Maternal effects of zygotic mutants affecting early neurogenesis inDrosophila

Summary The size of the neurogenic region ofDrosophila melanogaster is under the control of several genes of zygotic expression. Lack of function from any of those genes produces an increase of the size of the neurogenic region at the expense of the epidermal anlage. However, differences exist in the extent of neuralisation achieved by each of the genetic loci upon mutation. The present results show that in the case ofN andmam phenotype differences are due to different contributions of maternal gene expression. This could be shown by studying the phenotype which appeared in mutant embryos when the oocytes developed from homozygous mutant precursor cells. Clones of mutant cells were induced in the germ line of females heterozygous for the neurogenic mutationin trans over germ line dependent, dominant female sterile mutations. After removing maternal information the phenotype ofN andmam mutants became identical in both cases. Furthermore maternal information fromN ⁺ was found to be necessary for viability of the wildtype.     

Molecular organization of master mind, a neurogenic gene of Drosophila melanogaster

Summary The gene master mind (mam) is located in bands 50C23-D1 of the second chromosome of Drosophila melanogaster. mam is one of the neurogenic genes, whose function is necessary for a normal segregation of neural and epidermal lineages during embryonic development. Loss of function of any of the neurogenic genes results in a mis-routeing into neurogenesis of cells that normally would have given rise to epidermis. We describe here the molecular cloning of 198 kb of genomic DNA containing the mam gene. Ten different mam mutations (point mutants and chromosomal aberrations) have been mapped within 45 kb of the genomic walk. One of the mutations, an insertion of a P-element, was originally recovered from a dysgenic cross. Four different wild-type revertants of this mutation were characterized at the molecular level and, although modifications of the insertions were found, in no case was the transposon completely excised. An unusually high number of the repetitive opa sequence, and of an additional previously unknown element, which we have called N repeat, are scattered throughout the 45 kb where the mam mutations map. The functional significance of these repeats is unknown.     

Regulatory interactions during early neurogenesis in Drosophila

During neurogenesis in Drosophila, ectodermal cells are endowed with the capacity to become neuronal precursors. Following their selection, these cells initiate neuronal lineage development and differentiation. The processes of neuronal precursor specification and neuronal lineage development require the activities of several groups of genes functioning in a complex, hierarchical regulatory network. Whereas the proneural genes promote neurogenic potential, neurogenic genes restrict the acquisition of this identity to a subset of ectodermal cells. Following their selection, these cells express the pan neural neuronal precursor genes and a set of neuronal lineage identity genes. While lineage identity genes allow the various lineages to acquire specific identities, neuronal precursor genes presumably regulate functional and developmental characteristics common to all neuronal precursor cells. © 1996 Wiley-Liss, Inc.     

A model for the correlation of mutation rate with GC content and the origin of GC-rich isochores

Based on the biochemical kinetics of DNA replication and mutagenesis, including misincorporation and correction, a model has been developed for studying the relationships among the mutation rate (u), the G + C content of the sequence (f), and the G + C proportion in the nucleotide precursor pool (N). Also a measure for the next-nucleotide effect, called the maximum capacity of the next-nucleotide effect (MC), has been proposed. Under the normal physiological conditions of mammalian germ cells, our results indicate: (1) the equilibrium G + C content in a sequence is approximately equal to the G + C proportion in the nucleotide precursor pool, i.e., f ≈ N, which is independent of the next-nucleotide effect; (2) an inverted-V-shaped distribution of mutation rates with respect to G + C contents is predicted, when the next-nucleotide effect is week, i.e., MC ≈ 1; (3) the distribution becomes flatter (i.e., inverted-U-shaped) as MC increases, but the peak at 50% GC is still observed when MC < 2; and (4) the peak disappears when MC > 2.8, that is, when the next-nucleotide effect becomes strong. Our results suggest that changes in the relative concentrations of nucleotide precursors can cause variations among genes both in mutation rate and in G + C content and that compositional isochores (DNA segments with a homogeneous G + C content) can arise in a genome due to differences in replication times of DNA segments. Correspondence to: W.-H. Li     

The peripheral nervous system of mutants of early neurogenesis inDrosophila melanogaster

Summary Mutations previously known to affect early neurogenesis inDrosophila melanogaster have been found also to affect the development of the peripheral nervous system. Anti-HRP antibody staining has shown that larval epidermal sensilla of homozygous mutant embryos occur in increased numbers, which depend on the allele considered. This increase is apparently due to the development into sensory organs of cells which in the wild-type would have developed as non-sensory epidermis. Thus, neurogenic genes act whenever developing cells have to decide between neurogenic and epidermogenic fates, both in central and peripheral nervous systems. Different regions of the ectodermal germ layer are distinguished with respect to their neurogenic abilities.     

Adult neurogenesis in the crayfish brain: Proliferation,migration, and possible origin of precursor cells

The birth of new neurons and their incorporation into functional circuits in the adult brain is a characteristic of many vertebrate and invertebrate organisms, including decapod crustaceans. Precursor cells maintaining life‐long proliferation in the brains of crayfish (Procambarus clarkii, Cherax destructor) and clawed lobsters (Homarus americanus) reside within a specialized niche on the ventral surface of the brain; their daughters migrate to two proliferation zones along a stream formed by processes of the niche precursors. Here they divide again, finally producing interneurons in the olfactory pathway. The present studies in P. clarkii explore (1) differential proliferative activity among the niche precursor cells with growth and aging, (2) morphological characteristics of cells in the niche and migratory streams, and (3) aspects of the cell cycle in this lineage. Morphologically symmetrical divisions of neuronal precursor cells were observed in the niche near where the migratory streams emerge, as well as in the streams and proliferation zones. The nuclei of migrating cells elongate and undergo shape changes consistent with nucleokinetic movement. LIS1, a highly conserved dynein‐binding protein, is expressed in cells in the migratory stream and neurogenic niche, implicating this protein in the translocation of crustacean brain neuronal precursor cells. Symmetrical divisions of the niche precursors and migration of both daughters raised the question of how the niche precursor pool is replenished. We present here preliminary evidence for an association between vascular cells and the niche precursors, which may relate to the life‐long growth and maintenance of the crustacean neurogenic niche. © 2009 Wiley Periodicals, Inc. Develop Neurobiol, 2009     

The homeobox leucine zipper gene Homez plays a role in Xenopus laevis neurogenesis

The Homez gene encodes a protein with three atypical homeodomains and two leucine zipper motifs of unknown function. Here we show that during neurula stages, Xenopus Homez is broadly expressed throughout the neural plate, the strongest expression being detected in the domains where primary neurons arise. At later stages, Homez is maintained throughout the central nervous system in differentiating progenitors. In accordance with this expression, Homez is positively regulated by neural inducers and by Ngnr1 and negatively by Notch signaling. Interference with Homez function in embryos by injection of an antisense morpholino oligonucleotide results in the specific disruption of the expression of late neuronal markers, without affecting the expression of earlier neuronal and early neurectodermal markers. Consistent with this finding, Homez inhibition also interferes with the expression of late neuronal markers in Ngnr1 overexpressing animal cap explants and in Notch inhibited embryos. In gain of function experiments, Homez inhibits the expression of late neuronal markers but has no effect on earlier ones. These data suggest a role for Homez in neuronal development downstream of proneural/neurogenic genes.     

Location of the replication origin in the 9-kb repeat size class of rDNA in pea (Pisum sativum)

The replication origin of the 9-kb rDNA repeat size class of pea (Pisum sativum cv. Alaska) was identified by benzoylated naphthoylated DEAE-cellulose column chromatography and Southern blotting procedures. The origin is located at or near a 0.19-kb EcoR I fragment in the non-transcribed spacer region between the 25S and 18S rRNA genes. Identification of the origin was based on three criteria: (i) an enrichment of the 0.19-kb fragment in replicating rDNA from asynchronously dividing root meristematic cells, (ii) the scarcity of the 0.19-kb fragment in rDNA from non-dividing carbohydrate starved cells, and (iii) a 60-min periodic enrichment of the 0.19-kb fragment in replicating rDNA that temporally coincides with the sequential initiation of replication of replicon families in synchronized pea root cells.     

Eya1 and Six1 promote neurogenesis in the cranial placodes in a SoxB1-dependent fashion

Genes of the Eya family and of the Six1/2 subfamily are expressed throughout development of vertebrate cranial placodes and are required for their differentiation into ganglia and sense organs. How they regulate placodal neurogenesis, however, remains unclear. Through loss of function studies in Xenopus we show that Eya1 and Six1 are required for neuronal differentiation in all neurogenic placodes. The effects of overexpression of Eya1 or Six1 are dose dependent. At higher levels, Eya1 and Six1 expand the expression of SoxB1 genes (Sox2, Sox3), maintain cells in a proliferative state and block expression of neuronal determination and differentiation genes. At lower levels, Eya1 and Six1 promote neuronal differentiation, acting downstream of and/or parallel to Ngnr1. Our findings suggest that Eya1 and Six1 are required for both the regulation of placodal neuronal progenitor proliferation, through their effects on SoxB1 expression, and subsequent neuronal differentiation.     

Drive-selection equilibrium: Homopolymer evolution in the Drosophila gene mastermind

Interspecific sequence comparison of the highly repetitive Drosophila gene mastermind (mam) reveals extensive length variation in homopolymer domains. The length variation in homopolymers is due to nucleotide misalignment in the underlying triplet repeats, which can lead to slippage mutations during DNA replication or repair. In mam, the length variation in repetitive regions appears to be balanced by natural selection acting to maintain the distance between two highly conserved charge clusters. Here we report a statistical test of the null hypothesis that the similarity in the amino acid distance between the charge clusters of each species arose by chance. The results suggest that at mam there is a juxtaposition of length variability due to molecular drive and length conservation maintained by natural selection. The analysis of mam allows the extension of current theories of drive-selection interaction to encompass homopolymers. Our model of drive-selection equilibrium suggests that the physical flexibility, length variability, and abundance of homopolymer domains provide an important source of genetic variation for natural populations.Correspondence to: S.J. Newfeld 1072     

Glycosylation-related genes are variably expressed depending on the differentiation state of a bioaminergic neuronal cell line: implication for the cellular prion protein

A striking feature of the cellular prion protein (PrP^C) is the heterogeneity of its glycoforms, whose contribution to PrP^C function has yet to be defined. Using the 1C11 neuronal bioaminergic differentiation model and a glycomics approach, we show here a correlation between differential PrP^C N-glycosylations in 1C11^5-HT serotonergic and 1C11^NE noradrenergic cells compared to their 1C11 precursor cells and a variation of the glycogenome expression status in these cells. In particular, expression of genes involved in N-glycan synthesis or in the modeling of chondroitin and heparan sulfate proteoglycans appeared to be modulated. Our results highlight that, the expression of glycosylation-related genes is regulated during bioaminergic neuronal differentiation, consistent with a participation of glycoconjugates in neuronal development and plasticity. A neuronal regulation of glycosylation processes may have direct implications on some neurospecific functions of PrP^C and may participate in specific brain targeting of prion strains. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

The big brain gene of Drosophila functions to control the number of neuronal precursors in the peripheral nervous system.

big brain (bib) is one of the six known zygotic neurogenic genes involved in the decision of an ectodermal cell to take on the neurogenic or the epidermogenic cell fate. Previous studies suggest that bib functions in a pathway separate from the one involving Notch and other known neurogenic genes. For a better understanding of the bib function, it is essential first to characterize the mutant phenotype in detail. Our mutant analyses show that loss of bib function approximately doubles the number of neuronal precursors and their progeny cells in the embryonic peripheral nervous system. Mosaic studies reveal a hypertrophy of sensory bristles in bib mutant patches in adult flies. Our observations are compatible with a function of bib in specifying neuronal precursors of both the embryonic and adult sensory nervous system. This is in contrast to the function of Notch, which continues to be required at multiple stages of neural development subsequent to this initial determination event.     

scabrous Modifies Epithelial Cell Adhesion and Extends the Range of Lateral Signalling during Development of the Spaced Bristle Pattern in Drosophila

The role of scabrous (sca) in the evenly spaced bristle pattern of Drosophila is explored. Loss-of-function of sca results in development of an excess of bristles. Segregation of alternately spaced bristle precursors and epidermal cells from a group of equipotential cells relies on lateral inhibition mediated by Notch and Delta (Dl). In this process, presumptive bristle precursors inhibit the neural fate of neighbouring cells, causing them to adopt the epidermal fate. We show that Dl, a membrane-bound ligand for Notch, can inhibit adjacent cells, in direct contact with the precursor, in the absence of Sca. In contrast, inhibition of cells not adjacent to the precursor requires, in addition, Sca, a secreted molecule with a fibrinogen-related domain. Over-expression of Sca in a wild-type background, leads to increased spacing between bristles, suggesting that the range of signalling has been increased. scabrous acts nonautonomously, and we present evidence that, during bristle precursor segregation, Sca is required to maintain the normal adhesive properties of epithelial cells. The possible effects of such changes on the range of signalling are discussed. We also show that the sensory organ precursors extend numerous fine cytoplasmic extensions bearing Dl molecules, and speculate on a possible role for these structures during signalling.     

Sensillum development in the absence of cell division: the sensillum phenotype of the Drosophila mutant string

We have investigated sensillum development in Drosophila embryos homozygous for mutations in the locus string (stg). In these embryos, cell division is blocked following blastoderm formation. This permits a study of the differentiative fate of undivided precursor cells, in particular those giving rise to the larval sensory organs (sensilla). Of the different cell fates normally represented in the sensilla (i.e., sensory neuron, thecogen cell, trichogen cell, tormogen cell, glia cell), only the phenotype of sensory neurons is expressed morphologically in stg embryos, suggesting that the neuronal fate predominates over the fates of the nonneuronal accessory cells. Consistent with this finding, the P element-lacZ insertion A1-2nd-29, which is a marker for trichogen and tormogen cells in the wild-type embryo, is not expressed in the body wall of the stg embryo. Some sensillum precursor cells appear to express a mixed fate in stg mutants: They express antigens (recognized by the monoclonal antibodies 22C10 and 21A6) which in the wild-type appear in separate cells (sensory neurons and thecogen cell, respectively). The differentiation of undivided cells in stg embryos is not restricted to the peripheral nervous system; in all types of tissues analyzed in this study (e.g., epidermis, intestine, muscle, CNS), precursor cells express characteristics normally exhibited by their progeny.     

Restrictions of developmental capacities in the dorsal root ganglia during the course of development

By grafting ganglia from embryonic quails into the neural crest migration pathway of 2-day chick embryos, it was previously demonstrated that all type of ganglia possess more developmental potentialities than those normally expressed in the normal course of development. Namely autonomic neurones with catecholamine and adrenomedullary cells can be obtained from grafted spinal ganglia. The latter also yield sensory neurons to the host dorsal root ganglia (DRG) but only if they are taken from the donor before 8 days of incubation. In the present article we show that the capacity to differentiate sensory neurons in back-transplantation experiments can be correlated with the presence in the donor DRG of cycling neuronal precursors. Once all the neurons have been withdrawn from the cell cycle - an event which occurs first in the mediodorsal and then in the lateroventral area of the ganglion - the DRG cell population gives rise exclusively to autonomic ganglion cells in the host. It is concluded that in the conditions of the back-transplantation experiments, the postmitotic neurons contained in the donor ganglion do not survive. Therefore, the neurons and paraganglion cells which differentiate in the host arise from still undifferentiated precursor cells. This indicates that besides sensory neuron precursors the embryonic DRG cell population also contains precursor cells for the autonomic differentiation pathway.     

Altered neuronal lineages in the facial ganglia of Hoxa2 mutant mice

Neurons of cranial sensory ganglia are derived from the neural crest and ectodermal placodes, but the mechanisms that control the relative contributions of each are not understood. Crest cells of the second branchial arch generate few facial ganglion neurons and no vestibuloacoustic ganglion neurons, but crest cells in other branchial arches generate many sensory neurons. Here we report that the facial ganglia of Hoxa2 mutant mice contain a large population of crest-derived neurons, suggesting that Hoxa2 normally represses the neurogenic potential of second arch crest cells. This may represent an anterior transformation of second arch neural crest cells toward a fate resembling that of first arch neural crest cells, which normally do not express Hoxa2 or any other Hox gene. We additionally found that overexpressing Hoxa2 in cultures of P19 embryonal carcinoma cells reduced the frequency of spontaneous neuronal differentiation, but only in the presence of cotransfected Pbx and Meis Hox cofactors. Finally, expression of Hoxa2 and the cofactors in chick neural crest cells populating the trigeminal ganglion also reduced the frequency of neurogenesis in the intact embryo. These data suggest an unanticipated role for Hox genes in controlling the neurogenic potential of at least some cranial neural crest cells.