title: Comprehensive genetic dissection of the hemocyte immune response in the malaria mosquito Anopheles gambiae.
abstract: Reverse genetics in the mosquito Anopheles gambiae by RNAi mediated gene silencing has led in recent years to an advanced understanding of the mosquito immune response against infections with bacteria and malaria parasites. We developed RNAi screens in An. gambiae hemocyte-like cells using a library of double-stranded RNAs targeting 109 genes expressed highly or specifically in mosquito hemocytes to identify novel regulators of the hemocyte immune response. Assays included phagocytosis of bacterial bioparticles, expression of the antimicrobial peptide CEC1, and basal and induced expression of the mosquito complement factor LRIM1. A cell viability screen was also carried out to assess dsRNA cytotoxicity and to identify genes involved in cell growth and survival. Our results identify 22 novel immune regulators, including proteins putatively involved in phagosome assembly and maturation (Ca²⁺ channel, v-ATPase and cyclin-dependent protein kinase), pattern recognition (fibrinogen-domain lectins and Nimrod), immune modulation (peptidase and serine protease homolog), immune signaling (Eiger and LPS-induced factor), cell adhesion and communication (Laminin B1 and Ninjurin) and immune homeostasis (Lipophorin receptor). The development of robust functional cell-based assays paves the way for genome-wide functional screens to study the mosquito immune response to infections with human pathogens.
authors: Division of Cell and Molecular Biology, Department of Life Sciences, Imperial College London, London, United Kingdom. fabrizio.lombardo@uniroma1.it
authors: Fabrizio
authors: F
authors: Lombardo
authors: Yasmeen
authors: Y
authors: Ghani
authors: Fotis C
authors: FC
authors: Kafatos
authors: George K
authors: GK
authors: Christophides
mesh_terms: D000818:Animals; D000852:Anopheles / Q000235:genetics* / Q000276:immunology; D002470:Cell Survival; D002478:Cells, Cultured; D003165:Complement System Proteins / Q000235:genetics* / Q000276:immunology; D004926:Escherichia coli; D015870:Gene Expression; D020868:Gene Silencing; D055106:Genome-Wide Association Study; D006434:Hemocytes / Q000166:cytology / Q000276:immunology* / Q000382:microbiology; D054884:Host-Pathogen Interactions; D007110:Immunity, Active / Q000235:genetics*; D019476:Insect Proteins; D033661:Oocysts / Q000166:cytology / Q000276:immunology; D010587:Phagocytosis / Q000502:physiology; D034622:RNA Interference; D012330:RNA, Double-Stranded / Q000494:pharmacology; D034741:RNA, Small Interfering
chemical_list: D019476:Insect Proteins; D012330:RNA, Double-Stranded; D034741:RNA, Small Interfering; D003165:Complement System Proteins
references: Cell. 2008 May 30;133(5):916-27
references: 18510934
references: Immunol Rev. 2007 Oct;219:8-16
references: 17850478
references: J Immunol Methods. 2009 Mar 15;342(1-2):71-7
references: 19135446
references: Cell Microbiol. 2010 Jan;12(1):1-9
references: 19804484
references: Cell Microbiol. 2006 Sep;8(9):1392-405
references: 16922859
references: EMBO Rep. 2002 Sep;3(9):852-6
references: 12189180
references: Nat Rev Immunol. 2004 Aug;4(8):617-29
references: 15286728
references: Sci Signal. 2011 Oct 25;4(196):rs10
references: 22028469
references: Science. 2009 Apr 10;324(5924):258-61
references: 19264986
references: Science. 2010 Sep 10;329(5997):1353-5
references: 20829487
references: PLoS Biol. 2007 Sep;5(9):e247
references: 17880266
references: BMC Genomics. 2009 Jun 05;10:257
references: 19500340
references: Cell Host Microbe. 2009 May 8;5(5):498-507
references: 19454353
references: Science. 2009 Jul 17;325(5938):340-3
references: 19520911
references: Annu Rev Immunol. 2007;25:697-743
references: 17201680
references: Curr Opin Microbiol. 2008 Jun;11(3):262-70
references: 18539520
references: PLoS Pathog. 2009 Nov;5(11):e1000655
references: 19893628
references: PLoS Pathog. 2009 Aug;5(8):e1000542
references: 19662170
references: Nucleic Acids Res. 2009 Jan;37(Database issue):D583-7
references: 19028744
references: Cell. 2001 Mar 9;104(5):709-18
references: 11257225
references: Proc Natl Acad Sci U S A. 1997 Oct 14;94(21):11508-13
references: 9326640
references: PLoS Pathog. 2008 May 16;4(5):e1000069
references: 18483558
references: Science. 2007 Jun 22;316(5832):1738-43
references: 17588928
references: Immunity. 2006 Oct;25(4):677-85
references: 17045818
references: Cell. 2004 Mar 5;116(5):661-70
references: 15006349
references: Curr Biol. 2007 Apr 3;17(7):649-54
references: 17363253
references: EMBO J. 2002 Jun 17;21(12):3009-18
references: 12065414
references: BMC Dev Biol. 2010 Jun 11;10:65
references: 20540764
references: Proc Natl Acad Sci U S A. 2001 Oct 23;98(22):12590-5
references: 11606746
references: Insect Biochem Mol Biol. 2002 Oct;32(10):1295-309
references: 12225920
references: Dev Cell. 2002 Nov;3(5):711-22
references: 12431377
references: J Insect Physiol. 2009 Feb;55(2):87-103
references: 19071131
references: Curr Biol. 2005 Jul 12;15(13):1185-95
references: 16005290
references: J Biol Chem. 2007 Jul 27;282(30):21884-8
references: 17537726
references: EMBO J. 1999 Feb 15;18(4):959-67
references: 10022838
references: Genes Dev. 2005 Aug 15;19(16):1861-70
references: 16055650
references: Cell Host Microbe. 2008 Jun 12;3(6):364-74
references: 18541213
references: Cell. 2005 Oct 21;123(2):335-46
references: 16239149
references: J Leukoc Biol. 2010 Jul;88(1):57-68
references: 20400677
references: Proc Natl Acad Sci U S A. 2009 Oct 20;106(42):17841-6
references: 19805194
references: Genes Dev. 2006 Jul 15;20(14):1899-910
references: 16815999
references: Nucleic Acids Res. 2001 May 1;29(9):e45
references: 11328886
references: Mol Biol Evol. 2008 Nov;25(11):2337-47
references: 18703524
references: Genome Biol. 2006;7(7):R66
references: 16869968
references: Immunity. 2005 Jul;23(1):65-73
references: 16039580
references: Insect Biochem Mol Biol. 2006 Oct;36(10):769-78
references: 17027843
references: Proc Natl Acad Sci U S A. 2005 Aug 9;102(32):11420-5
references: 16076953
references: Nat Rev Genet. 2006 May;7(5):373-84
references: 16607398
references: Science. 2004 Feb 6;303(5659):832-5
references: 14764878
references: Genome Res. 2001 Dec;11(12):1996-2008
references: 11731489
references: PLoS Pathog. 2007 Oct 26;3(10):1446-58
references: 17967059
references: Proc Natl Acad Sci U S A. 2009 Dec 15;106(50):21270-5
references: 19940242
references: J Biol Chem. 2009 Apr 10;284(15):9835-44
references: 19193639
references: Mol Biol Cell. 2002 Jul;13(7):2448-60
references: 12134082
references: PLoS Biol. 2010 Jul 20;8(7):e1000434
references: 20652016
references: PLoS Pathog. 2006 Oct;2(10):e102
references: 17040126
references: Nucleic Acids Res. 2007 Jan;35(Database issue):D492-7
references: 17135194
references: Immunol Rev. 2004 Apr;198:127-48
references: 15199960
references: J Biol Chem. 2003 Feb 7;278(6):4184-93
references: 12456678
references: Nucleic Acids Res. 2010 Jan;38(Database issue):D448-52
references: 19910367
references: J Biol Chem. 1999 Apr 23;274(17):11727-35
references: 10206988
references: Cell Host Microbe. 2009 Mar 19;5(3):273-84
references: 19286136
references: Science. 2005 Aug 19;309(5738):1248-51
references: 16020693
references: Science. 2002 Oct 4;298(5591):159-65
references: 12364793
references: Proc Natl Acad Sci U S A. 2002 Jun 25;99(13):8814-9
references: 12077297
references: Nat Rev Genet. 2008 Jul;9(7):554-66
references: 18521077
references: PLoS Pathog. 2009 Apr;5(4):e1000366
references: 19343212
references: J Parasitol. 2003 Feb;89(1):62-9
references: 12659304
references: Dev Comp Immunol. 2010 Apr;34(4):387-95
references: 20026176
references: PLoS Pathog. 2006 Jun;2(6):e52
references: 16789837
references: Int J Parasitol. 2007 May;37(6):673-81
references: 17275826
references: PLoS Pathog. 2007 Mar;3(3):e41
references: 17381241