Secrets of the lac operon: A unified mechanism for aging, diabetes, and obesity

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Secrets of the lac operonA unified mechanism
for aging, diabetes, and obesity
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Lac operon Substrates induce their own
metabolism, and exhibit hysteresis
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The Metabolic Mystery Aging and Disease

• Obesity increases with age
• Obesity pre-disposes to age-related diseases
  (metabolic syndrome)
• Caloric restriction extends lifespan

WHY??
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WHY??

• Does obesity increase with age?
• Does protein oxidation increase with age?
• Does glycolysis increase with age?
• Does protein turnover decrease with age?
• Are effects of aging progressive and apparently
  irreversible?
• Are effects of diabetes progressive and
  apparently irreversible?
• Are effects of aging and diabetes really
  irreversible?

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Caloric (glucose?) restriction
Nutrient sensors (transcription factors?)
And then a miracle occurs
Increased lifespan
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How do (glucose) calories kill?Focus on brain
gene expression

• Diabetic complications mainly in
  insulin-insensitive tissues
• Neurons implicated in many models of aging
• Neurons regulate metabolism

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Dietary restriction alters characteristics of
glucose fuel use.Masoro, EJ, McCarter, RJ, Katz,
MS, and McMahan, C AJ. Gerontology (1992)
47(6)B202-B208
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Molecular hysteresis residual effects of
hormones and glucose on genes during aging.C.V.
MobbsNeurobiology of Aging (1994) 15523-534
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Glucose hysteresis
Glucose resistance
SIP down
VMH neurons glucose resistant
Inducedgenes
Inhibitedgenes
YoungThin
Old Fat
Many meals(glucoseexcursions)
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But is it glucose??

• Reducing glucose increases lifespan in yeast
• DR (including methionine) reduces glucose
• Every-other day feeding does not reduce calories,
  but reduces glucose and extends lifespan
• Elevated glucose causes toxicity (diabetic
  complications)

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What does glucose do?
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BUT- Low glucose blocks glycolysis

• Phosphofructokinase DOWN
• Transaldolase DOWN
• Ketohexokinase DOWN
• Moncarboxylate transporter 1 Down
• Pyruvate dehydrogenase kinase 4 UP

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AND- low glucose blocks NADH glycerol shuttle

• Mitochondrial glycerolphosphate dehydrogenase
  DOWN
• Cytoplasmic glycerolphosphate dehydrogenase
  UP

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Low glucose stimulates pentose pathway (makes
NADPH)

• Glucose-6-phosphate dehydrogenase UP
• Linked to aging and stress resistance

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Low glucose stimulates TCA cycle and
mitochondrial NADPH

• Isocitrate dehydrogenase 2 UP
• Linked to aging and stress resistance

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Low glucose activates hypothalamic mitochondrial
lipid oxidation

• Carnitine palmitoyl transferase UP
• Fatty acid transport protein UP
• Mitochondrial Acyl-CoA thioesterase UP

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AND- Low glucose activates peroxisomal fatty acid
oxidation

• Peroxisome membrane protein UP
• Peroxysomal Acyl-CoA peroxidase Up
• Peroxisomal integral membrane protein UP
• Peroxisome membrane protein UP
• Peroxisomal integral membrane protein UP
• Peroxisomal biogenesis factor UP

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AND- Low glucose activates protein and amino acid
degradation

• Multi-ubiquitin chain binding protein UP
• Ubiquitin fusion degradation protein 1 Up
• Proteasome delta subunit UP
• Proteasome (prosome) subunit UP
• Proteasome alpha subunit UP

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AND- Low glucose activates protein synthesis

• Ribosomal protein S7 UP
• Translation initiation factor eIF2 gamma Up
• Ribosomal protein S5 UP

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Low glucose induces the highly anti-oxidant
glucose switch profile of gene expression

• Glycolysis down (blocks NADH complex I)
• Complex I produces free radicals
• Pentose pathway up
• Makes antioxidant NADPH
• Lipid oxidation up (favors FADH2 complex II)
• Complex II does not produce free radicals
• Amino acid oxidation up
• Eliminates oxidatively damaged proteins
• Favors NAD
• Induces protective effects (e.g., SIR1)

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The glucose switch ATP-neutral substrate
repartitioning
Glucose Transport
GLUT1
Pentose Shunt
Cytoplasm
Glycolysis
PFK
Glucose-6-Phosphate DH
NADPH
Fatty Acid Oxidation
Glucose to Krebs
PDHK4
CPT1 PPaR?
NAD
Krebs Cycle
NADH
Complex I
Complex II
FADH2
Isocitrate DH2
NADPH
Mitochondria/Peroxisome
Respiratory Chain Shift to Complex II
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Conversely, high glucose blocks the glucose
switch profile, leading to an oxidative profile

• Glycolysis up (favors NADH complex I)
• Complex I produces free radicals
• Reduced Complex I, III, IV, and V increase
  lifespan
• Pentose pathway down
• Reducing antioxidant NADPH
• Lipid oxidation down (inhibits FADH2 complex II)
• Complex II does not produce free radicals
• Impaired Complex II reduces lifespan
• Amino acid oxidation down
• Allows accumulation of oxidatively damaged
  proteins
• Favors NAD
• Inhibits protective mechanisms (e.g., SIR1)

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Mechanistic studies Evidence that NADH mediates
toxic effects of glucose


a
a
a
a
cells alive (CCK)
100
b
50
b
0
5 15 0 15 0 15
Glucose Lactate Pyruvate 5 mM
Glucose
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High-throughput screen discovers drugs protective
against glucose toxicity
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High-throughput screen discovers drugs protective
against glucose toxicity
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OK, high glucose can cause oxidative damage
acutely, but what accounts for the progressive
and (apparently) irreversible nature of
age-related impairments?
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Hypothalamic glucose sensor similarities to and
differences from pancreatic beta-cell
mechanisms.Yang, Kow, Funabashi, MobbsDiabetes
1999 Sep48(9)1763-72
The pancreatic, but not hepatic, form of
glucokinase was expressed in the VMH..These data
suggest that glucose-responsive neurons sense
glucose through glycolysis, specifically the
production of NADH, not ATP.
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Implication of glucose switch

• Glucose activates its own metabolism (esp. NADH)
• Cytoplasmic NADH is the unique signature of
  glucose, thus particularly suited as a signal
• This implies a self-perpetuating, cumulative
  sensitization or priming effect HYSTERESIS!
• Could this be reversed?

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Multistability in the lactose utilization network
of Escherichia coliOzbudak et al.Nature (2004)
Feb 427737-740
The phase diagram of the wild-type network shows
that lac induction always takes place
hysteretically, with cells increasing their
expression levels discontinuously as a switching
threshold is reached.
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Molecular hysteresis in metabolic gene expression
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Implication Aging is reversible
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Mechanistic studies Evidence that NADH mediates
toxic effects of glucose


a
a
cells alive (CCK)
100
b
50
b
0
0 10 0 10
Glucose (mM)
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Obesity What is the question?
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How much variability in genetically identical
laboratory mice? About the same as in humans!
.
20
.
.
.
Body Wt Gain (gm)
15
.
.
10
.
5
0
Coefficent of variation 47!
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Variability in caloric intake is much less than
variability in weight gain
.
.
.
.
.
.
Caloric intake (kCal)
500
0
Coefficent of variation 8!
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20 of retired breeder C57Bl/6J mice dont gain
weight on a high-fat diet but eat each just as
much
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Diet-induced obesity (independent of food intake)
depends on genotype

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20
Body weight Gain on HF Diet (grams)
15
10
5
A/J C57
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Hypothalamus
According to glucostatic hypothesis A/J mice are
resistant to diet-induced obesity because they
have increased hypothalamic sensitivity to
glucose.

• This hypothesis implies that induction of genes
  by hypoglycemia will be less robust in
  (glucose-sensitive?) A/J mice than in
  (glucose-insensitive?)

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GLUT 1
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Ubp41
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CITED-1
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NPY and AGRP
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AGRP (in vitro)
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Liver and Cortex

• Evidence of a hypothalamic defect

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Liver Glut1
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Cortex Glut1
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Humanized monoclonal antibody reverses genetic
obesity
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Anti-obesity effect of FGF antibody is at least
partially independent of feeding