1. Impact of Amino Acid Depletion
on Streptococcus gordonii Gene
Expression
Jill Robinson, Centre for Oral Health Research,
Newcastle University (UK)
IADR, Boston, 13th March 2015

2. Background to project
• Streptococcus gordonii
– Early coloniser
– Important in binding to other cells
– Anchors biofilm foundations
Image credit: R. Holliday
• Sensing arginine linked to important phenotypes,
e.g.:
– Arginine biosynthesis in response to coaggregation
– ∆arcR mutant can’t regulate arginine catabolism, has
biofilm defect

3. Aims of project
• Does arginine depletion affect gene expression?
– Analyse samples by microarray and qRT-PCR
• Is this response specific to arginine?
– Compare responses to arginine depletion with
histidine/branched-chain amino acid depletion

4. Microarray sample preparation
Culture in
high
arginine to
mid-
exponential
phase.
Split
culture
and
harvest
Resuspend
High arginine
No arginine
37°C, 30 min
Extract RNA

5. Microarray findings
• > 400 genes regulated in response to arginine
– Approx. 20% of all predicted genes
• Includes genes involved in:
– Arginine biosynthesis (>200-fold up-regulated)
– Histidine biosynthesis
– bfb locus
– Receptor polysaccharide biosynthesis
– Hsa biosynthesis
Involved in
binding,
adhesion and
biofilm formation

6. Microarray validation using qRT-PCR
r2 = 0.947

7. Effect of amino acid deprivation on gene
expression
• Further qRT-PCR analysis on samples depleted of
other amino acids (arginine, histidine, branched-
chain amino acids)
• To check which differences in gene expression were
arginine-specific
• Cell growth in deplete media showed same growth
pattern as for arginine

8. Arrow shows point
of subculture to
depleted media
FMC growth curve graph

9. Heat map of
differential
gene
regulation in
response to
amino acid
deprivation:

10. Arginine-specific responses
Gene name Function/Locus function
Regulation in response
to arginine depletion
(fold-change values)
asp5 hsa locus, binding to oral cavity Down-regulated (-21.6)
hisC Histidine biosynthesis Up-regulated (19.3)
argC Arginine biosynthesis Up-regulated (33.9)
bfbC Bfb locus, necessary for S.
gordonii biofilm
Down-regulated (-19.6)
bfbF Down-regulated (-9.6)
SGO_1686 Fatty acid biosynthesis Down-regulated (-35.3)
wefE
Polysaccharide transport
protein
Down-regulated (-17.8)

11. Conclusions
• Arginine affects genes involved in binding,
adhesion and biofilm formation
• Some responses are specific to arginine,
although some are general response
• Arginine concentration and metabolism –
possible use as a signal for biofilm formation?
• Future potential target for biofilm control?

12. Thank you for listening!
Sponsored by:
No conflicts of interest are present.
With thanks to:
Dr Nick Jakubovics
Dr Alex Rickard
Prof Waldemar
Vollmer
Dr Matthew German
Mrs Lesley Old
Everyone in the OBL