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Development of the nervous system depends on signaling centers – specialized cellular populations that produce secreted molecules to regulate neurogenesis in the neighboring neuroepithelium. In some cases, signaling center cells also differentiate to produce key types of neurons. The formation of a signaling center involves its induction, the maintenance of expression of its secreted molecules, and cell differentiation and migration events. How these distinct processes are coordinated during signaling center development remains unknown. By performing studies in mice, we show that Lmx1a acts as a master regulator to orchestrate the formation and function of the cortical hem (CH), a critical signaling center that controls hippocampus development. Lmx1a co-regulates CH induction, its Wnt signaling, and the differentiation and migration of CH-derived Cajal–Retzius neurons. Combining RNAseq, genetic, and rescue experiments, we identified major downstream genes that mediate distinct Lmx1a-dependent processes. Our work revealed that signaling centers in the mammalian brain employ master regulatory genes and established a framework for analyzing signaling center development.

Large size and external development of the chicken embryo have long made it a valuable tool in the study of developmental biology. With the advent of molecular biological techniques, the chick has become a useful system in which to study gene regulation and function. By electroporating DNA or RNA constructs into the developing chicken embryo, genes can be expressed or knocked down in order to analyze in vivo gene function. Similarly, reporter constructs can be used for fate mapping or to examine putative gene regulatory elements. Compared to similar experiments in mouse, chick electroporation has the advantages of being quick, easy and inexpensive. This video demonstrates first how to make a window in the eggshell to manipulate the embryo. Next, the embryo is visualized with a dilute solution of India ink injected below the embryo. A glass needle and pipette are used to inject DNA and Fast Green dye into the developing neural tube, then platinum electrodes are placed parallel to the embryo and short electrical pulses are administered with a pulse generator. Finally, the egg is sealed with tape and placed back into an incubator for further development. Additionally, the video shows proper egg storage and handling and discusses possible causes of embryo loss following electroporation.

Mice homozygous for the dreher (dr) mutation are characterized by pigmentation and skeletal abnormalities and striking behavioral phenotypes, including ataxia, vestibular deficits, and hyperactivity. The ataxia is associated with a cerebellar malformation that is remarkably similar to human Dandy-Walker malformation. Previously, positional cloning identified mutations in LIM homeobox transcription factor 1 alpha gene (Lmx1a) in three dr alleles. Two of these alleles, however, are extinct and unavailable for further analysis. In this article we report a new spontaneous dr allele and describe the Lmx1a mutations in this and six additional dr alleles. Strikingly, deletion null, missense, and frameshift mutations in these alleles all cause similar cerebellar malformations, suggesting that all dr mutations analyzed to date are null alleles.

The cerebellum is the primary motor coordination center of the CNS and is also involved in cognitive processing and sensory discrimination. Multiple cerebellar malformations have been described in humans, however, their developmental and genetic etiologies currently remain largely unknown. In contrast, there is extensive literature describing cerebellar malformations in the mouse. During the past decade, analysis of both spontaneous and gene-targeted neurological mutant mice has provided significant insight into the molecular and cellular mechanisms that regulate cerebellar development. Cerebellar development occurs in several distinct but interconnected steps. These include the establishment of the cerebellar territory along anterior-posterior and dorsal-ventral axes of the embryo, initial specification of the cerebellar cell types, their subsequent proliferation, differentiation and migration, and, finally, the interconnection of the cerebellar circuitry. Our understanding of the basis of these developmental processes is certain to provide insight into the nature of human cerebellar malformations.

Ionizing radiation is a potent lung carcinogen, but the precise molecular damage associated with it is still unknown. In this study we investigated cancer-related molecular abnormalities including K-ras (codon 12) mutation, p16(INK4A) promoter hypermethylation and microsatellite alterations at seven chromosomal regions in successive biopsies obtained from former Chernobyl cleanup workers in comparison with smokers and nonsmokers who have never had radiation exposure. Our results indicate that prolonged persistence of inhaled radioactive particles is associated with appearance of allelic loss at 3p12, 3p14.2 (FHIT), 3p21, 3p22-24 (hMLH1) and 9p21 (p16INK4A) in bronchial epithelium of former Chernobyl clean-up workers. The prevalence of 3p14.2 allelic loss was associated with decreased expression of the FHIT mRNA in their bronchial epithelium in comparison with control group of smokers. During several years of our monitoring samples of epithelium were collected from the same area of bronchial tree. In epithelium exposed to carcinogens (tobacco smoke and/or radioactivity) the total number of molecular abnormalities was significantly higher in dysplasia and in morphologically normal foci progressed later to dysplasia than in these samples which never showed evidence of such progression. Our findings indicate that extensive cancer-related molecular abnormalities sequentially occur in radiation damaged bronchial epithelium of former Chernobyl clean-up workers.