IDGFs = Imaginal Disc Growth Factors
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Transcript of IDGFs = Imaginal Disc Growth Factors
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Growth factors in Drosophila:
Characterization of two novel families of growth factors,
IDGFs and ADGFs
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IDGFs = Imaginal Disc Growth FactorsIDGFs = Imaginal Disc Growth Factors
• secreted glycoproteins
• isolated from conditioned medium, where they are abundantly secreted by various Drosophila cell lines exhibiting macrophage properties (Cl8+, S2, Kc) (Kirkpatrick et al. 1995, Kawamura et al. 1999)
Detection of DS47 secreted by Dm cell lines1 - M3 media + serum2 - supernatant from S2 culture, serum-free3 - supernatant from S2 culture, serum+4 - supernatant from KcO culture, serum-free5 - supernatant from 1006-2 culture, serum+6 - supernatant from 1006-2 M culture, serum+7 - supernatant from Ox736-3 M culture, serum+8 - supernatant from 5697-5 culture, serum+9 - supernatant from 5697-7 culture, serum+
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IdgfIdgf genes in genes in Drosophila melanogasterDrosophila melanogaster
~ 55 % seq. similarity at cDNA level~ 50-60 % seq. similarity at aa level
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IDGF = Imaginal Disc Growth FactorsIDGF = Imaginal Disc Growth Factors
• IDGF proteins in cooperation with insulin are able to stimulate proliferation, polarization and motility of Cl8+ cells in culture (Kawamura et al.1999 – recombinant IDGF1 and 2)
Cl8+ in CM
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Human homolog HC-gp39Human homolog HC-gp39
• stimulate proliferation of fibroblasts and synovial cells in culture (Recklies et al.2002)
• in vivo is connected with differentiation of peripheral macrophages (Kirkpatick et al. 2007)
• is highly expressed in human body during inflammation (autoimmunity deseases like arthritis, final stage of cancer, viral infections)
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IDGFs probably act as lectinsIDGFs probably act as lectins
• binding domain is slighly modified
• Do they bind chitin or different oligosaccharide motive?
A – IDGF2 binding domainB – Chitinase A binding domainC – Ym1 binding domain (murine homolog of insect IDGFs)Varela et al. 2002
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IDGFs can be specialized to different functionIDGFs can be specialized to different function
Western blot (Bryant and Zurovec)
2 main groups: IDGFs with constitutive and induced expression
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IDGFs 1-3 were identified IDGFs 1-3 were identified as a part of as a part of DrosophilaDrosophila immune response immune response
(DeGregorio et al. 2001, 2002, Irving et al. 2001, Levy et al. 2004,…)
• various MA and proteomic analysis identified IDGF 1-3 as a part of Drosophila immune response in both larvae and adults
• IDGF 1 and 3 are induced with septic injury
• IDGF3 expression is affected with Toll and Imd pathway, however IDGF1 expression is independent on both of them.
• IDGF2 expression is upregulated early after Gram-positive bacterial infection as well as after infection with yeasts.
• All IDGFs were found as a part of haemolymphal clots
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ADG F-D ADG F-C CG 9599
9460 9465 9470
ADG F-E
ADA CG 18005
P-EP
P-EP
3L;74E2
3R;87F12
2R;51C1
3R;85C7
ADG F-A ADG F-B
96409635
4870 4872
M SI
ADA-RELATED GENES IN DROSOPHILA
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The effect of recombinant ADGF-A on the growth and morphology of the Cl.8+ imaginal disc cells
Dying cells Living cells
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It is possible to mimic the effect of ADGFAon cell growth by removal of adenosine
The mutant ADGF has no effect
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Expression of different ADGFs ADGF-A is expressed in embryo, larva and adult (arrows)
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Gene targeting by homologous recombination
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Gene targeting by homologous recombination
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Gene targeting by homologous recombination
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Gene targeting by homologous recombination
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Gene targeting by homologous recombination
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Gene targeting by homologous recombination
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Gene targeting by homologous recombination
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Gene targeting by homologous recombination
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Gene targeting by homologous recombination
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Gene targeting by homologous recombination
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Mutagenesis of the gene ADGF-A17650 17660
3L;74E2
ADGF-A ADGF-B
MSI
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ADGF-A ADGF-B
MSI ADGF-B
MSI ADGF-B
MSI
ADGF-A
3L;74E2
2L;
ADGF-A White
WT
The ADGFA mutant
A rescue construct
Stop kódón
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Tests for somatic mutagenesis inDrosophila based on a recessive mutation in a marker gene (left), or a mutant tumor suppressor gene (right)
y = yellow, yellow bristles instead of black normal ones;wts = warts, warts-like patches of overgrown tissue
Heterozygous wild type cells at four-chromatide stage
Mutagen Mutagen
Tumor cell Normal cell
Clone with yellow bristles
Tumorous clone
Wild type homozygousclone
Surrounding heterozygous tissue, phenotypically normal