BACKGROUND: Leishmania major infection induces robust interleukin-12 (IL12) production in human dendritic cells (hDC), ultimately resulting in Th1-mediated immunity and clinical resolution. The surface of Leishmania parasites is covered in a dense glycocalyx consisting of primarily lipophosphoglycan (LPG) and other phosphoglycan-containing molecules (PGs), making these glycoconjugates the likely pathogen-associated molecular patterns (PAMPS) responsible for IL12 induction.
METHODOLOGY/PRINCIPAL FINDINGS: Here we explored the role of parasite glycoconjugates on the hDC IL12 response by generating L. major Friedlin V1 mutants defective in LPG alone, (FV1 lpg1-), or generally deficient for all PGs, (FV1 lpg2-). Infection with metacyclic, infective stage, L. major or purified LPG induced high levels of IL12B subunit gene transcripts in hDCs, which was abrogated with FV1 lpg1- infections. In contrast, hDC infections with FV1 lpg2- displayed increased IL12B expression, suggesting other PG-related/LPG2 dependent molecules may act to dampen the immune response. Global transcriptional profiling comparing WT, FV1 lpg1-, FV1 lpg2- infections revealed that FV1 lpg1- mutants entered hDCs in a silent fashion as indicated by repression of gene expression. Transcription factor binding site analysis suggests that LPG recognition by hDCs induces IL-12 in a signaling cascade resulting in Nuclear Factor κ B (NFκB) and Interferon Regulatory Factor (IRF) mediated transcription.
CONCLUSIONS/SIGNIFICANCE: These data suggest that L. major LPG is a major PAMP recognized by hDC to induce IL12-mediated protective immunity and that there is a complex interplay between PG-baring Leishmania surface glycoconjugates that result in modulation of host cellular IL12.
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This work was supported by National Institutes of of Allergy and Infectious Diseases grants R01AI056242 (MAM) and NIH RO1AI31078 (SMB, SJT). MAF was a fellow of the Chemistry-Biochemistry-Biology Interface (CBBI) Program at the University of Notre Dame, supported by training grant T32GM075762 from the National Institute of General Medical Sciences. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
Favila, Michelle A.; Geraci, Nicholas S.; Jayakumar, Asha; Hickerson, Suzanne; Mostrom, Janet; Turco, Salvatore J.; Beverley, Stephen M.; and McDowell, Mary Ann, "Differential Impact of LPG-and PG-Deficient Leishmania major Mutants on the Immune Response of Human Dendritic Cells" (2015). Molecular and Cellular Biochemistry Faculty Publications. 83.
S1 Fig. Confirmation of <em>L. major</em> FV1 <em>lpg1−</em> mutant.
journal.pntd.0004238.s002.EPS (946 kB)
S1 Fig. Confirmation of <em>L. major</em> FV1 <em>lpg2−</em> mutant.
journal.pntd.0004238.s003.EPS (1545 kB)
S3 Fig. Generation of <em>L. major</em> FV1 LPG null mutant (FV1 <em>lpg1−</em>) and PG null mutant (FV1 <em>lpg2−</em>) mutant.
journal.pntd.0004238.s004.EPS (1754 kB)
S4 Fig. Validation of microarray expression by qRT-PCR.
journal.pntd.0004238.s005.EPS (593 kB)
S5 Fig. Purified LPG stimulates <em>IL12B</em> expression in hDCs.
journal.pntd.0004238.s006.EPS (1023 kB)
S6 Fig. <em>L. major</em> LV39c5 induced hDC IL12 responses do not differ between LV39c5 mutants.
journal.pntd.0004238.s007.DOCX (15 kB)
S1 Table. Primers used for molecular generation of <em>L. major</em> FV1 mutants.
journal.pntd.0004238.s008.DOCX (15 kB)
S2 Table. Human Primer Sequences for qRT-PCR analysis.