Cell cycle and Morphology Transition

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Cell cycle and Morphology Transition
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Cell shape

• Candida can grow in a variety of morphological
  forms
• Yeast -unicellular budding
• True hyphae - parallel-sided walls
• Pseudohyphae - daughter bud elongation and
  remaining attached to the mother cell
• The ability to switch between morphologies is
  necessary for virulence

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• Hyphae and pseudohyphae are invasive - could
  promote tissue penetration, colonization of
  organs
• Yeast form might be more suited for dissemination

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The effect of environmental conditions on
morphology
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Signal transduction pathways govern morphology
transition

• cAMP protein kinase A pathway
• Mitogen activated protein kinase pathway
• pH response Rim101 pathway
• Solid matrix related Czf1 pathway
• Negative regulators -Tup1 Nrg1, Rpg1

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Colony switching white-opaque switching

• Epigenetic phenomenon
• Colony
• Yeast cells form smooth white dome-shaped
  colonies, oval shape yeast cells
• Opaque cells form opaque flat colonies,
  elongated oblong shape, twice the size of white
  cells, competent to undergo mating

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• Only cells with homozygous mating type locus form
  opaque cells
• Opaque cells signal white cells to form a a
  biofilm
• facilitates mating between opaque cells.

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Cell cycle

• Cell cycle is regulated in a different way in
  each cell type
• The subcellular components are rearranged/localize
  d differentially during the cell cycle
• Actin patch
• Actin cable
• Septin ring
• Septum
• Nucleus

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Actin patch/cable

• Core components of actin cytoskeleton
• Critical for establishment of cell polarity
• Actin cables bundles of actin that align along
  the axis of polarization
• Actin patches cortical membrane zones invested
  with F-actin and actin binding/regulating
  proteins
• Actin patches are enriched at sites of polarized
  cell surface growth

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Septin

• septin ring forms during cell division
• septin ring serves as a "dock" and proteins bind
  to it in an asymmetrical fashion
• Known functions
• Scaffold recruit other proteins
• Cytokinesis septum formation
• Anchoring site for bud site selection factors

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Septum

• Cross wall between mother and daughter cell
• Prevent lysis during separation of the two cells
• Chitin ring
• Contractile actomyosin ring, invagination of the
  plasma membrane
• Chitin disk formed (primary septum)
• Secondary septum formed (similar composition to
  the cell wall)

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Yeast
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Pseudohyphae
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Hyphae
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(No Transcript)
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Polarized growth

• Polarized growth is associated with polarization
  of actin cytoskeleton
• Polarized growth in yeast form
• Bipolar at least 2 scars on opposite ends of
  cell
• Unipolar 1 scar on one end, 3 or more on
  opposite end
• Axial 4 or more scars grouped together on one
  side
• Random scars occuring randomly and ungrouped
  around cell
• Polarized growth in hyphae form continuously
  in a polarized fashion
• yeast isotropic growth , Hyphae apical
  growth

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Virulence Factors of Candida albicans
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Early events in the pathogenesis of candidiasis
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Detection methods

• Gene disruption C. albicans is diploid,
  transformation yield is low
• Phenotypic evaluation in vitro (adherence,
  growth rates, morphogenesis, etc)
• Animal study murine model of hematogenously
  disseminated disease
• survival
• Tissue colony count

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Virulence factors Adhesins

• Adhesins biomolecule that promotes the
  adherence of C. albicans to host cells/ligands
• Function host recognition/binding

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Als1p and Als5p

• Agglutinin like sequence
• In S. cerevisiae, a-agglutinin is required for
  cell-cell recognition during mating
• In C. ablcans, 7 Als proteins are known
• Three domain 5 domain (relatively conserved),
  central domain (tandem repeat), 3 domain (serine
  threonine rich)
• Attached to the cell wall by glycosylphosphatidyli
  nositol anchor
• Als1p and Als5p appear to provide an adhesin
  function
• Als1p is essential for virulence in
  hematogenously disseminated murine model

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Hwp1p

• Hyphal and germ tube specific protein
• Outer surface mannoprotein
• Covalently linked to cell wall glucan
• N-terminal proline and glutamine-rich domain
• Resemble transglutaminase substrates
• Binding of C. albicans hyphae to human buccal
  epithelial cells by stable and covalent bonding
• KO reduced virulence in HDC

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Int1p

• C. albicans binds to several ECM ligands
  (fibronectin, laminin and collagen I and IV)
• Int1p Integrin like protein (18 homology )
• First identified by immunological analysis using
  human anti integrin antibody
• Functional analysis of Int1p
• Antibody
• Heterologous expression
• Gene knockout analysis ( for adherence and
  virulence)

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Adhesins of C. albicans and C. glabrata
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Morphogenesis

• Transition between yeast and filamentous growth
  form
• The mutant strains lack of the ability to switch
  morphology between either form are less virulent
  in animal study
• Form-specific gene expression
• Morphogenesis is governed by several signal
  transduction pathways

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Other protein kinases also govern
morphogenesis PKC, HOG1,SLN1, CHK1
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Transcriptional proteins of C. albcans

• Table 2

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Enzymes that contribute to invasiveness

• Phospholipases (PLA - D)
• PLB1 is associated with virulence
• 84kDa glycoprotein, secreted
• Mutants has 60 reduced host cell killing ability
• Plb1p activity localize at hyphal tips

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Enzymes that contribute to invasiveness

• Secreted aspartyl proteinases (SAP)
• At least nine proteins comprise in SAP family
• Sap1-6 deletions attenuated virulence during
  invasive disease
• In vitro in vivo expression on SAP production
  demonstrated stage specific expression of SAPs

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• In vitro oral epithelial cell
• SAP1 and 3 expressed within 42hr post infection
• SAP6 48hr (most extensive tissue damage)
• SAP2 and 8 60 hr
• SAP4 and 5 never detected
• In vivo human epidermis model
• SAP 1, 2 early invasion
• SAP8, 6 and 3 extensive penetration and
  extensive hyphal growth
• Vaginitis models SAP2 is required for disease
  development

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Phenotypic switching

• White-opaque switching
• Opaque normally dont germinate except if the
  opaque cells were grown on human skin epithelial
  cells
• Differential gene expression
• OPA SAP1 is opaque-specific
• SAP2,3 is white-specific
• Role in virulence ??
• Freshly isolated strains have much
• higher frequencies of switching
• Opaque skin infection, less virulent in
• systemic animal model