title: A forward genetic screen in mice identifies Sema3A(K108N), which binds to neuropilin-1 but cannot signal.
abstract: We have performed a three-generation, forward genetic screen to identify recessive mutations that affect the patterning of the peripheral nervous system. Using this assay, we identified Sema3A(K108N), a novel loss-of-function allele of Sema3A. Class 3 semaphorins, which include Sema3A, are structurally conserved secreted proteins that play critical roles in the development and function of the nervous system. Sema3A(K108N) mutant mice phenocopy Sema3A-null mice, and Sema3A(K108N) protein fails to repel or collapse DRG axons in vitro. K108 is conserved among semaphorins, yet the loss-of-function effects associated with K108N are not the result of impaired expression, secretion, or binding of Sema3A to its high-affinity receptor Neuropilin-1 (Npn-1). Using in silico modeling and mutagenesis of other semaphorin family members, we predict that Sema3A(K108N) interacts poorly with the Npn-1/PlexA holoreceptor and, thus, interferes with its ability to signal at the growth cone. Therefore, through the use of a forward-genetic screen we have identified a novel allele of Sema3A that provides structural insight into the mechanism of Sema3A/Npn-1/PlexinA signaling.
authors.affiliation: The Solomon H. Snyder Department of Neuroscience and Howard Hughes Medical Institute, Johns Hopkins University, Baltimore, Maryland 21205, USA.
authors.forename: Janna
authors.initials: J
authors.lastname: Merte
authors.forename: Qiang
authors.initials: Q
authors.lastname: Wang
authors.forename: Craig W
authors.initials: CW
authors.lastname: Vander Kooi
authors.forename: Sarah
authors.initials: S
authors.lastname: Sarsfield
authors.forename: Daniel J
authors.initials: DJ
authors.lastname: Leahy
authors.forename: Alex L
authors.initials: AL
authors.lastname: Kolodkin
authors.forename: David D
authors.initials: DD
authors.lastname: Ginty
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution / Q000235:genetics*; D000818:Animals; D001216:Asparagine / Q000235:genetics; D002460:Cell Line; D005260:Female; D005808:Genes, Recessive; D005820:Genetic Testing / Q000379:methods; D006801:Humans; D008239:Lysine / Q000235:genetics; D008297:Male; D051379:Mice; D008809:Mice, Inbred C3H; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D008969:Molecular Sequence Data; D039942:Neuropilin-1 / Q000235:genetics* / Q000378:metabolism*; D017354:Point Mutation; D011485:Protein Binding / Q000235:genetics; D040121:Semaphorin-3A / Q000235:genetics* / Q000378:metabolism*; D015398:Signal Transduction / Q000235:genetics*
chemical_list: C488217:Sema3a protein, mouse; D040121:Semaphorin-3A; D039942:Neuropilin-1; D001216:Asparagine; D008239:Lysine
references.citation: Neuron. 2003 Aug 14;39(4):589-98
references.pmid: 12925274
references.citation: Mech Dev. 2000 May;93(1-2):95-104
references.pmid: 10781943
references.citation: Mamm Genome. 2000 Jul;11(7):484-8
references.pmid: 10886010
references.citation: Cell. 1997 Aug 22;90(4):753-62
references.pmid: 9288754
references.citation: Cell. 1999 Oct 1;99(1):59-69
references.pmid: 10520994
references.citation: Cell. 1999 May 28;97(5):551-2
references.pmid: 10367884
references.citation: Neuron. 1997 Sep;19(3):519-30
references.pmid: 9331345
references.citation: Genome Biol. 2006;7(3):211
references.pmid: 16584533
references.citation: Neuron. 1997 Nov;19(5):995-1005
references.pmid: 9390514
references.citation: Hum Mol Genet. 1999;8(10):1955-63
references.pmid: 10469849
references.citation: Science. 2005 Jan 14;307(5707):265-8
references.pmid: 15550623
references.citation: Nucleic Acids Res. 2006 Jul 1;34(Web Server issue):W239-42
references.pmid: 16845001
references.citation: Neuron. 2000 Jan;25(1):29-41
references.pmid: 10707970
references.citation: Proc Natl Acad Sci U S A. 1998 Jun 23;95(13):7485-90
references.pmid: 9636176
references.citation: Annu Rev Cell Dev Biol. 2007;23:263-92
references.pmid: 17539753
references.citation: Genesis. 2000 Apr;26(4):230-3
references.pmid: 10748459
references.citation: J Neurosci. 2005 Dec 14;25(50):11787-95
references.pmid: 16354937
references.citation: Neuron. 2005 Feb 17;45(4):513-23
references.pmid: 15721238
references.citation: Neuron. 1997 Sep;19(3):531-7
references.pmid: 9331346
references.citation: Dev Cell. 2003 Jul;5(1):45-57
references.pmid: 12852851
references.citation: J Biol Chem. 2002 May 17;277(20):18069-76
references.pmid: 11886873
references.citation: Cell Mol Life Sci. 2009 Feb;66(4):649-66
references.pmid: 18953684
references.citation: Neuron. 2001 Oct 25;32(2):249-63
references.pmid: 11683995
references.citation: Neuron. 2000 Jan;25(1):43-56
references.pmid: 10707971
references.citation: Nature. 1996 Oct 10;383(6600):525-8
references.pmid: 8849723
references.citation: Neuron. 1995 May;14(5):949-59
references.pmid: 7748562
references.citation: Nat Protoc. 2007;2(8):2005-11
references.pmid: 17703212