Dr' Lloyd Old

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???????? ??? - 1975 ?.
Dr. Lloyd Old
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Plan for the minicourse.

• Lecture 1.
• Introduction to TNF and TNFR superfamilies.
• Signaling paradigms. Evolution.
• Introduction to genetic knockout.
• Lecture 2.
• Tumor Necrosis Factor.
• Genomics, regulation.
• Physiological functions good and bad TNF.
• Lecture 3.
• Lymphotoxins.
• Organization of lymphoid tissues.
• Physiological functions. LT produced by
  lymphocytes.

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Cytokine classification/nomenclature.

• According to the type of the receptor/intracellula
  r signal
• EGF, PDGF, VEGF internal tyrosine kinases.
• TGFb (1-3) internal serine-threonine kinases.
  
• Hemopoietins (except M-CSF) and most of
  interleukins (except IL8, 10, 17, 18) class I
  cytokine receptors associated with JAK kinases.
• M-CSF (CSF-1), SCF tyrosine kinases.
• Interferons and IL10 class II receptors
  associated with JAK kinases.
• IL1 family (IL1a,b, IL1RA, IL18) Toll-like
  signalling (adaptor molecules, TRAFs).
• TNF family (2 subfamilies) special classes of
  adapter molecules acting based on homotypic
  interactions (DD, DED, CARD), TRAFs.
• Chemokines 7-transmembrane domain receptors,
  G-proteins.

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TNF and TNFR superfamilies.
???????? ???, ?? ? ?? ?????????, ? ?????
????????? CD40 ? FAS (Apo-1, CD95) ???? ??????? ?
?????????? ???????????????? ? ?????????????
?????????????. ???????? ? ?????????????? ????
???????? -?????? ????????????? ??????????
??????????? ???????? ???????? ?
??????????????. ????????????? ?????? ????????? ?
???? ???????, ? ?????????, ??????????.
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TNF and TNFR superfamilies
Modified from Locksley et al, 2002
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TNF Superfamily

• The prototypic members of this superfamily are
  TNF and lymphotoxin (LT-a)
• About 20 ligands and 30 receptors. Over 40
  distinct ligand-receptor pairs are known (and
  more are likely to be found).
• These cytokines regulate immune response and host
  defense, as well as development, organization
  and homeostasis of lymphoid, neuronal, ectodermal
  and other tissues.
• Most TNF family ligands are membrane-anchored
  trimers. Some of them may be cleaved
  extracellularly by metalloproteinases (e.g., TACE
  for TNF), and the released soluble trimeric
  ligands trigger receptor responses.

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TNF Receptor Superfamily.

• Extracellular portions contain 1-4 cystein-rich
  domains CXXCXXC, with three disulfide bonds
  creating an elongated molecule.
• Forms (homo) trimers which interact with ligand
  trimers through clefts.
• Intracellular portions do not possess kinase
  domains, and are not docking tyrosine kinases in
  response to ligand binding.
• Transmit signal through distinct class of adapter
  molecules.
• Can transmit two types of signals activation
  (proliferation, differentiation, costimulation)
  and programmed cell death (PCD).

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TNF-TNFR additional complications

• Some cytokines can engage more than one receptor
  (ex. TNF, LT, LIGHT, TRAIL, etc).
• Cross-utilization of receptors by the TNF family
  ligands is also common (ex. TNF/LT and TNFR1,2).
• Moreover, mixed trimeric ligands can exist (ex.
  LTa/LTb) and this changes the receptor
  specificity (from TNFR1 to LTbR).
• Finally, TWEAK can form fusion with APRIL at the
  level of RNA splicing. Function of the fused
  ligand TWEAPRIL is unknown.

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?????????? NFkB, IkB ? IKK ??????????
From Barnhart Peter, 2003 Cell 114148
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??????? ? ????????? ????????? ??? ???????
LTa3
TNFR1
TNFR1
LTa3
TNF3
TNFR1
TNFR1
TNFR1
TRAF3
TNFR1
TNFR1
LTa
TRAF
TRAF
TNFR1
CD40 peptide
TRAF
Modified from Locksley et al, 2002
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Chen Goeddel, 2002
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???????? ????????? ?????????????? ????????.

• FAS-L (Apo-1L) ????????? ????? FAS (Apo-1, CD95).
• TRAIL (Apo-2L) ????????? ????? DR4, DR5.
• TL1a ????????? ????? DR3.
• TNF ????????? ????? TNFR1 (p55, CD120a).
• ??? ??? ????????? ???????? ? ???? ???????????????
  ????? ?????? ?????? ??? ??????????????
  ?????-???????? ?????????????? ????? ??????.

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Dempsey et al., 2003
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Micheau Tschopp, Cell 2003
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??-????????????? ???????? ? ????????? ?????????
??? ? ???-?

• ??????? ????? ?????????? ??????????????? ?? ?
  ?????????? ? ???????????? ????????
  ??-?????????????? ??????? ?? ???????-?????????????
  ? ? ???? ??????.
• CD40 ??????????????? ?? ? ??????? ? ??????????
  ???????, ? ???????? ??-?????????????
  (?????????) ?????? ?? ? ??????.
• ?????????, ??? ?????? ????? GITR ????? ???
  ???????? ???????????? ? ??????, ?? ???????
  ?????????? GITR ????????. ?? ????????? ???? ????
  ?????? ?? ?????????.

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??-????????????? ????????? ?? ? ???????
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??????? ??-????????????? ????????
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?????????, ?? ?????????? ????? ??????, ?????
????? ?????????? ???-?????????????? ??????
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???????? ????????? ??? ??????? ????? ???
????????? ? ??????

• CD40L-CD40.
• BAFF (Blys)-BAFF-R.
• TNF, LTa TNFRI (p55)

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In situ EDAR expression in hair follicles, E15
embryo
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Fig. 1. Ectodysplasin-EDAR signaling in hair
placodes. (Top) Molecular detail of
ectodysplasin-EDARsignaling. The expression of
ED1 gene in the interfollicular cell is induced
by Wnt signals transduced by LEF-1. ED1 encodes
the TNF molecule ectodysplasin, which is
synthesized as an integral membrane protein.
Cleavage is required for the biological activity
of ectodysplasin, which binds to EDAR in the
placodal cell. EDAR signaling activates the NF-?B
pathway. Only those molecules that have been
shown to be essential for EDAR signaling are
indicated. EDAR uses the death domain adaptor
EDARADD for signal transduction. IKK? (NEMO) is
required for the release of cytoplasmic NF-?B,
allowing its transport to the nucleus. EDAR
expression is stimulated by mesenchymal signals
including activin, but the downstream targets of
EDAR signaling are currently not known. (Bottom)
Whole mountin situ hybridization analysis of
embryonic mouse skin reveals complementary
expression of the ligand ectodysplasin (left) and
its receptor EDAR (right). EDAR expression is
confined to hair placodes (ectodermal thickenings
developing into hair follicles), whereas the
ligand ectodysplasin, encoded by the ED1 gene, is
expressed in the interfollicular domains.
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??? ? ?? ??????????, ????? ????????? ? ???????? ?
???????? ?/??? ? ???????? ? ???????? ???????

• ???????, ?????????? ? ?????????? ? ???????
  ???????? ??????? (??????? gld, lpr, tabby,
  downless, crinkled).
• ???????????? ??????? (????????, ? ??????????
  ?????).
• ??????????? ??????? ????????? ???????
  ???????????? ????? ???? ? ????????????, ?? ?????
  ????? ????????? ? ???????????????????? ?????????.
  

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• ?????????????? ????????? (? ?????), ??????? tabby
  (? ?????)
• ??????? ??????? ????? ? ????????? ??????????
• ??????? ??????????? ?????? ????????
• ????????????? ?????? ?????? ??????? ???
• ??????? ???????? ?????????? ??????????? ?????????
  ????
• ???????? ??????? Eda -gt Edar -gt EDARADD.

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??? ? ????????
TNF
TNF
-R
-R
In Drosophila, TNF Eiger, TNFRWengen, adapter
proteindTRAF2 FADD/DREDD signaling in Drosophila
is in Toll/IMD pathway, not in TNFR pathway.
Apparently, vertebrates borrowed this pathway
for TNFR signaling later in evolution.
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TRAFs ? ????????
H- human Ce- C. elegans Hy- Hydra D-
Drosophila
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??????? ???????? ? ??????? ???????? ????????

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• ???????? ?1 ??-?? ????????????? ???????
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  ???????? ????.
• ???????? ?2 ??????????? ??? ????? ??????????
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  ????????? ????????? ???????? ?????.
• ???????? ?3 ???????? ??????? ????? ?????? ??????
  ???????, ??? ??????. (?????????? ???-??
  ????????? ?????????? ???????? ? ?????????
  ???????? ????????).