A Hemocyte-like Cell Line Established from the Malaria VectorAnopheles gambiae Expresses Six Prophenoloxidase Genes*
Abstract
Cell lines from the malaria vectorAnopheles gambiae have been established as a tool for the study of the mosquito innate immune system in vitro. Here, we describe the first continuous insect cell line that produces prophenoloxidase (PPO). This cell line (4a-3B) expresses constitutively six PPO genes, three of which are novel (PPO4, PPO5, andPPO6). The PPO genes show distinct temporal expression profiles in the intact mosquito, spanning stages from the embryo to the adult in an overlapping manner. Transient induction of larva-specific PPO genes in blood-fed adult females suggests that the developmental hormone 20-hydroxyecdysone may be involved in PPO gene regulation. Indeed, exposure of 4a-3B cells to 20-hydroxyecdysone in culture results in induction of those PPO genes that are mainly expressed in early developmental stages, and repression of PPO5, which is preferentially expressed at the adult stage. The cell line shows bacteria-induced immune transcripts that encode defensin and Gram-negative bacteria-binding protein, but no induction of PPO transcripts. This cell line most likely derives from a hemocyte lineage, and represents an appropriate in vitro model for the study of the humoral and cellular immune defenses of A. gambiae.
Footnotes
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↵* This work was funded by grants from the John D. and Catherine T. MacArthur Foundation, the Human Frontiers Science Program, and the European Union Training and Mobility of Researchers Network on Insect-Parasite Interactions.The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
The nucleotide sequence(s) reported in this paper has been submitted to the GenBank™/EMBL Data Bank with accession number(s) AJ010193 (PPO4; clone L46), AJ010194 (PPO5; clone A1) andAJ010195 (PPO6; clone A2).
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↵‡ Supported by a fellowship of the Deutsche Forschungsgemeinschaft. To whom correspondence should be addressed. Tel.: 49-6221-387-440; Fax.: 49-6221-387-306; E-mail: hmueller{at}embl-heidelberg.de.
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↵§ Supported by a European Union Training and Mobility of Researchers postdoctoral fellowship.
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↵2 P. Brey, personal communication.
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↵3 H.-M. Müller, unpublished data.
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↵4 C. Blass, unpublished data.
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↵5 M. Gorman, personal communication.
- Abbreviations:
- PPO
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prophenoloxidase
- 20-HOE
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20-hydroxyecdysone
- BAC
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bacterial artificial chromosome
- FCS
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fetal calf serum
- PBS
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phosphate-buffered saline
- PO
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phenoloxidase
- PCR
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polymerase chain reaction
- RT-PCR
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reverse transcription-PCR
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- Received December 15, 1998.
- Revision received February 2, 1999.
- The American Society for Biochemistry and Molecular Biology, Inc.
