LIM domain-binding protein 1 (Ldb1) is a nuclear cofactor that interacts with LIM homeodomain proteins to form multiprotein complexes that are important for transcription regulation. Ldb1 has been shown to play essential roles in various processes during mouse embryogenesis. To determine the role of Ldb1 in mid- and hindbrain development, we have generated a conditional mutant with a specific deletion of the Ldb1 in the Engrailed-1-expressing region of the developing mid- and hindbrain. Our study showed that the deletion impaired the expression of signaling molecules, such as fibroblast growth factor 8 (FGF8) and Wnt1, in the isthmic organizer and the expression of Shh in the ventral midbrain. The midbrain and the cerebellum were severely reduced in size, and the midbrain dopaminergic (mDA) neurons were missing in the mutant. These defects are identical to the phenotype that has been observed previously in mice with a deletion of the LIM homeodomain gene Lmx1b. Our results thus provide genetic evidence supporting that Ldb1 and Lmx1b function cooperatively to regulate mid- and hindbrain development. In addition, we found that mouse embryonic stem cells lacking Ldb1 failed to generate several types of differentiated neurons, including the mDA neurons, serotonergic neurons, cholinergic neurons, and olfactory bulb neurons, indicating an essential cell-autonomous role for Ldb1 in the development of these neurons.
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References
1.
AngSL. (2006). Transcriptional control of midbrain dopaminergic neuron development. Development, 133:3499–3506.
2.
PrakashN and WurstW. (2006). Development of dopaminergic neurons in the mammalian brain. Cell Mol Life Sci, 63:187–206.
3.
ArenasE, DenhamM and VillaescusaJC. (2015). How to make a midbrain dopaminergic neuron. Development, 142:1918–1936.
4.
BlaessS and AngSL. (2015). Genetic control of midbrain dopaminergic neuron development. Wiley Interdiscip Rev Dev Biol, 4:113–134.
5.
AdamsKA, MaidaJM, GoldenJA and RiddleRD. (2000). The transcription factor Lmx1b maintains Wnt1 expression within the isthmic organizer. Development, 127:1857–1867.
6.
O'HaraFP, BeckE, BarrLK, WongLL, KesslerDS and RiddleRD. (2005). Zebrafish Lmx1b.1 and Lmx1b.2 are required for maintenance of the isthmic organizer. Development, 132:3163–3173.
7.
GuoC, QiuHY, HuangY, ChenH, YangRQ, ChenSD, JohnsonRL, ChenZF and DingYQ. (2007). Lmx1b is essential for Fgf8 and Wnt1 expression in the isthmic organizer during tectum and cerebellum development in mice. Development, 134:317–325.
8.
DengQ, AnderssonE, HedlundE, AlekseenkoZ, CoppolaE, PanmanL, MillonigJH, BrunetJF, EricsonJ and PerlmannT. (2011). Specific and integrated roles of Lmx1a, Lmx1b and Phox2a in ventral midbrain development. Development, 138:3399–3408.
9.
YanCH, LevesqueM, ClaxtonS, JohnsonRL and AngSL. (2011). Lmx1a and lmx1b function cooperatively to regulate proliferation, specification, and differentiation of midbrain dopaminergic progenitors. J Neurosci, 31:12413–12425.
10.
AnderssonE, TryggvasonU, DengQ, FrilingS, AlekseenkoZ, RobertB, PerlmannT and EricsonJ. (2006). Identification of intrinsic determinants of midbrain dopamine neurons. Cell, 124:393–405.
11.
LinW, MetzakopianE, MavromatakisYE, GaoN, BalaskasN, SasakiH, BriscoeJ, WhitsettJA, GouldingM, KaestnerKH and AngSL. (2009). Foxa1 and Foxa2 function both upstream of and cooperatively with Lmx1a and Lmx1b in a feedforward loop promoting mesodiencephalic dopaminergic neuron development. Dev Biol, 333:386–396.
12.
NakataniT, KumaiM, MizuharaE, MinakiY and OnoY. (2010). Lmx1a and Lmx1b cooperate with Foxa2 to coordinate the specification of dopaminergic neurons and control of floor plate cell differentiation in the developing mesencephalon. Dev Biol, 339:101–113.
13.
HobertO and WestphalH. (2000). Functions of LIM-homeobox genes. Trends Genet, 16:75–83.
14.
ZhaoY, MalikN and WestphalH. (2006). Functions of LIM-homeodomain proteins in the development of the nervous system. In: Transcription Factors in the Nervous System. KriegH, ed. Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim, Germany, pp 75–94.
15.
AgulnickAD, TairaM, BreenJJ, TanakaT, DawidIB and WestphalH. (1996). Interactions of the LIM-domain-binding factor Ldb1 with LIM homeodomain proteins. Nature, 384:270–272.
16.
JurataLW, KennyDA and GillGN. (1996). Nuclear LIM interactor, a rhombotin and LIM homeodomain interacting protein, is expressed early in neuronal development. Proc Natl Acad Sci U S A, 93:11693–11698.
17.
BachI, CarriereC, OstendorffHP, AndersenB and RosenfeldMG. (1997). A family of LIM domain-associated cofactors confer transcriptional synergism between LIM and Otx homeodomain proteins. Genes Dev, 11:1370–1380.
18.
MukhopadhyayM, TeufelA, YamashitaT, AgulnickAD, ChenL, DownsKM, SchindlerA, GrinbergA, HuangSP, DorwardD and WestphalH. (2003). Functional ablation of the mouse Ldb1 gene results in severe patterning defects during gastrulation. Development, 130:495–505.
19.
TzchoriI, DayTF, CarolanPJ, ZhaoY, WassifCA, LiL, LewandoskiM, GorivodskyM, LovePE, et al. (2009). LIM homeobox transcription factors integrate signaling events that control three-dimensional limb patterning and growth. Development, 136:1375–1385.
20.
ZhaoY, KwanKM, MaillouxCM, LeeWK, GrinbergA, WurstW, BehringerRR and WestphalH. (2007). LIM-homeodomain proteins Lhx1 and Lhx5, and their cofactor Ldb1, control Purkinje cell differentiation in the developing cerebellum. Proc Natl Acad Sci U S A, 104:13182–13186.
21.
ZhaoY, FlandinP, VogtD, BloodA, HermeszE, WestphalH and RubensteinJL. (2014). Ldb1 is essential for development of Nkx2.1 lineage derived GABAergic and cholinergic neurons in the telencephalon. Dev Biol, 385:94–106.
22.
KimmelRA, TurnbullDH, BlanquetV, WurstW, LoomisCA and JoynerAL. (2000). Two lineage boundaries coordinate vertebrate apical ectodermal ridge formation. Genes Dev, 14:1377–1389.
23.
HwangM, GorivodskyM, KimM, WestphalH and GeumD. (2008). The neuronal differentiation potential of Ldb1-null mutant embryonic stem cells is dependent on extrinsic influences. Stem Cells, 26:1490–1495.
24.
WilkinsonDG, BhattS, ChavrierP, BravoR and CharnayP. (1989). Segment-specific expression of a zinc-finger gene in the developing nervous system of the mouse. Nature, 337:461–464.
25.
GorivodskyM and LonaiP. (2003). Novel roles of Fgfr2 in AER differentiation and positioning of the dorsoventral limb interface. Development, 130:5471–5479.
26.
YingQL, StavridisM, GriffithsD, LiM and SmithA. (2003). Conversion of embryonic stem cells into neuroectodermal precursors in adherent monoculture. Nat Biotechnol, 21:183–186.
27.
LiL, JothiR, CuiK, LeeJY, CohenT, GorivodskyM, TzchoriI, ZhaoY, HayesSM, et al. (2011). Nuclear adaptor Ldb1 regulates a transcriptional program essential for the maintenance of hematopoietic stem cells. Nat Immunol, 12:129–136.
28.
SorianoP. (1999). Generalized lacZ expression with the ROSA26 Cre reporter strain. Nat Genet, 21:70–71.
29.
ChiCL, MartinezS, WurstW and MartinGR. (2003). The isthmic organizer signal FGF8 is required for cell survival in the prospective midbrain and cerebellum. Development, 130:2633–2644.
30.
MeyersEN, LewandoskiM and MartinGR. (1998). An Fgf8 mutant allelic series generated by Cre- and Flp-mediated recombination. Nat Genet, 18:136–141.
31.
McMahonAP and BradleyA. (1990). The Wnt-1 (int-1) proto-oncogene is required for development of a large region of the mouse brain. Cell, 62:1073–1085.
32.
ThomasKR and CapecchiMR. (1990). Targeted disruption of the murine int-1 proto-oncogene resulting in severe abnormalities in midbrain and cerebellar development. Nature, 346:847–850.
33.
McMahonAP, JoynerAL, BradleyA and McMahonJA. (1992). The midbrain-hindbrain phenotype of Wnt-1-/Wnt-1- mice results from stepwise deletion of engrailed-expressing cells by 9.5 days postcoitum. Cell, 69:581–595.
34.
SmidtMP, AsbreukCH, CoxJJ, ChenH, JohnsonRL and BurbachJP. (2000). A second independent pathway for development of mesencephalic dopaminergic neurons requires Lmx1b. Nat Neurosci, 3:337–341.
35.
ThalerJP, LeeSK, JurataLW, GillGN and PfaffSL. (2002). LIM factor Lhx3 contributes to the specification of motor neuron and interneuron identity through cell-type-specific protein-protein interactions. Cell, 110:237–249.
36.
MariniM, BongersEM, CusanoR, Di DucaM, SeriM, KnoersNV and RavazzoloR. (2003). Confirmation of CLIM2/LMX1B interaction by yeast two-hybrid screening and analysis of its involvement in nail-patella syndrome. Int J Mol Med, 12:79–82.
37.
ZhaoY, MarinO, HermeszE, PowellA, FlamesN, PalkovitsM, RubensteinJL and WestphalH. (2003). The LIM-homeobox gene Lhx8 is required for the development of many cholinergic neurons in the mouse forebrain. Proc Natl Acad Sci U S A, 100:9005–9010.
38.
DingYQ, MarklundU, YuanW, YinJ, WegmanL, EricsonJ, DenerisE, JohnsonRL and ChenZF. (2003). Lmx1b is essential for the development of serotonergic neurons. Nat Neurosci, 6:933–938.
39.
KolterudA, AleniusM, CarlssonL and BohmS. (2004). The Lim homeobox gene Lhx2 is required for olfactory sensory neuron identity. Development, 131:5319–5326.
40.
HirotaJ and MombaertsP. (2004). The LIM-homeodomain protein Lhx2 is required for complete development of mouse olfactory sensory neurons. Proc Natl Acad Sci U S A, 101:8751–8755.
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