differential gene expression in heterosigma akashiwo in response to model flue gas: where does the...

Differential Gene Expression in Heterosigma akashiwo in Response to

Model Flue Gas:Where Does the Carbon Go?

8th Annual Algae Biomass SummitAlgal Strain Development

October 2, 2014

Jennifer J. Stewart, Ph.D.Scientist, NSF SEES Fellow

302-645-4371Jen@udel.edu

R83-3221

Koonin et al. (2010) Genome Biology 11:209

What is a Raphidophyte?

Hara & Chihara 1987

3

Heterosigma akashiwo Raphidophyte Metabolizes NO gas Optimum growth

maintained over a wide range of salinity (10-30 psu) and temperature (16-30°C)

Survives nutrient limitation and high light stress

Exhibits no strong preference for nitrogen source

(NO3- , NO2

-, NH4+)Photos by Elif Demir-Hilton and Kirk Czymmek

Jennifer J. Stewart, Ph.D.Scientist, NSF SEES Fellow

302-645-4371Jen@udel.edu

Theoretical Mechanism of NR2-2/2HbN

Dual NO dioxygenase and Nitrate Reductase Activities

5 e- when fully reduced: 2 e- accepted by FAD, 1e- by heme-Fe, and 2e- by Mo-MPT

Univalent reduction of both heme-Fe centers possible

Nitrate captured(Stewart & Coyne 2011)

Jennifer J. Stewart, Ph.D.Scientist, NSF SEES Fellow

302-645-4371Jen@udel.edu

5

Air 10.5mM Flue Gas 12.4 mM

12% CO2, 150 ppm NO, N2 balance

Heterosigma akashiwo on a Model Flue Gas

Jennifer J. Stewart, Ph.D.Scientist, NSF SEES Fellow

302-645-4371Jen@udel.edu

_x0003_Air _x0008_Flue Gas0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Carbohydrate LipidProtein

Carb Lipid Protein0

1

2

3

4

5

6

7

8

9 Air Flue Gas

Pro

du

cti

vit

y (

gra

ms

m-2

da

y-1

)

Compositional Breakdown-Per Cell Basis

6

1 3 5 7 9 11 13 15 17

-10

-8

-6

-4

-2

0

2

4

6

8

10

-log(p-value)

log

2 (

fold

ch

an

ge

)

1641 Transcripts Were Significantly Differentially Expressed

524 Received KEGG Orthology (KO) Identifiers

274 Received Enzyme Commission (EC) Numbers

149 Mapped to KEGG Reference Pathways

Global Gene Expression Analysis

7

Metabolic Overview of Differentially Expressed Genes

8

Photosynthesis

Carbohydrate

Carbohydrate

TCA

Lipid Syn

N-Metabolism

AA

Isoprenoids

Abiotic Stress

Redox

Nucleotides

RNA Processing

DNA Synthesis

Protein SynthesisSignaling

Cell Cycle

Transporters

Unknowns

Metabolic Overview of Differentially Expressed Genes

9

For NR, Only NR2 sequences were Found to be Differentially Expressed The Highest Up-regulated Transporter Genes were Phosphate Transporters Flue Gas is a Significant Source of Nitrogen for Protein Synthesis

Nitrogen Uptake and Utilization

10

C14:0

C15:0

C15:1

C16:0

C16:1

C16:1

C17:0

C17:1

C18:0

C18:1

C18:2n

6

C18:3n

3C18

:4

C20:1n

9

C20:5n

3

C22:1n

9C22

:2

C22:6n

30

2

4

6

8

10

12

14

16

18Air Flue Gas

FA

ME

(p

g/ce

ll)Fatty Acid Biosynthesis

Increase in FAs involved in plastid membrane composition

11

Fatty Acid Biosynthesis

12

Carbonate Chemistry

13

CO2 Fixation During Growth on Model Flue Gas

a-D-Glucose-6P

b-D-Glucose-6P b-D-Fructose-6P

b-D-Fructose-1,6P2

Glyceraldehyde-3PGlyceraone-P

Glycerate-1,3P2

Glycerate-3P

Glycerate-2P

Phosphoenolpyruvate Pyruvate

Carbon Fixation

AA Synthesis Fatty Acid Synthesis

Up-Regulation Seen Throughout the

Glycolysis Pathway

_x0003_Air _x0008_Flue Gas0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Carbohydrate LipidProtein

Heterosig

ma aka

shiw

o

Nannoch

loropsis

sp.

0%

20%

40%

60%

80%

100%

Carbohydrate LipidProtein

Pathway for Storage Carbohydrates?12%CO2 + 150 ppm NO 2%CO2 – Day 5 Batch Growth

Pathway for Storage Carbohydrates?

16

Green algae = StarchDiatoms = Chrysolaminarin

Jennifer J. Stewart, Ph.D.Scientist, NSF SEES Fellow

302-645-4371Jen@udel.edu

17

Glycolytic and Glucan Biosynthesis in the Diatom Phaeodactylum tricornutum

Chauton et al. (2013) Plant Physiology 161:1034

18

All Roads Lead to b-D-Fructose-6P

19

Storage Carbohydrate metabolism of Ectocarpus siliculosus. Michel et al. (2010) New Phytologist 188:67

Dittami et al. (2011) Plant Signaling & Behavior 6:8

Fate of b-D-Fructose-6P in Brown Macro-Algae

Acknowledgements

20

R83-3221 R83-3221

• UD: Kathy Coyne, Mark Warner, & Tom Hanson • SU: Katherine Miller• ASU: John McGowen, Tom Dempster, Hank Gerken, and

Crew• Lab Members: Colleen Bianco, Chris Main, Kaytee P.,

Josee Nina Bouchard• DNREC Division of Air Quality: Ali Mirzakhalili & Mark

Lutrzykowski• The following funding sources:

Jennifer J. Stewart, Ph.D.Scientist, NSF SEES Fellow

302-645-4371Jen@udel.edu