A postmitotic role for Isl-class LIM homeodomain proteins in the assignment of visceral spinal motor neuron identity

LIM homeobox genes have a prominent role in the regulation of neuronal subtype identity and distinguish motor neuron subclasses in the embryonic spinal cord. We have investigated the role of Isl-class LIM homeodomain proteins in motor neuron diversification using mouse genetic methods. All spinal motor neuron subtypes initially express both Isl1 and Isl2, but Isl2 is rapidly downregulated by visceral motor neurons. Mouse embryos lacking Isl2 function exhibit defects in the migration and axonal projections of thoracic level motor neurons that appear to reflect a cell-autonomous switch from visceral to somatic motor neuron character. Additional genetic mutations that reduce or eliminate both Isl1 and Isl2 activity result in more pronounced defects in visceral motor neuron generation and erode somatic motor neuron character. Thus, an early phase of high Isl expression and activity in newly generated motor neurons permits the diversification of visceral and somatic motor neuron subtypes in the developing spinal cord.     

Expression of Lhx6 in the adult and developing mouse retina

PurposeTo investigate the expression patterns of LIM Homeobox 6 (Lhx6) in the adult and developing mouse retina.MethodsThe Lhx6-GFP knock-in allele was used to activate constitutive expression of a GFP reporter in Lhx6 expressing cells. Double labeling with GFP and retinal markers in the mouse retina at postnatal day 56 (P56) was performed to identify the cell types expressing Lhx6. To determine the neuronal cell types that express Lhx6, double labeling with GFP and various retinal markers was employed in the differentiating retina at P7 and P15.ResultsGFP + Lhx6 lineage cells were determined in Brn3a + retinal ganglion cells (RGCs), ChAT + amacrine cells (ACs), and Islet-class LIM-homeodomain 1 (Isl1+) ACs in the mouse retina at P56. In the ganglion cell layer (GCL), Lhx6 was expressed in Brn3a + RGCs but not Brn3b + RGCs at P15. Moreover, in the inner nuclear layer (INL), Lhx6 was not expressed in Bhlhb5+ ACs at P15. However, Lhx6 was weakly expressed in Glyt1+ ACs and Pax6+ ACs, and strongly expressed in Isl1+ and ChAT + ACs at P15.ConclusionLhx6 was expressed in RGCs and ACs in both the adult and developing mouse retina.     

Isl1 Directly Controls a Cholinergic Neuronal Identity in the Developing Forebrain and Spinal Cord by Forming Cell Type-Specific Complexes

The establishment of correct neurotransmitter characteristics is an essential step of neuronal fate specification in CNS development. However, very little is known about how a battery of genes involved in the determination of a specific type of chemical-driven neurotransmission is coordinately regulated during vertebrate development. Here, we investigated the gene regulatory networks that specify the cholinergic neuronal fates in the spinal cord and forebrain, specifically, spinal motor neurons (MNs) and forebrain cholinergic neurons (FCNs). Conditional inactivation of Isl1, a LIM homeodomain factor expressed in both differentiating MNs and FCNs, led to a drastic loss of cholinergic neurons in the developing spinal cord and forebrain. We found that Isl1 forms two related, but distinct types of complexes, the Isl1-Lhx3-hexamer in MNs and the Isl1-Lhx8-hexamer in FCNs. Interestingly, our genome-wide ChIP-seq analysis revealed that the Isl1-Lhx3-hexamer binds to a suite of cholinergic pathway genes encoding the core constituents of the cholinergic neurotransmission system, such as acetylcholine synthesizing enzymes and transporters. Consistently, the Isl1-Lhx3-hexamer directly coordinated upregulation of cholinergic pathways genes in embryonic spinal cord. Similarly, in the developing forebrain, the Isl1-Lhx8-hexamer was recruited to the cholinergic gene battery and promoted cholinergic gene expression. Furthermore, the expression of the Isl1-Lhx8-complex enabled the acquisition of cholinergic fate in embryonic stem cell-derived neurons. Together, our studies show a shared molecular mechanism that determines the cholinergic neuronal fate in the spinal cord and forebrain, and uncover an important gene regulatory mechanism that directs a specific neurotransmitter identity in vertebrate CNS development.     

人同源框基因Lhx4 cDNA序列的获得,染色体定位和基因组分析

Lhx4基因是LIM同源框基因家族的一员 ,它在运动神经元的发育过程中发挥着重要的作用 .从人脊髓cDNA文库中筛选到了 1个人源性Lhx4基因的cDNA全长序列 ,它与鼠源性的Lhx4基因的cDNA序列有 92 %同源性 .它的基因被定位在 1号染色体 1q 2 4 .1- 1q 2 4 .3的位置 ,并包含有 6个外显子 .其中同源框结构域由外显子 4和 5表达 ,LIM结构域 1由外显子 2表达 ,LIM结构域2由外显子 3表达 .     

LIM homeobox gene-dependent expression of biogenic amine receptors in restricted regions of the C. elegans nervous system

Biogenic amines regulate a variety of behaviors. Their functions are predominantly mediated through G-protein-coupled 7-transmembrane domain receptors (GPCR), 16 of which are predicted to exist in the genome sequence of the nematode Caenorhabditis elegans. We describe here the expression pattern of several of these aminergic receptors, including two serotonin receptors (ser-1 and ser-4), one tyramine receptor (ser-2), and two dopamine receptors (dop-1 and dop-2). Moreover, we describe distinct but partially overlapping expression patterns of different splice forms of the ser-2 tyramine receptor locus. We find that each of the aminergic receptor genes is expressed in restricted regions of the nervous system and that many of them reveal significant overlap with the expression of regulatory factors of the LIM homeobox (Lhx) gene family. We demonstrate that the expression of several of the biogenic amine receptors is abrogated in specific cell types in Lhx gene mutants, thus establishing a role for these Lhx genes in regulating aspects of neurotransmission. We extend these findings with other cell fate markers and show that the lim-4 Lhx gene is required for several but not all aspects of RID motor neuron differentiation and that the lim-6 Lhx gene is required for specific aspects of RIS interneuron differentiation. We also use aminergic receptor gfp reporter fusions as tools to visualize the anatomy of specific neurons in Lhx mutant backgrounds and find that the development of the elaborate dendritic branching pattern of the PVD harsh touch sensory neuron requires the mec-3 Lhx gene. Lastly, we analyze a mutant allele of the ser-2 tyramine receptor, a target of the ttx-3 Lhx gene in the AIY interneuron class. ser-2 mutants display none of the defects previously shown to be associated with loss of AIY function.     

Cofactor CLIM2 promotes the repressive action of LIM homeodomain transcription factor Lhx2 in the expression of porcine pituitary glycoprotein hormone alpha subunit gene

We have cloned a porcine orthologue of cofactor CLIM2 (Ldb1/NLI) from the porcine pituitary cDNA library by protein-protein interaction with the Yeast Two-Hybrid System using porcine Lhx2 as a bait protein. Porcine CLIM2 shows a high identity (99%) in the dimerization domain, nuclear localization signal and LIM binding domain with those of man and mouse. The expression of CLIM2 gene in the anterior pituitary lobe was detected during the porcine fetal and postnatal period by RT-PCR analysis, suggesting that this protein is constitutively expressing and plays a basic role in the anterior pituitary. Transfection assay to the pituitary tumor derived LbetaT2 cells, and the Chinese hamster ovary cells demonstrated that CLIM2 acts as a corepressor of the porcine Lhx2 function. Interestingly, CLIM2 alone apparently repressed the high level of alphaGSU gene expression in LbetaT2 cells. These data suggest that CLIM2 is a basic factor in the pituitary development and function, and plays the role of repressor to modify the function of Lhx2 on the alphaGSU gene expression.