title: Activation of human mast cells by retrocyclin and protegrin highlight their immunomodulatory and antimicrobial properties.
abstract: Preclinical evaluation of Retrocyclins (RC-100, RC-101) and Protegrin-1 (PG-1) antimicrobial peptides (AMPs) is important because of their therapeutic potential against bacterial, fungal and viral infections. Human mast cells (HMCs) play important roles in host defense and wound healing but the abilities of retrocyclins and protegrin-1 to harness these functions have not been investigated. Here, we report that chemically synthesized RC-100 and PG-1 caused calcium mobilization and degranulation in HMCs but these responses were not blocked by an inhibitor of formyl peptide receptor-like 1 (FPRL1), a known receptor for AMPs. However, RC-100 and PG-1 induced degranulation in rat basophilic leukemia (RBL-2H3) cells stably expressing Mas related G protein coupled receptor X2 (MrgX2). Chemical synthesis of these AMPs is prohibitively expensive and post-synthesis modifications (cyclization, disulfide bonds, folding) are inadequate for optimal antimicrobial activity. Indeed, we found that synthetic RC-100, which caused mast cell degranulation via MrgX2, did not display any antimicrobial activity. Green-fluorescent protein (GFP)-tagged RC-101 (analog of RC-100) and GFP-tagged PG-1 purified from transgenic plant chloroplasts killed bacteria and induced mast cell degranulation. Furthermore, GFP-PG1 bound specifically to RBL-2H3 cells expressing MrgX2. These findings suggest that retrocyclins and protegrins activate HMCs independently of FPRL1 but via MrgX2. Harnessing this novel feature of AMPs to activate mast cell's host defense/wound healing properties in addition to their antimicrobial activities expands their clinical potential. Low cost production of AMPs in plants should facilitate their advancement to the clinic overcoming major hurdles in current production systems.
authors.affiliation: Department of Pathology, School of Dental Medicine, University of Pennsylvania, Philadelphia, PA, USA.
authors.forename: Kshitij
authors.initials: K
authors.lastname: Gupta
authors.affiliation: Department of Biochemistry, School of Dental Medicine, University of Pennsylvania, Philadelphia, PA, USA.
authors.forename: Akhil
authors.initials: A
authors.lastname: Kotian
authors.affiliation: Department of Pathology, School of Dental Medicine, University of Pennsylvania, Philadelphia, PA, USA.
authors.forename: Hariharan
authors.initials: H
authors.lastname: Subramanian
authors.affiliation: Department of Biochemistry, School of Dental Medicine, University of Pennsylvania, Philadelphia, PA, USA.
authors.forename: Henry
authors.initials: H
authors.lastname: Daniell
authors.affiliation: Department of Pathology, School of Dental Medicine, University of Pennsylvania, Philadelphia, PA, USA.
authors.forename: Hydar
authors.initials: H
authors.lastname: Ali
mesh_terms: D000818:Animals; D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D023181:Antimicrobial Cationic Peptides / Q000096:biosynthesis / Q000235:genetics / Q000494:pharmacology*; D001419:Bacteria / Q000187:drug effects / Q000254:growth & development; D020013:Calcium Signaling / Q000187:drug effects; D015550:Cell Degranulation / Q000187:drug effects*; D045744:Cell Line, Tumor; D023082:Defensins / Q000096:biosynthesis / Q000235:genetics / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D006801:Humans; D007155:Immunologic Factors / Q000096:biosynthesis / Q000235:genetics / Q000494:pharmacology*; D008407:Mast Cells / Q000187:drug effects* / Q000276:immunology / Q000378:metabolism; D050296:Microbial Viability / Q000187:drug effects; D009419:Nerve Tissue Proteins / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D030821:Plants, Genetically Modified / Q000235:genetics / Q000378:metabolism; D051381:Rats; D043562:Receptors, G-Protein-Coupled / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D018013:Receptors, Neuropeptide / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000494:pharmacology; D015398:Signal Transduction / Q000187:drug effects; D013997:Time Factors; D014162:Transfection
chemical_list: D000900:Anti-Bacterial Agents; D023181:Antimicrobial Cationic Peptides; D023082:Defensins; D007155:Immunologic Factors; C480039:MRGPRX2 protein, human; D009419:Nerve Tissue Proteins; D043562:Receptors, G-Protein-Coupled; D018013:Receptors, Neuropeptide; D011993:Recombinant Fusion Proteins; C094880:protegrin-1; C450142:theta-defensin
keywords: Immune response; Immunity; Immunology and Microbiology Section; MrgX2; antimicrobial peptides; chloroplast; mast cells; protegrin; retrocyclin