Summary Class notes - HAP-31806 Molecular regulations of health and disease

- HAP-31806 Molecular regulations of health and disease
- Vincent de Boer
- 2016 - 2017
- Wageningen University (Wageningen University, Wageningen)
- Biology
201 Flashcards & Notes
2 Students
  • This summary

  • +380.000 other summaries

  • A unique study tool

  • A rehearsal system for this summary

  • Studycoaching with videos

Remember faster, study better. Scientifically proven.

PREMIUM summaries are quality controlled, selected summaries prepared for you to help you achieve your study goals faster!

Summary - Class notes - HAP-31806 Molecular regulations of health and disease

  • 1473026400 Theme 1: Molecular regulation of energy and nutrient metabolism

  • 3-bromopyruvaat can be used as an anticancer agent. It is a glucose blocker and targets the proteins: Hexokinase, GAPDH, pyruvate kinase and phosphoglycerate kinase. It can also target: Succinate dehydrogenase, malate dehydrogenase and glutemate dehydrogenase. Those are needed in different steps during the glycolysis from glucose to pyruvate. The 3-bromopyruvaat then forms an alkylated protein.

    3-bromopyruvaat enters the cell via MCT1 which is presented on cancer cells (but not likely on normal cells).
  • What are the hallmarks of cancer (name all 10)?
    The hallmarks of cancer are the underlying principles of the complexity of cancer on a cellular level.

    1. Sustaining proliferative signalling.
    2. Resisting cell death.
    3. Evading growth supressors.
    4. Enabling replicative immortality.
    5. Inducing angiogenesis.
    6. Activating invasion and metastases.

    7. Deregulating cellular energetics.
    8. Avoiding immune destruction.
    9. Tumor promoting inflammation.
    10. Genome instability and mutation.
  • Cancer cell metabolism is programmed to optimize cell growth, this means that a major goal of cancer cells is to produce the three major constituents of the cell: protein, DNA and membranes. The major fuels that are consumed by cancer cells are glucose, glutamine and fatty acids. ATP, NADH and NADPH are needed for all catabolic and anabolic processes to proceed. (PPP --> pento phosphate pathway)
  • Cancer cell vs normal cell:

    Do not require growth signals.

    Loss of contact inhibition. Reduced adhesion. Loss of anchorage dependence.          

    More mobile surface proteins. 
    Altered secreted protein profile.
    Resistant to programmed cell death (apoptosis) and necrotic cell death.

    Altered nutrient and energy metabolism:
    Glycolytic metabolism (facilitates growth). Sustained angiogenesis (facilitate nutrient supply). Negative cell surface (facilitates nutrient import).

    Cancer cells are not fundamentally different
    Cancer cells are re-programmed such that optimal growth of the individual cell is facilitated (at the expense of the organism to which the cancer cell belongs). Reprogramming results in improved fitness to provide a selective advantage during tumorigenesis

    Reprogrammed activities related to metabolism:
    • Altered bioenergetics
    • Enhanced biosynthesis
    • Optimized redox balance
  • What are the two types of phases in cancer cells and what is their metabolic geprogrammed for?
    Metabolic reprogramming to maximize:
    • Biomass
    • Energy (ATP)

    Metabolic reprogramming to maximize:
    • Alternative fuels
    • Anti-oxidant defense
  • What is the Warburg effect?
    Rate of glucose uptake increases and lactate is produced even in the presence of oxygen and fully functioning mitochondria.
  • How does the Warburg effect benifit cancer?
    • Rapid ATP synthesis: glycolysis to pyruvate is faster then al the way through the mitochondria.
    • Biosynthesis: glycolysis to pyruvate has multiple products that can be used for DNA and proteins. Reductive carboxylation allows biosynthesis during hypoxia. 
    • Tumor microenvironment: accidification promotes invasiveness. Taking away the glucose from native immune cells.
    • Cell signalling: allows for signalling transduction through ROS and/or chromatin modulation.

  • Mitochondria
    • Mitochondria are not impaired in cancer cells, but provide intermediates for biosynthesis.
    • Aerobic glycolysis provides the cancer cell with sufficient ATP and precursors for biomass production.
    • NADPH production is needed for lipogenesis and anti-oxidantdefense.
    • Glutamine plays a crucial role in sustaining cancer cell growth.
    • Reductive carboxylation occurs during hypoxia for lipid biosynthesis.
  • Cataplerosis: efflux of TCA cycle intermediates.
    Anaplerosis: influx of (other) TCA cycle intermediates.

    Several anaplerotic (replenishing) mechanisms exist. Predominant is the use of various amino acids to replenish the TCA cycle, in particular alanine and glutamine/glutamate: glutaminolysis. Glytaminolysis is the breakdown of glutamine to pyruvaat/lactate.

    Also reductive carboxylation occurs during hypoxia for lipid biosynthesis. (see other note)
  • Functions of mitochondria:
    • ATP production.
    • Respond dynamically to cellular energy demand.
    • Regulate use of energy substrates (lipids, sugars, amino acid).
    • Regulate urea cycle.
    • Essential for iron metabolism (heme and FeS synthesis)
    • Regulate calcium homeostasis.
    • Regulate amino acid metabolism.
    • Mediate apoptosis and programmed necrosis.
    • Mediate innate immune defence.
    • Regulate oxidative signalling, mediated by reactive oxygen species (ROS)

    Mitochondrial function is substantially altered in cancer cells, which determines cell death resistance and altered metabolism.

    Summary mitochondria:
    • Mitochondria are complex organelles
    • Mitochondria are controlling metabolism in many different ways
    • Mitochondria in normal cells provide them efficiently with ATP
    • In cancer cells mitochondria are needed for biosynthesis
    • Oxidative phosphorylation in cancer cells is needed to oxidize NADH with oxygen
  • TCA cycle = tricarboxylic acid cycle = citric acid cycle = Krebs cycle: Substrate oxidation.
  • Oxidative Phosphortylation (OXPHOS). Electron transport chain(s) generates membrane potential. FOF1 complex (ATP synthase) uses membrane potential to generate energy (ATP).
  • OXPHOS detailed.
  • Overview of mitochondrial energy metabolism (OXPHOS and TCA cycle).
  • Do cancer cells need mitochondria?
    Pyruvaat, (NADH-->NAD+), AKB, nucleotides and aspartate rescue tumor cells with OXPHOS deleted mitochondria. Aspartate biosynthesis is essential for mitochondrial defective cells. Oxidative phosphorylation in cancer cells is needed to oxidize NADH with oxygen.

    Mitochondria are needed for biosynthesis and oxidative phosphorylation.
  • Aspartate synthesis.
    NADH is generated in mitochondria, by substrate oxidation, isocitrate ---> a-KG. O2 oxidizes NADH to NAD+ but if there is no oxygen then pyruvate broken down into lactate generates NAD+. This is needed for aspartate biosynthesis.
  • Regulation.
    Growth and proliferation:
    • AKT/PI3K
    • mTOR
    • PKM2

    Survival and metabolic adaptation:
    • AMPK
    • Beclin 1 autophagy
    • HIF1alpha

    Other transcription regulators:
    • MYC
    • P53
  • Metabolism is controlled by signalling. (and vica versa)
    Regulation of metabolism in normal cells is dependent on external signals. regulation of metabolism in cancer cells is largely autonomous (not dependent on growth factors). The same pathways operate, but external signalling pathways are often mutated in cancer cells. Among most frequently mutated "cancer" genes are TP53, LKB1, MYC, genes of AKT/mTORC1 signalling.

    Regulation of cancer cell metabolism can be altered on different levels.
    • DNA (mutations: altered genes --> proteins, epigenetics, DNA methylation, histone modifications)
    • mRNA (transcription factors: genes)
    • Protein (Protein signalling (phosphorylation))
    • Metabolites (fatty acids, glucose, amino acids, etc.)

    Proliferation state of cancer cells can be regulated by AKT, mTOR and PKM2. Survival state of cancer cells can be regulated by HIF1alpha, AMPK and autophagy. (see picture)
  • AKT signalling.
    Mutations in PI3K/AKT signalling occurs in >70% of all breast cancers.

    PI3K phosphorylates a lipid signalling molecule. (PIP2---> PIP3). PIP3 brings PDK1 and AKT in proximity. AKT can inhibit FOXO which helps in cell cycle and cell survival and it kan also inhibit mTORC1, which helps with protein synthesis.

    AKT controls cell growth, but also:
    • Metabolism
    • Angiogenesis
    • Glucose uptake
    • Survival and
    • Proliferation.
  • mTOR.
    mTOR = mechanistic target of rapamycin = protein complex.
    You have mTORC1 and mTORC2 (not much known about 2).

    • Signal integrator complex
    • Characterized by mTOR and RAPTOR
    • Regulates balance between cell survival and cell proliferation.
    • mTOR is a phospho-inositide-3-kinase (PI3K)-related kinase
    • Nutriënt (amino acid, glucose, oxygen), energy and growth factor sensitive

    mTORC1 has effect on:
    Translation, proliferation, survival, lipid biogenesis, autophagy and angiogenesis.

    Most wel known targets are 4EBP and S6K. 4EBP inhibits elongation factors and S6K activates translation of mRNA.
  • TSC1/TSC2 complex regulates mTORC1. Genetic mutation in TSC1/TSC2 lead to tuberous sclerosis and cells can lose controle of cell growth and division. This has effect on entire body, known parts it has an effect on are: brain, kidney, heart and skin. It forms benign tumors.
  • GTPase.
    • GTPase is active when bound to GTP
    • GEF and GAP control this activity
    • GDI binds GDP bound GTPase and prevents translocation to membrane
  • mTOR signalling is integrated at lysosomes. (See picture) in the first part GTPases are inactive. When amino acids signal to Rag it is bound to GTP which brings mTORC1 to membrane (of lysosome). Growth factors activate AKT, phosphorylation of TSC relieves TSC from Rheb, Rheb can be activated. There is only full activation of mTORC1 when both signals (amino acids and PI3K (growth signals)) are available.
  • mTORC1 summary:
    • mTORC1 controls cell growth
    • mTORC1 is regulated by nutrients and growth factors
    • mTORC1 signalling is integrated at the lysosomes
    • GTPases serve as switches for mTORC1 activity
    • mTORC1 activates protein synthesis via S6K and 4EBP1
    • mTORC1 inhibits autophagy
    • mTORC1 is active in cancer cells
  • AMPK
    • Cellular energy master regulator
    • Trimeric protein
    • Regulated by AMP/ATP (LKB1 =  kinase)
    • Regulated by phosphorylation
    • Downstream it controls mTORC1 and lipid metabolism
    • (many downstream targets (complex mechanism))

    Lipid metabolism
    ACC stimulates:    acetyl-CoA --> malonyl-CoA --> fatty acid --> lipid
    fatty acid --> acylcarnitine (cytosol) --> (via CPT1) acylcarnitine (mitochondria) --> acetyl-CoA
    Malonyl-CoA directly inhibits CPT1.

    When AMPK is active, ACC is inactivated and therfore lipid breakdown is stimulated.

    When AMPK is inactive, TSC2 and Raptor is active and activate mTORC1 which inhibits autophagy. Thanks to the inactive AMPK, ULK1 is inactive and does not stimulate autophagy.

    When AMPK is active, TSC1 is active and Raptor is inhibited. Also ULK1 is activated and therefore autophagy is stimulated.

    Upstream regulation
    Growth factor with PI3 Kinase which stimulates AKT. AKT inactivates AMPK. Calcium, DNA damage and TGF signalling stimulate AMPK (together with AMP/ATP (LKB1)).

    AMPK can expert pro- or anti-tumor effects based on context (see picture).
  • Hypoxia inducible factor (HIF)
    • HIF is a complex of HIF1A (regulated), ARNT, EP300, CREBBP
    • HIF recognizes and binds to hypoxia response elements (HRE) in DNA
    • HIF regulates transcription of genes under control of HRE (metabolic adaptation, oxygen supply, stress resistance, angiogenesis)

    1. HIF1A is recognized by EGLN and HIF1AN
    2. EGLN and HIF1AN hydroxylate HIF1A
    3. HIF1AN hydroxylation prevents binding of cofactors EP300 and CREBBP (no transcription)
    4. EGLN hydroxylation facilitates VHL binding
    5. VHL recruits E3 ubiquitin which polyubiquitinates HIF1A
    6. Ubiquination of HIF1A labels it for proteasomal degradation (HIF1A degradation)

    Function of EGLNs and HIF1AN critically dependent on
    • Fe2+
    • 2-oxoglutarate (
  • Summary HIF1
    • HIF1alpha is turned on in cancer cells even if there are high oxygen levels
    • EGLN and VHL normally keeps HIF1alpha out of the nucleus
    • Succinate, fumarate and 2-HG interfere with EGLN2 protein (a-KG dependent reaction)
    • HIF1alpha mainly transcribes glycolytic genes. 
  • PKM 1/2
    Pyruvate kinase, PEP ---> pyruvate (ADP ---> ATP)
    There are pyruvate kinase isoforms, PKM1 is found in muscle and PKM2 is found in tumors.

    PKM1 has high constitutive activity
    is not activated by upstream fructose 1,6-biphosphate (F1,6BP)

    PKM2 has low activity
    is activated by upstream fructose 1,6-biphosphate (F1,6BP)
  • PKM2 activity is tightly controlled
  • PKM2 summary
    • Switching between isoforms can be beneficial for cancer cells
    • Tumor cells adopt PKM2 for optimal growth
    • PKM1 is expressed in differentiated, ATP-demanding cells (brain, muscle)
    • PKM2 gives tumors cells more control over glycolysis and PPP

    PKM2 regulation is different from transcription factor regulation or phoshporylation regulator.
  • Gliomas are the most common primary brain tumors. They arise from glial cells (supporting neurons). Brain tumors are difficult to treat because of the blood brain barrier.

    Isocitrate dehydrogenase (IDH) is mutated in glioblastomas and is a TCA cycle enzyme converting isocitrate into alpha ketoglutarate (a-KG). The mutant instead converts a-KG to 2-hydroxyglutarate (2HG). 2HG is highly elevated in tumors.

    Oncometabolites = metabolites whose abnormal accumulation causes metabolic or non-metabolic dysregulation and potential transformation of normal cells to cancer cells.
  • What are the 3 (known) oncometabolites?
    • 2-hydroxyglutarate
    • Succinate
    • Fumerate
  • 2-HG inhibits DNA demethylases and histone demethylases of anti-cancer proteins/metabolites and has effect on prolyl hydroxylases (enhanced degradation of HIF1alpha in gliablastoma, but inhibited degradation of HIF1alpha in other tumors).

    2-HG, succinate and fumarate have effect on DNA demethylases, histone demethylase, prolyl hydroxylases (2-HG). Fumarate and succinate also have effect on succination of antioxidant defense proteins and succinylation.

    • 2-hydroxygluterate, succinate and fumarate are oncometabolites
    • oncometabolites can drive cancer progression by:
    1. activating HIF1alpha (or 2-HG deactivating it)
    2. epigenetic alteration
    3. altering ROS and ROS signalling
    • Oncometabolites discovery opens up possibilities for therapeutic targeting
  • Alternative fuels.
    Cancer cells need to adapt to different matabolic environments. Low glucose and hypoxic conditions can let cancer cells choose different substrates.

    Alternative fuels via:
    • Fatty acid oxidation
    • Protein uptake (macropinocytosis)
    • Autophagy
  • Fatty acid oxidation
    CPT1 = carnitine palmitoyl-transferase. It is used to get palmitoyl-carnitine from cytosol into mitochondria.

    CPT1c is highly expressed in human brain tumors. CPT1c knockdown slows tumor growth. CPT1c protects cells from apoptosis, embryonic stem cells are protected from apoptosis during hypoxia by cpt1c.
  • Cancer stem cells.
    • Quescent cells, in tumors that are resistant to therapy
    • Low metabolic rates
    • They are set to survive instead of proliferation
    • Eradicating these cells would prevent disease relapse and residual disease
    • Are able to renew the cancer cell pool
    • Tumor initiating cells (TICs), dormant tumor cells

    Cancer stem cell metabolism
    normal cancer cells use energy metabolism for growth, work, division. Cancer stem cells use energy metabolism for defense, work and survival. It mainly uses fatty acids and autophagy as substrates.
  • Metabolism loss of attachment
    Loss of attachment normally induces apoptosis (anoikis).Cancer cells have anoikis resistance. Detached cells cannot take up glucose, therefore cancer cells need other things to survive. Glucose is needed for PPP (pento phosphate pathway). Metabolism of detached cancer cells is altered. Which can make these cells resistance to cell death (apoptosis).
  • Protein uptake (macropinocytosis)
    Macropinocytosis is the uptake and catabolism of extracellular proteins to intracellular amino acids. (see picture)

    There are three types of vesicle formation:
    • Lamelipoda-like
    • Circular ruffle
    • Bleb
  • Summary
    • Tumors are heterogenous and do not only contain cells that are rapidly proliferating
    • Fatty acid oxidation is used as alternative source for ATP, when glucose is scarce
    • fatty acid oxidation is an important metabolic pathway in cancer stem cells
    • Anoikis resistance can be overcome by anti-oxidant or upregulation of FAO
    • Macropinocytosis is an alternative fuel pathway when amino acids are scarce
  • Autophagy
    Autophagy is one of the breakdown pathways. First there is autophagosome formation, then there is an autophagosome, this fuses with a lysosome and the content gets degraded and released. Macroautophagy can be selective or non-selective. Non-selective autophagy degrades cytosolic proteins and organelles. Mitophagy selectively degrades mitochondria.
  • Normal role of autophagy
    Autophagy is a process for:
    • Removal of damaged parts
    • Recycling/redistribution of building blocks
    • Emergency energy supply

    Cell growth requires:
    • Growth factors
    • Nutrients
    • ATP production and biosynthesis

    Lack of these growth requirements --> starvation --> autophagy

    Autophagy is a pro-survival process, but continued autophagy leads to cell death.
  • Phagophore formation has three phases.

    Phase 1:
    • ATG1 (ULK1) complex formation
    • Extraction of lipid membranes by ATG9

    Phase 2:
    • Beclin-1 complex formation
    • Recruitment of Atg5-Atg12-Atg16 and LC3

    Phase 3:
    • Membrane elongation by Atg5-12-16 and
    • coating (protecting) membrane
  • Phagophore formation Phase 1
  • Phagophore formation phase 2
  • Phagophore formation phase 3
  • AMPK and mTORC1 regulate autophagy

    • Autophagy is a survival mechanism of the cell
    • Autophagy is mediated through a large set of Atg proteins (eg. Atg1=ULK1)
    • Beclin1 complex initiates phagophore formation
    • Autophagy is primarily controlled by mTOR and AMPK
  • What are the four kind of cancer therapies?
    Immune therapy
  • During WWII they developed new agents and protective mechanisms --> mustard gas decreased lymphomas. "Alkylating agents" block DNA replication --> cisplatin (currently used in clinic).

    1957 Sidney Farber (Boston) --> Methotrexate --> blocks DNA synthesis and is an anti-folate, also called anti-metabolites. Folate is cofactor for methyltranferases. Methotrexate is an inhibitor of dehydrofolate reductase (DHFR).

    Folate --> (via DHFR) THF --> Thymine --> DNA

    Other widely used anti-metabolites:
    Purine/pyrimidine analogues block DNA synthesis
    5-fluorouracil (5-FU) --> inhibits thymidine synthesis

    PD1 ligand blockade
    PD1= receptor for activating T cells
    PD1L= ligand that blocks the activation of T cells, highly expressed on cancer cells not on normal cells.
    effective therapy when compared to other drugs 

    • Chemotherapy is non-specific and also targets normal cells
    • Search for new types of drugs that are more selective
    • Targeted therapy development has proven useful in CML
    • Immunotherapy is a promising new treatment area
  • Rapamycin
    1977 it was discovered that rapamycin has immunosuppressant function in rat models. It is now used clinically against tranplantation rejection. It blocks T cell and B cell proliferation, blocks IL-2 function. It blocks tumor growth in rats, blocks tumors in mice. Rapamycin analogues are used in the clinic since 1999 (everolimus (afinitor) and sirolimus)

    • Prevention of allograft rejection
    • Anti-restenosis in blood vessel stents
    • Advanced kidney cancer
    • Mantle cell lymphoma

    Advanced breast cancer after treatment with aromatoase inhibitors. Upregulation of mTOR was seen. Everolimus blocks in combination with a non-steroidogenic drug. (progression free survival 6.9 mo vs 2.8 mo) But also many clinical trials failed --> it is not effective by itself.

    Mechanisms why rapamycin doesn't work:
    1. Rapamycin is not cytotoxic but more cytostatic, tumors regrow after drug is taken away
    2. Fails to complete inhibit protein synthesis, 4E-BP1 phosphorylation is less affected than S6K phosphorylation
    3. Feedback loops are activated when mTOR is inhibited
    4. Other kinases take over regulation of S6K
    5. RNA expression of eIF4E (elongation factor) is upregulated upon chronic mTOR inhibition
    6. Decrease mTOR leads to increased macropinocytosis

    Blocking mTOR activates AKT and Ras signaling (growth) because feedback loop is gone.(see picture)
  • Metformin
    • Used since 1957 for its glucose lowering effects in diabetic patients
    • Metformin inhibits gluconeogenesis
    • Metformin activates AMPK
    • Number one anti-diabetic drug in the world

    Metformin increases reductive carboxylation. Combination therapy with glutaminase inhibitor was succesful. CB-839 inhibits glutaminase, (glutamate --> glutamine).
Read the full summary
This summary. +380.000 other summaries. A unique study tool. A rehearsal system for this summary. Studycoaching with videos.

Latest added flashcards

What are Weibel-Palade bodies?
They are cells that store and release von Willebrand factor and P-selectin.
What are the functions of healthy endothelium?
  1. Endothelium dependent dilatation
  2. Anti-inflammatory (inhibition of leukocyte adhesion)
  3. Anti-coagulant and anti-thrombotic (inhibition of platelet adhesion and aggregation)
  4. Anti-hypertrophic (inhibition on vascular smooth muscle cells proliferation)   
In which step of the butyrate kinase pathway is ATP formed?
From butyryl-P to butyrate. see picture. Acetate to acetyl-CoA with butyryl-CoA to butyrate has no direct ATP production.
What are specific outer membrane substances for gram-positive and what for gram-negative bacteria?
Gram-positive: Lipoteichoic acid
Gram-negative: Lipopolysasscharide (LPS)
Name the indicated substances/parts of gram-negative cell envelope.
  • Cell wall
  • Outer membrane
  • Periplasm
  • Cytoplasmic membrane

  • O-polysaccharide
  • Core polysaccharide
  • Lipid A
  • Protein
  • Porin
  • Lipopolysaccharide (LPS)
  • Phospholipid
  • Lipoprotein            
Name the indicated substances.
  • Wall-associated protein
  • Teichoic acid
  • Peptidoglycan
  • Lipoteichoic acid
  • Cytoplasmic membrane    
What does the cell envelope of bacteria consist of?
  • Cell wall
  • One or two membranes (one membrane gram-positive, two membranes gram-negative)
  • proteins
  • other complex molecular structures   

Possible other envelope structures:
  • Flagella (for motility)
  • Fimbria (for attachment)
  • Pili (for attachment and conjugation)
What are the three hypotheses for "universal" functions of microbiota?
  1. All of us contain microbes that we al share: core microbiota
  2. Different microbes have similar functions
  3. Enterotypes  
Why is the ecosystem in the large intestine driven by conversion of complex indigestible polysacharides?
Because the transit time is slow (24-72 hours) therefore there is more time to ferment the products and the host metabolic absorbtion is lower. Upstream the simple sugars have been converted and absorbed by the host.
Name some functions of GI tract bacteria.
  • Fermentation of indigestible carbohydrates
  • Development GI tract structure and immune system
  • Resistance against pathogens
  • Improved metabolite absorption
  • Production short chain fatty acids (SCFA)
  • Production of vitamins
  • Deconjugation bile acids

many more....