Distinct roles of Hand2 in initiating polarity and posterior Shh expression during the onset of mouse limb bud development.
The polarization of nascent embryonic fields and the endowment of cells with organizer properties are key to initiation of vertebrate organogenesis. One such event is antero-posterior (AP) polarization of early limb buds and activation of morphogenetic Sonic Hedgehog (SHH) signaling in the posterior mesenchyme, which in turn promotes outgrowth and specifies the pentadactylous autopod. Inactivation of the Hand2 transcriptional regulator from the onset of mouse forelimb bud development disrupts establishment of posterior identity and Shh expression, which results in a skeletal phenotype identical to Shh deficient limb buds. In wild-type limb buds, Hand2 is part of the protein complexes containing Hoxd13, another essential regulator of Shh activation in limb buds. Chromatin immunoprecipitation shows that Hand2-containing chromatin complexes are bound to the far upstream cis-regulatory region (ZRS), which is specifically required for Shh expression in the limb bud. Cell-biochemical studies indicate that Hand2 and Hoxd13 can efficiently transactivate gene expression via the ZRS, while the Gli3 repressor isoform interferes with this positive transcriptional regulation. Indeed, analysis of mouse forelimb buds lacking both Hand2 and Gli3 reveals the complete absence of antero-posterior (AP) polarity along the entire proximo-distal axis and extreme digit polydactyly without AP identities. Our study uncovers essential components of the transcriptional machinery and key interactions that set-up limb bud asymmetry upstream of establishing the SHH signaling limb bud organizer.
Galli
Antonella
A
Developmental Genetics, Department of Biomedicine, University of Basel, Basel, Switzerland. ag2994@columbia.edu
Robay
Dimitri
D
Osterwalder
Marco
M
Bao
Xiaozhong
X
Bénazet
Jean-Denis
JD
Tariq
Muhammad
M
Paro
Renato
R
Mackem
Susan
S
Zeller
Rolf
R
D000818:Animals; D051792:Basic Helix-Loop-Helix Proteins / Q000235:genetics* / Q000378:metabolism; D019521:Body Patterning / Q000235:genetics*; D002843:Chromatin / Q000378:metabolism; D004622:Embryo, Mammalian / Q000378:metabolism; D018507:Gene Expression Regulation, Developmental; D053823:Hedgehog Proteins / Q000235:genetics* / Q000378:metabolism; D018398:Homeodomain Proteins / Q000235:genetics / Q000378:metabolism; D018878:Limb Buds / Q000196:embryology*; D051379:Mice; D008822:Mice, Transgenic; D009154:Mutation; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism
D051792:Basic Helix-Loop-Helix Proteins; D002843:Chromatin; D053823:Hedgehog Proteins; D018398:Homeodomain Proteins; D014157:Transcription Factors; C498766:Hand2 protein, mouse; C084118:Hoxd13 protein, mouse; C465570:Shh protein, mouse
Nature. 2000 Aug 31;406(6799):1005-9
10984056
Nature. 2008 Apr 17;452(7189):882-6
18354396
Dev Cell. 2006 Jan;10(1):93-103
16399081
Development. 2005 Feb;132(4):797-803
15677727
Hum Mol Genet. 2003 Jul 15;12(14):1725-35
12837695
Nat Rev Genet. 2009 Dec;10(12):845-58
19920852
Nature. 2008 May 15;453(7193):401-5
18449196
J Biol Chem. 1999 Jan 8;274(2):1040-9
9873049
Nature. 2005 Jul 14;436(7048):277-81
16015334
Dev Biol. 2007 Apr 1;304(1):9-21
17300775
Genes Dev. 2002 Feb 15;16(4):421-6
11850405
Dev Cell. 2009 Jan;16(1):47-57
19097946
Dev Cell. 2008 Apr;14(4):624-32
18410737
Development. 1999 Jan;126(1):13-21
9834182
Nature. 1982 Apr 8;296(5857):564-6
7070499
Development. 1997 Nov;124(22):4523-36
9409670
Curr Opin Cell Biol. 2006 Dec;18(6):723-9
17049221
Development. 2007 May;134(10):1977-89
17442700
Science. 2002 Oct 25;298(5594):827-30
12215652
Development. 2005 Apr;132(8):1961-70
15790970
Development. 2002 Jul;129(13):3077-88
12070084
Am J Med Genet A. 2006 Jul 1;140(13):1407-13
16688725
Dev Biol. 2001 Aug 15;236(2):421-35
11476582
Nat Genet. 2005 Apr;37(4):373-81
15735646
Nature. 2009 Feb 12;457(7231):818-23
19212399
Nature. 1999 Oct 7;401(6753):598-602
10524628
Nature. 2002 Aug 29;418(6901):979-83
12198547
Cell. 2000 Feb 18;100(4):423-34
10693759
Dev Biol. 2007 Oct 15;310(2):430-41
17714700
Nat Genet. 2000 Dec;26(4):460-3
11101846
Development. 2000 May;127(10):2133-42
10769237
Nature. 1997 Aug 14;388(6643):639-48
9262397
Development. 2009 Mar;136(6):933-42
19211672
Genesis. 2002 Jun;33(2):77-80
12112875
Mech Dev. 1997 Mar;62(2):175-82
9152009
Nat Genet. 1993 Mar;3(3):241-6
8387379
Dev Growth Differ. 2007 Aug;49(6):439-48
17661738
J Biol Chem. 2002 Apr 12;277(15):12604-12
11812799
Science. 2009 Feb 20;323(5917):1050-3
19229034
Development. 2006 Sep;133(17):3419-28
16908629
Development. 1996 Oct;122(10):2997-3011
8898214
Genes Dev. 2008 Oct 1;22(19):2651-63
18832070
Mech Dev. 2001 Jan;100(1):45-58
11118883
Cell. 2004 Aug 20;118(4):517-28
15315763
Development. 2006 Jun;133(11):2263-73
16672333
Development. 2000 Jun;127(12):2573-82
10821756
Nature. 2002 Aug 1;418(6897):501-8
12152071
Int J Dev Biol. 2005;49(4):443-8
15968591
Nature. 2005 Jun 23;435(7045):1113-6
15973411
Nature. 2006 Oct 26;443(7114):985-8
17066034
Mamm Genome. 2004 Jan;15(1):23-34
14727139
Proc Natl Acad Sci U S A. 2006 Mar 21;103(12):4511-5
16537395
Development. 2003 May;130(10):2263-73
12668638
Development. 2004 May;131(10):2339-47
15102708
Cell. 2004 Aug 20;118(4):505-16
15315762
Development. 2000 Jun;127(11):2461-70
10804186
Dev Biol. 2007 Jun 15;306(2):883-93
17467687
Development. 2007 Jan;134(1):85-92
17138667
Methods. 1997 Feb;11(2):205-14
8993033
Science. 2004 Jul 16;305(5682):396-9
15256670
Nat Genet. 1997 Jun;16(2):154-60
9171826
Mech Dev. 2002 Jun;114(1-2):51-9
12175489
Development. 2002 Aug;129(15):3563-74
12117807