EXPLORING EXPLORING BIOCHEMICAL BIOCHEMICAL ADAPTATION: ADAPTATION: Synthetic IntuitionSynthetic Intuition on a Family Farm on a Family Farm

www.carleton.ca/~kbstoreywww.carleton.ca/~kbstorey

Biochemical AdaptationBiochemical Adaptation- How Animals Work- How Animals Work

• Search to identify principles of biochemical regulation across the animal kingdom

• Biochemical unity• Chemistry bkgd., NO Biology (ever) • Elastic limits of biochemistry

There are so many songs There are so many songs and only the three cords. and only the three cords.

- John Hiatt - John Hiatt

A Life of Science A Life of Science Adventure Adventure

Mentors Mentors

Knut Schmidt-Neilsen, Ladd Prosser, Knut Schmidt-Neilsen, Ladd Prosser, Kjell Johansen, Kjell Johansen, Ken DaveyKen Davey, Hans Krebs,, Hans Krebs,

Gene Huber, Les Copley, Gene Huber, Les Copley, Geoff EalesGeoff Eales, A. Helix, A. Helix

Peter Peter HochachkHochachk

aa

DaveDave JonesJones

DO YOUR WORK, THEN STEP BACK.THE ONLY PATH TO SERENITY.

- “Tao Te Ching” by Lao Tzu

Storey Family FarmStorey Family Farm

SYNTHETIC INTUITIONSYNTHETIC INTUITION1) A new mix of ideas leads to a new field:

Chemistry

Mol. Biol.

Biochem.

NMR, MRI, DSC, AFM, DSF, UV difference, Protein purification, etc.

2) Models: Zoology, Ecology, Field Trips

3) Any new set of data reorganizes itselfthrough UNRELATED pieces of the puzzleand returns as UNIQUE.

4) Salvage Solutions from Chaos:

ex. Thesis Ideas, *DATA KNOTS*

5) Time Vampire: “Students Match Projects”

Comparative Experiments

Diverse Techniques

SYNTHETIC INTUITIONSYNTHETIC INTUITIONSomething Old,Something New,Something Borrowed, Some Glue

SYNTHETIC INTUITION INPUTINPUT

~ Ecology

~ Natural History

~ PHYSIOLOGY

~Chemistry Analytical

~ Biochem methods

~ Mol Biol methods

~ Genes – BioInform.

- NOT Metabolism

~ Equipment Expos

~ Catalogues

~ Tools:

- PHONE (1980)

- EMAIL (1990)

~ Jan Writing

~ Touring ( R&R)

OUTPUT OUTPUT

~ Metabolic Arrangement

~ Reorganization of Metabolism

~ Adaptive Change at Pathway Level

~ Integration: multi-levels of biol. organization

~ Model Organisms as raw material

~ Intracellular: from membrane to Nucleus

BIOCHEMICAL UNITY BIOCHEMICAL UNITY

FILTERFILTER

~ Transducer

~ Organizer

~ *Revamp*

~ IDEA LENS

~ Non-sci. Stability

~ Never “Circle the Wagons”

~ COST / BENEFIT

Why Am *I* Here “ “ My lab is full”My lab is full” >> Said to KBS when turned

down by PWH for a spot in his lab.

…15 minutes ( discussion) later …

“ “ Take that desk” Take that desk” >> Said to KBS when Peter

discovered that Ken had a scholarship that paid both salary and research expenses.

And, StudentsAnd, Students50 MSc, 36 PhD, 11 PDFs50 MSc, 36 PhD, 11 PDFs

MSc MSc K Male, D Miller, S Korycan, R Cole, A Chakrabarti, J Duncan, R WhitwamD Kelly, D Schafhauser, C Holden, E Russell, D Douglas, A Muise, T Pannunzio,P Schade, S Greenway, M de la Roche, T English, D Lobsinger, L Jurgensen, S Lautru T Pfister, S Saeedi, M Castellarin, E Sepehr, K Yan, J Ni, L Zhenhong, A Woods, S McFadyen, J Du, J Zhang, J Zhou, L Xie, M Bouffard, J Niles, L-H Yao, J Lama, R Bell, M Mathialagan, O Aguilar, N Dawson, C Brooks, R Roufayel, S Tessier, A Letourneau, M Allan, A Holden, K Sullivan, A Mahmud, J Wu

PDFPDFJ BallantyneS RahmanB MichaelidisR FergusonSPJ BrooksM Hermes-LimaC FrankQ CaiI MoineauM BatrukovaE Kotani

PhD PhD W PlaxtonT Churchill D Joanisse Y Su J Grundy H Mehrani C Holden W WillmoreB Thatcher K Cowan J MacDonald T Bilgen S-B Wu T EnglishA FahlmanD McNallyK Larade S Eddy

D Hittel D McMullen A de Croos P Morin K AbnousM HapsatouC RamnananC Dieni A MalikA KrivoruchkoB Lant K BiggarJ Zhang C-W WuN DawsonR Bell A LetourneauS Tessier

+ 105+ 105UndergradUndergrad

ss

WHY I PERSISTED WHY I PERSISTED • Synthetic Intuition Synthetic Intuition • JANET JANET • Talented Team (PhD, MSc, UG) Talented Team (PhD, MSc, UG) • Strategy (Student Wrangling) Strategy (Student Wrangling) • PDF ( Few but Key) + PDF ( Few but Key) + Luck Luck • Constant NSERC fundingConstant NSERC funding• Occasional Equipment grants Occasional Equipment grants • Big Scores (timely) CFI-OIT, NIH Big Scores (timely) CFI-OIT, NIH • Cardboard Enemies / Cardboard Enemies /

Big Talent Support Big Talent Support

My Presentation : My Presentation : R&R ? Or Comedy ? R&R ? Or Comedy ?

• At Rock Shows, you want the group At Rock Shows, you want the group to play the to play the OLDOLD songs just like on songs just like on the album the album

• At Comedy Shows, if you hear an At Comedy Shows, if you hear an OLDOLD joke, you boo. You want the joke, you boo. You want the NEWNEW, fresh material. , fresh material.

Answer: Science talks are about Answer: Science talks are about tomorrowtomorrow

Greatest Hits: (Vol 1)Greatest Hits: (Vol 1)“High Tides & Green Grass” “High Tides & Green Grass”

The early days The early days • Comparative regulation of glycolysisComparative regulation of glycolysis• Methods in Enzymology (purification) Methods in Enzymology (purification) • Exercise muscle metabolism: insects & cephalopods Exercise muscle metabolism: insects & cephalopods • Octopine (Octopine (Storey cycleStorey cycle), alanopine & strombine ), alanopine & strombine • Glycolytic Enzyme ComplexGlycolytic Enzyme Complex• Bound water in metabolism Bound water in metabolism • Reversible phosphorylation control of Reversible phosphorylation control of

metabolic enzymes ( Novel) metabolic enzymes ( Novel) • Phosphagen effects on enzymes Phosphagen effects on enzymes

• PFK polymers & F2,6PPFK polymers & F2,6P22

Greatest Hits: (Vol 2) Greatest Hits: (Vol 2) Anoxia ToleranceAnoxia Tolerance

• Anti-Pasteur Effect (Anti-Temperature !) Anti-Pasteur Effect (Anti-Temperature !) • Coordinated Metabolic Rate Depression (MRD)Coordinated Metabolic Rate Depression (MRD)• Anabolism Anabolism followsfollows Catabolism in MRD Catabolism in MRD • Reversible phosphorylation of glycolytic Reversible phosphorylation of glycolytic

enzymes: Gly phos, PFK, PK, etc. enzymes: Gly phos, PFK, PK, etc.• Protein kinases: PKA, PKG, MAPKs, AMPK, etc.Protein kinases: PKA, PKG, MAPKs, AMPK, etc.• Anoxia-induced gene expressionAnoxia-induced gene expression• Antioxidants – adaptation to variable oxygenAntioxidants – adaptation to variable oxygen

Greatest Hits: (Vol. 3) Greatest Hits: (Vol. 3) Cold & Freezing SurvivalCold & Freezing Survival

• Deep supercooling by cold-hardy insects Deep supercooling by cold-hardy insects • Freeze tolerance:Freeze tolerance:

- insects & molluscs - insects & molluscs

- frogs, turtles, lizards, snakes- frogs, turtles, lizards, snakes• Biochemistry of cryoprotectants Biochemistry of cryoprotectants • Adaptive evolution from underlyingAdaptive evolution from underlying

anoxia & dehydration tolerances anoxia & dehydration tolerances• Freeze responsive gene expressionFreeze responsive gene expression• Novel proteinsNovel proteins

Unique Animal Stress ModelUnique Animal Stress ModelVertebrate whole-

body freeze tolerance

Tissue cryopreservation

Tolerance of extreme ischemia and

hyperglycemia

Greatest Hits: (Vol. 4) Greatest Hits: (Vol. 4) HibernationHibernation

• Regulated MRD drives body temperature downRegulated MRD drives body temperature down• Reversible enzyme control in torpor-arousal cyclesReversible enzyme control in torpor-arousal cycles• Cold adaptation of mammalian enzymesCold adaptation of mammalian enzymes• Polysome dissociation, mRNA storagePolysome dissociation, mRNA storage• Micro RNA Micro RNA • Gene screening, epigenetic controlsGene screening, epigenetic controls• Antioxidant defense (Tf) Antioxidant defense (Tf) • Atrophy resistance Atrophy resistance

Greatest Hits: (Vol. 5) Greatest Hits: (Vol. 5) EstivationEstivation

• MRD parallelsMRD parallels estivation = low oxygen / anoxia estivation = low oxygen / anoxia

• Phosphorylation also controlsPhosphorylation also controls - ion motive ATPases (NaK, Ca) - ion motive ATPases (NaK, Ca) - novel enzymes - novel enzymes - protein synthesis flux generators - protein synthesis flux generators - transcription factors - transcription factors

• Involvement of protein kinases – Akt,Involvement of protein kinases – Akt, AMPK, MAPKs, PKA/PKG AMPK, MAPKs, PKA/PKG

• Role of FOXO transcription factorsRole of FOXO transcription factors• Urea effects on enzymesUrea effects on enzymes

www.carleton.ca/kbstoreywww.carleton.ca/kbstorey

• Animal stories• Publications• Research Interests• Students & projects• Media•Textbook with former students

Greatest Greatest

Hits: Hits: Full Full CollectionCollection

Best Experiment EVER ! Best Experiment EVER !

CSZ U. Ottawa

1983

J.E.StoreyCarleton U.

Estivation

METABOLIC RATE METABOLIC RATE DEPRESSIONDEPRESSION

Diapause

Freezing

Anoxia

Hibernation

Crayfish

Principles of Principles of Metabolic Rate DepressionMetabolic Rate Depression

• Transcriptional + translational + PTM Transcriptional + translational + PTM • Anabolism Anabolism followsfollows Catabolism in MR Catabolism in MR • Coordinated suppression: gene expression, Coordinated suppression: gene expression,

protein synthesis, biosynthesis, cell cycle, etc. protein synthesis, biosynthesis, cell cycle, etc.• Reversible phosphorylation of key enyzmes,Reversible phosphorylation of key enyzmes,

ion channels, transcription factors, etc. ion channels, transcription factors, etc.• Stress-activated protein kinases Stress-activated protein kinases

Cell preservation in hypometabolismCell preservation in hypometabolism• Antioxidant defenses, iron sequesteredAntioxidant defenses, iron sequestered• Chaperone proteins Chaperone proteins

GENESGENES

Transcription

RNAsRNAs

Control byControl bytranscriptional regulationtranscriptional regulation

Control byControl bytranslational regulationtranslational regulationTranslation

PROTEINSPROTEINS(ENZYMES)(ENZYMES)

Control byControl byproteasesproteases INACTIVEINACTIVE

ENZYMEENZYME

NoModification

FUNCTIONALFUNCTIONALENZYMESENZYMES

Covalentmodification

Degradation

Control by post-Control by post-translationaltranslationalmodificationmodification

ACTIVEACTIVEENZYMESENZYMES

Inhibitionand

Activation

Control at level ofControl at level ofenzyme functionenzyme function

www.carleton.ca/~kbstoreywww.carleton.ca/~kbstorey

GENE CHIPS TRANSCRIPTION FACTORPROFILING

Data LeadsData Leads

ELISAs in plates

Confirm by EMSA

Confirm byRT-PCR,

Northern blots

TfDownstream genes

ROLE & CONTROL OF SYSTEM

Transgenics

Cell AssayRNAi Knock out Epigenetics

FUNCTIONAL ASSAYS

Protein levels - enzyme assay - antibodies : protein - functional analysis e.g. HIF EPO

Where do we go from Where do we go from here? here?

• Human applications of MRD research • Novel phosphorylations• Atrophy, hypertrophy -- autophagy for survival • Turning it all off -- microRNA• Epigenetics & adaptation • Life span extension• Antioxidant defense• Cell cycle suppression• Unity through evolution

NOW: finally !!! NEW NOW: finally !!! NEW DIRECTIONS DIRECTIONS

Hibernation and medicineHibernation and medicine

Primates !!

Novel phosphorylations Novel phosphorylations

PPPP PP

PP

PiPiProtein

Phosphatase

ATPATP ADPADPProtein Kinase

Atrophy – HypertrophyAtrophy – Hypertrophy

Turning it all Turning it all offoff

Epigenetics in AdaptationEpigenetics in Adaptation

Life span Life span extensionextension

Ukrainian connectionUkrainian connection- Metal & herbicideMetal & herbicide

effects on fisheffects on fish- Antioxidant defense- Antioxidant defense

Unavoidable metabolic costs Unavoidable metabolic costs

Unity through Evolution Unity through Evolution

WWCeDWWCeD

ONWARDS !ONWARDS !

pH low high

MW

low

hig

h