A novel Leishmania infantum nuclear phosphoprotein Lepp12 which stimulates IL1-beta synthesis in THP-1 transfectants.
BACKGROUND
We report cloning and characterization of a novel Leishmania infantum protein which we termed Lepp12, and we examine its possible implication in the interference with intramacrophage signaling pathways.


RESULTS
The protein Lepp12 contains 87 amino acid sequence and exhibits 5 potential phosphorylation sites by protein kinase C (PKC). Recombinant GST-Lepp12 is phosphorylated in vitro by exogenous PKC and by PKC-like activities present in promastigote and in the myelomonocytic THP-1 cell line, indicating that at least one phosphorylation site is functional on the recombinant Lepp12. The natural Lepp12 protein is present in L. infantum promastigotes, as evidenced using specific anti-Lepp12 antibodies produced by immunopurification from acute phase VL patient sera. Interestingly, human patient sera are strongly reactive with GST-Lepp12, demonstrating immunogenic properties of Lepp12 in man, but no immune response to Lepp12 is detectable in experimentally infected animals. When isolated from promastigotes, Lepp12 migrates as two species of apparent MW of 18.3 kDa (major) and 14 kDa (minor), localizes in the nuclear fraction and appears constitutively phosphorylated. Natural Lepp12 is phosphorylable in vitro by both exogenous PKC and PKC-like activity present in THP-1 extracts. The intracellular Lepp12 transfected into THP-1 cells activates these cells to produce IL-1beta and induces an enhancing effect on PMA stimulated IL-1beta synthesis, as demonstrated using GST-Lepp12 transfectants.


CONCLUSIONS
Together these results indicate that Lepp12 represents a substrate for PKC or other PKC-like activities present in the promastigote form and the host cell and therefore may interfere with signal transduction pathways involving PKC.
Fragaki
Konstantina
K
Groupe de Recherche en Immunopathologie de la Leishmaniose (EA 2675), Faculté de Médecine, Nice, France. k.fragaki@afssa.fr
Ferrua
Bernard
B
Mograbi
Baharia
B
Waldispühl
Julie
J
Kubar
Joanna
J
D000818:Animals; D002478:Cells, Cultured; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000032:analysis; D006801:Humans; D007375:Interleukin-1 / Q000096:biosynthesis*; D018314:Leishmania infantum / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000235:genetics* / Q000378:metabolism / Q000494:pharmacology; D010766:Phosphorylation; D011493:Protein Kinase C / Q000378:metabolism; D015800:Protozoan Proteins / Q000235:genetics* / Q000378:metabolism / Q000494:pharmacology; D011994:Recombinant Proteins / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects*
D018076:DNA, Complementary; D007375:Interleukin-1; C481965:Lepp12 protein, Leishmania infantum; D009687:Nuclear Proteins; D015800:Protozoan Proteins; D011994:Recombinant Proteins; D011493:Protein Kinase C