m8, E(spl)m8, E(spl), Enhancer of split, E(spl)-m8
transcription factor - bHLH - Hairy/E(spl) class - neurogenic gene - target of Notch pathway - Casein kinase II regulates lateral-inhibition during eye and bristle development by targeting E(spl) repressors
Gene model reviewed during 5.42
Gene model reviewed during 5.48
There is only one protein coding transcript and one polypeptide associated with this gene
179 (aa); 19.7 (kD predicted)
The orange domain and the basic helix-loop-helix motif mediate repression of specific transcriptional activators, such as basic helix-loop-helix protein dimers.
The C-terminal WRPW motif is a transcriptional repression domain necessary for the interaction with Groucho, a transcriptional corepressor recruited to specific target DNA by Hairy-related proteins.
Click to get a list of regulatory features (enhancers, TFBS, etc.) and gene disruptions (point mutations, indels, etc.) within or overlapping Dmel\E(spl)m8-HLH using the Feature Mapper tool.
Comment: reported as procephalic ectoderm primordium
Comment: reported as procephalic ectoderm primordium
Comment: reported as procephalic ectoderm primordium
Comment: reported as head epidermis primordium
Comment: reported as head epidermis primordium
E(spl) genes were found to be differentially expressed during metamorphosis. Unlike other E(spl) genes, E(spl)m8 maintains farily steady levels during metamorphosis.
The distribution of embryonic E(spl) transcripts was compared in D. melanogaster and D. hydei. The patterns of embryonic gene activity were found to be nearly indistinguishable. Mesectodermal expression was observed earlier in melanogaster embryos.
The peak of E(spl) expression during embryogenesis occurs at 2-10 hours. In the late blastoderm, expression is detected in a 2-3 cell-wide stripe on each side of the embryo, in groups of cells over the dorsal half of the poles, and abundantly in a dorsomedian band spanning the anterioposterior axis. During germ band extension, ectodermal expression is detected, and at the extended germ band stage, epidermal expression is abundant. In late stage 11, epidermal expression becomes patchy. At stage 10, the primordia of the supraoesophageal ganglion and the posterior midgut express E(spl). From stage 11 through late stage 12, expression is detected in the entire mesodermal layer. From late stage 11 through stage 14, expression is also detected in the primordia of the stomatogastric nervous system and in the optic lobes.
GBrowse - Visual display of RNA-Seq signalsView Dmel\E(spl)m8-HLH in GBrowse 2
Please Note FlyBase no longer curates genomic clone accessions so this list may not be complete
Please Note This section lists cDNAs and ESTs that fall within the genomic extent of the gene model, which may include cDNAs and ESTs of genes within introns, or of overlapping genes. Please see GBrowse for alignment of the cDNAs and ESTs to the gene model.
For each fully sequenced cDNA the DGRC maintains various forms of the cDNA (e.g tagged or untagged) in several different host vectors for subsequent cloning and expression in Drosophila and Drosophila cell lines.
Source for identity of: E(spl)m8-HLH E(spl)
dsRNA has been made from templates generated with primers directed against this gene. RNAi of E(spl) results in reduced arborization of ddaD and ddaE neurons, alterations in the number of MD neurons, defects in dendrite morphogenesis and reproducible defects in da dendrite development.
Mutant allele fails to complement a QTL affecting male mating behaviour.
Enhancement of the Nspl-1 phenotype by E(spl)1 occurs within the remaining proneural cells, operating primarily at the protein level due to altered protein-protein interactions between E(spl)1 gene product and the proneural proteins.
The distinct expression patterns of genes of the E(spl) complex in imaginal tissues depend to a significant degree on the capacity of their transcriptional cis-regulatory apparatus to respond selectively to direct proneural and Su(H)-mediated activation, often in a subset of the territories and cells in which proneural and Su(H) regulation is occurring.
Candidate gene for quantitative trait (QTL) locus determining bristle number.
The bHLH domains of the gene products encoded by the E(spl)-C and AS-C differ in their ability to form homo- and/or heterodimers. The interactions established through the bHLH link the products of the two complexes in a single interaction network which may function to ensure that a given cell retains the capacity to choose between epidermoblast and neuroblast fates until the cell becomes definitively determined.
Proneural and neurogenic genes control specification and morphogenesis of stomatogastric nerve cell precursors.
Clones mutant for E(spl)-C bHLH-encoding genes or for gro display bristle hyperplasia. The E(spl)-C genes participate in the N signalling pathway. E(spl)-C mutants are epistatic over a gain of function mutant of N and ac-sc mutants are epistatic over E(spl)-C mutants. Expression in Schneider cells demonstrates HLHm5 and E(spl) mediate transcriptional repression of an ac Ecol\CAT reporter gene, gro potentiates this effect.
Mutations show no interaction with high and low selection lines, abdominal and sternopleural bristle numbers are not affected. Results suggest E(spl) is not a candidate for bristle number quantitative trait loci (QTL) in natural populations or is in the same genetic pathway.
The genes of the E(spl) complex mediate only a subset of N activities during imaginal development. Comparisons of mutant phenotypes suggests that the N pathway bifurcates after the activation of Su(H) and that E(spl) activity is not required when the consequence of N function is the transcriptional activation of downstream genes. Transcriptional activation mediated by Su(H) and transcriptional repression mediated by E(spl) could provide greater diversity in the response of individual genes to N activity.
The yeast interaction system has been used to study the E(spl) complex genes, results suggest that E(spl) and gro form a highly interconnected interacting network involved in transcriptional regulation.
Almost all E(spl)-complex bHLH proteins can homo-hetero-dimerise, but not with the same efficiency. All E(spl)-complex bHLH proteins interact with gro protein via their C-terminal domain. E(spl)-complex bHLH proteins interact with proneural proteins, with members of the E(spl) family exhibiting distinct preferences for different proneural proteins.
Proximal upstream region contains multiple specific binding sites for Su(H). Integrity of these sites and Su(H) activity are required not only for normal levels of E(spl) complex gene expression in imaginal disc proneural clusters but also for their transcriptional response to hyperactivity of the N receptor. Su(H) is a direct regulatory link between N receptor activity and the expression of E(spl) complex genes, extending the known lineage of the N cell-cell signaling pathway.
The bristle loss phenotype of H mutants can be suppressed by deleting components of the E(spl)-complex. The degree of suppression depends on both the number and identity of E(spl)-complex transcription units removed.
The E(spl) and ASC complexes interact with each other through the HLH domains of their components.
Transcriptional repression by the h/E(spl) family of bHLH proteins involves two separable mechanisms: repression of specific transcriptional activators, such as sc, through the bHLH and orange domains and repression of other activators via interaction of the C-terminal WRPW motif with corepressors, such as the gro protein.
E(spl) bHLH proteins are turned on in cells which are inhibited from becoming neural by signals from the delaminating neuroblast.
In late stages of development E(spl) BHLH gene products are part of the same signalling pathway and are expressed in cells where N is activated. Loss of N function leads to a reduction in E(spl) bHLH protein expression and the presence of ubiquitous activated N result in high levels of E(spl) bHLH throughout the developing wing disc, effects are independent of genes of the AS-C.
E(spl) protein expression in ectodermal cells commences in the neuroectoderm when neuroblasts have began to delaminate.
Experiments with a chimeric E(spl) derivative with a heterologous transcriptional activation domain support the idea that E(spl) proteins normally act as direct repressors of transcriptional activation of regulatory genes, possibly including the ASC genes.
Gel retardation experiments demonstrate the 5' regulatory region from position -1166 to +87 contains in vitro binding sites for Su(H).
E(spl) complex basic helix loop helix genes inhibit neural fate during the selection of neural precursors, and play a role in restricting the neuronal fate to one of the four progeny cells of the bristle precursor.
The gene products of ac, sc and l(1)sc together with vnd act synergistically to specify the neuroectodermal E(spl) and HLHm5 expression. Negative cross- and autoregulatory interactions of the E(spl) complex contribute, directly or indirectly tot he regulation.
DNaseI footprinting analysis of bacterially expressed E(spl) and HLHm5 demonstrates the gene products can bind as homo- and heterodimers to a sequence in the promoters of the E(spl) and ac genes, called the N-box, which differs slightly from the consensus binding site for other bHLH proteins.
Electrophoretic mobility shift assays demonstrate that E(spl) is directly activated in proneural clusters of the late third-instar wing imaginal disc by protein complexes that include the ac and sc bHLH proteins.
Arrangement and sequence of E(spl)-complex genes in D.melanogaster and D.hydei revealed that the E(spl)-gene, and the structure of complex are highly conserved, suggesting that each individual gene, as well as the organization of the complex, is of functional importance.
Conclusion based on interactions with Df(3R)E(spl)-rv27, as opposed to point mutations in E(spl) locus.
NM1 defines a new class of Notch allele: similarity with and lack of specificity of interaction of N- and NM1 with H, mam, gro and E(spl) suggest that the NM1 effect is due to modification in the intracellular signalling of the activated N receptor.
On basis of cross-hybridization and sequence data the E(spl) HLH genes can be placed into 3 groups. The first includes E(spl) and HLHm5, the second includes HLHm7, HLHm3, HLHmA and HLHmB and the last includes HLHmC.
The embryonic phenotype of neurogenic mutations was examined in most tissues using Ecol\lacZ enhancer trap lines. All alleles examined show defects in many organs from all three germ layers. At least for ectodermally and endodermally derived tissues, neurogenic gene function is primarily involved in interactions among cells that need to acquire or maintain an epithelial phenotype.
Genes of the E(spl) complex act as a functional unit composed of redundant genes which can partially substitute for each other. Eight E(spl)-region genes are required for the development of neurectodermal cells: HLHmδ, HLHmβ, HLHmγ, HLHm3, HLHm5, HLHm7, E(spl) and gro. The E(spl)-region gene m4 may also play a role in this process.
E(spl) acts in the last step of lateral inhibition (de la Concha, Genetics 111: 499--508) and the E(spl) region encodes several HLH proteins (Klambt, EMBO J. 8: 203--210).
In vitro DNA binding assays demonstrate that the basic domain of the E(spl) gene product is necessary for DNA binding. Dominant enhancement of Nspl-1 is caused by the truncation of the E(spl)1 protein in combination with a deletion of a putative regulatory element.
E(spl) is needed for proper mesoderm differentiation prior to the onset of nau expression: mutant alleles cause hypertrophy in nau expressing cells.
The neurogenic phenotype of various embryonic combinations have been studied and include extreme neurogenic embryos, moderate extreme embryos, intermediate neurogenic embryos, weak intermediate neurogenic embryos and weak neurogenic embryos.
Genetic analysis demonstrates that Dl, neu, E(spl), HLHm5, HLHm7 and m4 are functionally related. Spatial distribution of mRNA in neurogenic mutant embryos suggests that some of the functional interactions take place at the transcriptional level.
Ecol\lacZ reporter gene constructs demonstrate that neurogenic loci are required to restrict the number of competent cells that will become sensory mother cells, SMCs.
A synergistic interaction is observed between E(spl) and emc alleles with regard to the ectopic posterior macrochaetae.
E(spl)- cells fail to differentiate chaetae in the step of sensory organ mother cell singularization and/or later during epidermal sublineage specification.
DNA sequence analysis reveals four E box binding sites, for the binding of hetero-oligomeric complexes composed of da or AS-C proteins, in the first 877 bp of the ac upstream region. Electrophoretic mobility shift assays demonstrate that the emc protein can specifically antagonise DNA binding of the da/AS-C complexes in vitro in a dose-dependent manner, h and E(spl) proteins fail to exhibit this inhibitory effect.
E(spl) mutations show no interaction with dx.
Genetic analysis demonstrates that Dl mutations can modify the imaginal phenotypes that result from heterozygosity for E(spl) mutations.
The Notch phenotypic group includes neurogenetic mutations involved in cell communications. Some alleles are embryonic lethal.
N, Dl and E(spl) gene products interact directly during embryonic and imaginal development. Morphogenesis of the ectodermally derived adult eye is sensitive to the combined action of the N, Dl and E(spl) gene products.
A study of the interactions between N, Dl, H and E(spl) suggest that the effects of H, Dl and E(spl) on N are allele specific and occurring at the protein level.
Neural hyperplasia, caused by mutations in E(spl), can be prevented by the presence of another neurogenic mutation.
Molecular and cytogenetic analysis of the E(spl) locus has led to the characterization of a 14kb deletion that affects E(spl) functions.
The genetic organization of the chromosome interval 96F8 has been determined. Results suggest that several genetic functions are related to E(spl) and that the role E(spl) plays in neurogenesis requires the participation of more than one of these genetic functions.
Mutant analysis of E(spl) indicates that low levels of E(spl)+ gene activity result in hyperplasia of both the CNS and PNS, fewer neurons are produced in genotypes where the activity of the locus has increased. Results suggest that E(spl) acts as a genetic switch directing the decision to become either a neural or epidermal progenitor.
Regions of E(spl) cross-hybridize to the opa sequence.
E(spl) mutants display no ventral cuticle and hypertrophy of the central nervous system.
Locus involved in the differentiation of the neural ectoderm into neuroblasts and epidermoblasts. Increased levels of gene product favor epidermal differentiation, whereas decreased levels favor neuronal differentiation. Locus originally identified by the split-enhancing feature of a dominant gain of function mutation. Loss of function mutations are lethals and are described separately. E(spl) causes spl/+ to display a split phenotype and elicits a more extreme phenotype in spl/spl and spl/Y. The spl-enhancing effect of E(spl)1 is suppressed in flies heterozygous for Dl (Shepard et al., 1989). With respect to enhancement, +/+/+ < +/E(spl) < E(spl)/+/+ < E(spl)/E(spl), in accord with expectations from a hypermorphic allele; duplication for E(spl)+ achieved with Dp(3;3)M95A+16. In the absence of spl, E(spl) causes slight roughening of the eyes; furthermore, depending on parental constitution, varying percentages of embryos display defects in central- and peripheral-nervous-system development and irregular cuticular defects. A fraction of these fail to develop; percentages vary from 25% neural hypoplasia and 8% embryonic mortality in crosses between homozygous E(spl) parents to 100% death and 78% neural hypoplasia when both parents are E(spl)/Dp(3;3)M95A+16. Both of these effects are sensitive to maternal genotype. That E(spl) is not simply a hypermorph is indicated by the fact that although heterozygous E(spl)- deletions are viable, hemizygous E(spl) is virtually lethal, especially when the deficiency is maternally inherited. Embryos homozygous for loss-of-function alleles vary in phenotype from weak to very strong neural hyperplasia, with concomitant aplasia of the epidermal sheath. Heterozygotes for stronger hypomorphic alleles may show terminal thickening of wing veins L4 and L5 and may have adventitious vein segments in the posterior wing membrane. Double heterozygotes for E(spl) loss-of-function alleles and either N or Dl are lethal. In the adult, increasing levels of E(spl) function result in increasing levels of split enhancement and in decreased numbers of sensilla as measured by the number of costal bristles on the wing. Conversely, decreased E(spl) function results in larger eyes and more sensilla plus ectopic sensory neurons appearing in the wing blade, especially along the posterior margin. Hemizygosity for E(spl)+ completely suppresses spl. Three doses of E(spl)+ increase the severity of the effects of the absence of function of Dl, reduce the severity of the absence of function of N, neu and mam and are without effect on the phenotype of bib-; conversely, absence of function of E(spl) is not affected by hyperploidy for any of the neurogenic loci or by loss of H function; from this De la Concha et al. infer that E(spl) is positively controlled by N and negatively controlled by H and Dl. Unlike the results using other neurogenic mutants, single vitally stained cells taken from the neurogenic ectoderm of E(spl)- embryos and transplanted into wild-type host embryos fail to give rise to clones containing epidermal cells; only neuronal elements are produced. This observation is interpreted to indicate that the E(spl)+ product serves a receptor rather than a signalling function (Technau and Campos-Ortega, 1987).