene database
• There were 772 errors detected when converting the raw
data.
• We customized our data by selecting new color sets.
– Pink showed increased expression
– Blue showed decreased expression
• Criterion 0 represented increased.
• Criterion 1 represented decreased.
• Statistical Criteria:
–
–
–
–
Z Score > 2
P value < 0.05
Number Changed Ranged from <3-5
% Changed Ranged < 15-25
Increased Gene Expression for Infectivity
Increased Metabolic Processes and Organization
resulted in more virulent strain
• Metabolic Processes
– Amino acid
– Cellular
– Nitrogen
• Amino Acid Processes
Cellular Metabolic Process
GO Term
– Are relative to cell metabolism
• Movement via Flagellum
– Process carried out at cellular level which controls
assembly, arrangement, or disassembly of flagellum
Decreased Gene Expression for Infectivity
Decreased in Overall Organization
• Nucleotide binding
• Protein functions
– Folding
– Binding
• Molecular binding
– Zinc
– Cation
– Vitamin
Protein Folding GO Term
Increases in metabolic, amino acid
processes and flagellum organization show
overall virulence in V. cholerae
• Increased metabolic processes play a role in energy
distribution within the bacteria.
• Increase amino acid metabolic processes present
increase in storage molecules.
• Increased movement via flagellum plays a role in
increased overall virulence of the bacterial cells.
• Amino acid processes effect protein structure, which
has an overall effect on protein function.
• These increases focus on the pathogenicity of V.
cholerae and the structural component of the cells.
Decreased overall organization of the cell
results in increased pathogenicity
• Decreased protein binding will effect the protein
structures which has an overall effect on the protein
function.
• Utilizes the increased storage molecules to decrease
protein synthesis.
• Decreases molecular binding in order to focus on
increasing its’ virulence.
Significantly changed genes in comparison
to Merrell et al. (2002)
• According to the Merrell et al. study, these genes were
significantly changed
– VC0028, VC0941, VC0869, VC0051, VC0647, VC0468, VC2350,
VCA0583
• We found that only 2 of these 8 genes showed to be
significantly changed in our results.
• These genes were VC0647 and VCA0583
– VC0647: mRNA catabolic processes, RNA processing, cytoplasm,
RNA binding, 3'5'exoribonuclease activity, transferase activity,
nucleotidyltransferase polyribonucleotide
nucleotidyltransferase activity
– VCA0583: transport, cell envelope outermembrane bounded
periplasmic space, transport activity
References
• Merrell DS, Butler SM, Qadri F, Dolganov NA, Alam A,
Cohen MB, Calderwood SB, Schoolnik GK, and Camilli
A. Host-induced epidemic spread of the cholera
bacterium. Nature 2002 Jun 6; 417(6889) 642-5.
• Todar, Kenneth. Online Textbook of Bacteriology “Vibrio
cholerae” http://www.textbookof
bacteriology.net/cholera.html. 25 April 2010.
• http://www.zdravstvena.info/vsznj/wpcontent/uploads/2009/01/kolera.jpg