Scute Genes Research Articles

Juan Modolell (1937-2023).

Juan Modolell (1937-2023).

The bristle pattern development in Drosophila melanogaster: the prepattern and achaete-scute genes

The external drosophila mechanoreceptors, residing on the head and body of imago, are represented by bristles of different sizes (macrochaetes and microchaetes). Macrochaetes are arranged in the species-specifc bristle pattern, where each of them is strictly positioned. The bristle pattern is formed starting from its prototype (prepattern) in the imaginal disc. The position specifcity of future mechanoreceptors is determined by local expression of two proneural genes, achaete ( ac ) and scute ( sc ) belonging to the AS-C complex, in response to the action of certain factors, referred to as prepattern factors, nonuniformly distributed in the ectoderm of imaginal discs. The topography of their total distribution defnes the bristle prepattern. Thus, the full-fledged adult bristle pattern is the result of interaction of two systems – the prepattern and the system responding to prepattern, i. e., the achaete and scute genes. A considerable volume of miscellaneous experimental data related to various aspects in development of the bristle pattern has been so far accumulated; however, any formalized and detailed representation of the molecular genetic interaction of the prepattern factors with both each other and the achaete-scute genes is yet absent. This review systematizes the available data on the regular patterns of this interaction and shows that local expression of these genes is determined by hierarchical two-level control system comprising both direct and indirect regulators of their activities. A generalized scheme of the system containing the functional interactions of its components is proposed. The structural organization and principles of operation of the hierarchical molecular genetic system enabling the local expression of ASC genes and the resulting formation of ordered bristle pattern are described.

Analysis of the neurogenesis: Prepattern gene network controlling first stage of bristle pattern development in Drosophila melanogaster

The insect chitinous cover is not able to respond to stimuli and specialized receptors serve for them as environmental signal receivers. The drosophila perceives tactile stimuli by external sensor organs (microand macrochaetae) located on the head and back (notum). The microchaetae are numerous and generate regular rows oriented along the body. The number of macrochaetae is relatively small, and their location on the head and notum is strictly determined. Macrochaetae function as mechanoreceptors maintaining the balance in flight for a fly. The set of macrochaetae is called a bristle pattern. The full bristle pattern in the adult Drosophila melanogaster develops as a result of a multistage process. Its fundamental stage consists in the creation of a prototype of the future location of macrochaeta (prepattern presented by proneural clusters). The proneural clusters are separated from the mass of the wings’ imaginal disc cells at the stage of the third instar larvae and early prepupa under the influence of the prepattern factors identified with the transcription factors that directly regulate the expression of its target genes in the appropriate regions. The results of the reconstruction and analysis of the gene network, which provides the process of the prepattern formation, are for the first time given in the article; the principles of its organization and functioning are considered. The network includes 80 objects connected by 109 regulatory interactions. The proneural ASC proteins encoded by the achaete and scute genes, as well as the Decapentaplegic (Dpp) and Wingless (Wg) proteins, are the key objects of the network demonstrating the largest connectivity with its other components. The network structure is characterized by a hierarchic organization and has at least three management levels. The net functioning as an ensemble of the genes is achieved in general by the coordinated work of the regulatory circuits performing both the intra- and interlevel control for the gene activity. The resulting network effect consists in the activation of proneural AS-C complex genes, whose expression distinguishes the proneural cluster cells from the surrounding ectoderm cells.

Analysis of the Neurogenesis: Prepattern gene network сontrolling the first stage in Drosophila melanogaster bristle pattern development

The external insect chitinous skeleton is unable to respond to stimuli; the external signals are received by specialized receptors. Drosophila perceives the tactile stimuli by its external sensory organs, the microchaetes and macrochaetes residing on the head and back (notum). The microchaetes (hairs) are numerous and arranged in perfect rows along the body. The macrochaetes (bristles) are rather few and are strictly positioned on the head and notum, being referred to as bristle pattern. Bristles act as mechanoreceptors, providing balance for flying drosophila. The proper bristle pattern of an adult fly develops through several stages. Its basic stage is formation of prepattern for the future bristles, represented by proneural clusters. The proneural clusters separate from the ectodermal cells in imaginal discs in the third instar larvae and early prepupae. They are induced by prepattern factors, identified with the transcription factors driving expression of their target genes in certain disc regions. Reconstruction of the gene network controlling prepattern development and its analysis are for the first time described as well as the principles underlying arrangement and function of this network. The hierarchical structure of the network, its key components, and regulatory circuits are identified. The network comprises 80 entities interconnected via 109 regulatory interactions. The key objects of the network, displaying the greatest connectivity with its other components, are the ASC proneural proteins encoded by the achaete and scute genes, and the proteins Decapentaplegic (Dpp) and Wingless (Wg). The structure of the network is hierarchical and has at least three control levels. The network acts as a gene ensemble owing to coordinated functioning of the regulatory circuits controlling activities of the corresponding genes both within and between the levels. The resulting effect of the network operation consists in activation of the AS-C, proneural genes, the expression of which distinguishes the cells of proneural cluster from the surrounding ectodermal cells.

Transcriptional interactions between the pannier isoforms and the cofactor U-shaped during neural development in Drosophila

The pannier ( pnr) gene of Drosophila melanogaster encodes two isoforms that belong to the family of GATA transcription factors. The isoforms share an expression domain in the wing discs where they exhibit distinct functions during regulation of the proneural achaete/ scute ( ac/ sc) genes. We previously identified two regions in the pnr locus that drive reporter expression in transgenic lines in patterns that recapitulate the essential features of expression of the two isoforms. Here, we identify promoter regions driving isoform expression, showing that pnr-α regulatory sequences are close to the transcription start site while pnr-β expression requires functional interactions between proximal and distal regulatory elements. We find that the promoter domains necessary for reporter expression also mediate autoregulation of Pnr-β and repression of pnr-α by Pnr-β. The cofactor U-shaped (Ush), which is known to down-regulate the function of Pnr during thorax patterning postranscriptionally, in addition represses pnr-β required for ac/ sc activation. Moreover, Ush negatively regulates its own expression, while the pnr isoforms positively regulate ush. Our study uncovers complex transcriptional interactions between the pnr isoforms and the cofactor Ush that may be important for regulation of proneural expression and thorax patterning.

HASH1 expression is closely correlated with endocrine phenotype and differentiation extent in pulmonary neuroendocrine tumors

The human homolog 1 of the Drosophila neurogenic achaete–scute genes, hASH1, is specifically expressed in fetal pulmonary neuroendocrine cells and in some neuroendocrine tumor cell lines. However, no data have been gathered regarding its in vivo expression in tumors. hASH1 mRNA expression was investigated by in situ hybridization in 238 surgically resected lung carcinomas, and the correlations between hASH1 expression status and immunostaining results of neuroendocrine markers chromogranin A, neural cell adhesion molecule, gastrin-releasing peptide and calcitonin, and clinical outcome were analyzed. hASH1 expression was detected in 2/20 (10%) adenocarcinomas, 4/30 (13.3%) typical carcinoids, 11/13 (84.6%) atypical carcinoids, 38/67 (56.7%) large-cell neuroendocrine carcinomas and 56/78 (71.8%) small-cell carcinomas, respectively, but not in any squamous cell carcinoma (0/21) or large-cell carcinoma (0/9). The 2 hASH1+ adenocarcinomas also expressed multiple neuroendocrine markers. Thus, hASH1 expression was restricted to lung cancers with neuroendocrine phenotypes. However, not all neuroendocrine tumors expressed hASH1. Within the entities of large-cell neuroendocrine carcinoma and small-cell carcinoma, hASH1 expression correlated very closely with chromogranin A, gastrin-releasing peptide and calcitonin expression (P<0.0001, r=0.852), but was not related to neural cell adhesion molecule expression (P=0.8892), suggesting that hASH1 expression, at least in lung cancer, is associated with endocrine phenotype expression other than ‘neuroendocrine differentiation’ in a broad sense. The fact that hASH1 was virtually absent in almost fully differentiated typical carcinoids, but was expressed in most, if not all, less differentiated atypical carcinoids as well as large-cell neuroendocrine carcinomas and small-cell carcinomas, suggests that hASH1 expression in lung cancer imitates its early and transient expression in fetal development, and that hASH1 is instrumental in the establishment, but not in the maintenance, of a cellular endocrine phenotype. Finally, hASH1 expression correlated with a significantly shortened survival in small-cell carcinoma patients (P=0.041).

HASH1 expression is closely correlated with endocrine phenotype and differentiation extent in pulmonary neuroendocrine tumors

The human homolog 1 of the Drosophila neurogenic achaete–scute genes, hASH1, is specifically expressed in fetal pulmonary neuroendocrine cells and in some neuroendocrine tumor cell lines. However, no data have been gathered regarding its in vivo expression in tumors. hASH1 mRNA expression was investigated by in situ hybridization in 238 surgically resected lung carcinomas, and the correlations between hASH1 expression status and immunostaining results of neuroendocrine markers chromogranin A, neural cell adhesion molecule, gastrin-releasing peptide and calcitonin, and clinical outcome were analyzed. hASH1 expression was detected in 2/20 (10%) adenocarcinomas, 4/30 (13.3%) typical carcinoids, 11/13 (84.6%) atypical carcinoids, 38/67 (56.7%) large-cell neuroendocrine carcinomas and 56/78 (71.8%) small-cell carcinomas, respectively, but not in any squamous cell carcinoma (0/21) or large-cell carcinoma (0/9). The 2 hASH1+ adenocarcinomas also expressed multiple neuroendocrine markers. Thus, hASH1 expression was restricted to lung cancers with neuroendocrine phenotypes. However, not all neuroendocrine tumors expressed hASH1. Within the entities of large-cell neuroendocrine carcinoma and small-cell carcinoma, hASH1 expression correlated very closely with chromogranin A, gastrin-releasing peptide and calcitonin expression (P<0.0001, r=0.852), but was not related to neural cell adhesion molecule expression (P=0.8892), suggesting that hASH1 expression, at least in lung cancer, is associated with endocrine phenotype expression other than ‘neuroendocrine differentiation’ in a broad sense. The fact that hASH1 was virtually absent in almost fully differentiated typical carcinoids, but was expressed in most, if not all, less differentiated atypical carcinoids as well as large-cell neuroendocrine carcinomas and small-cell carcinomas, suggests that hASH1 expression in lung cancer imitates its early and transient expression in fetal development, and that hASH1 is instrumental in the establishment, but not in the maintenance, of a cellular endocrine phenotype. Finally, hASH1 expression correlated with a significantly shortened survival in small-cell carcinoma patients (P=0.041).

U-shaped encodes a zinc finger protein that regulates the proneural genes achaete and scute during the formation of bristles in Drosophila.

The pattern of the large sensory bristles on the notum of Drosophila arises as a consequence of the expression of the achaete and scute genes. The gene u-shaped encodes a novel zinc finger that acts as a transregulator of achaete and scute in the dorsal region of the notum. Viable hypomorphic u-shaped mutants display additional dorsocentral and scutellar bristles that result from overexpression of achaete and scute. In contrast, overexpression of u-shaped causes a loss of achaete-scute expression and consequently a loss of dorsal bristles. The effects on the dorsocentral bristles appear to be mediated through the enhancer sequences that regulate achaete and scute at this site. The effects of u-shaped mutants are similar to those of a class of dominant alleles of the gene pannier with which they display allele-specific interactions, suggesting that the products of both genes cooperate in the regulation of achaete and scute. A study of the sites at which the dorsocentral bristles arise in mosaic u-shaped nota, suggests that the levels of the u-shaped protein are crucial for the precise positioning of the precursors of these bristles.

A chimeric enhancer-of-split transcriptional activator drives neural development and achaete-scute expression.

Drosophila melanogaster neurogenesis requires the opposing activities of two sets of basic helix-loop-helix (bHLH) proteins: proneural proteins, which confer on cells the ability to become neural precursors, and the Enhancer-of-split [E(spl)] proteins, which restrict such potential as part of the lateral inhibition process. Here, we test if E(spl) proteins function as promoter-bound repressors by examining the effects on neurogenesis of an E(spl) derivative containing a heterologous transcriptional activation domain [E(spl) m7Act (m7Act)]. In contrast to the wild-type E(spl) proteins, m7Act efficiently induces neural development, indicating that it binds to and activates target genes normally repressed by E(spl). Mutations in the basic domain disrupt m7Act activity, suggesting that its effects are mediated through direct DNA binding. m7Act causes ectopic transcription of the proneural achaete and scute genes. Our results support a model in which E(spl) proteins normally regulate neurogenesis by direct repression of genes at the top of the neural determination pathway.

The human Achaete-Scute homologue 2 (ASCL2,HASH2) maps to chromosome 11p15.5, close to IGF2 and is expressed in extravillus trophoblasts.

Here we describe the cloning of the human Achaete Scute Homologue 2 (HASH2) gene, officially designated ASCL2 (Achaete Scute complex like 2), a homologue of the Drosophila Achaete and Scute genes. In mouse, this gene is imprinted and maps to chromosome 7. We mapped the human homologue close to IGF2 and H19 at 11p15.5, the human region syntenic with mouse chromosome 7, indicating that this imprinted region is highly conserved in mouse and man. HASH2 is expressed in the extravillus trophoblasts of the developing placenta only. The lack of HASH2 expression in non-malignant hydatidiform (androgenetic) moles indicates that HASH2 is also imprinted in man.

Drosophila development: A prepattern for sensory organs

The sensory bristles of Drosophila arise in stereotyped positions from small clusters of cells that express achaete–scute genes. A set of genes has now been identified that regulate achaete–scute expression and form a prepattern for sensory bristle development.

Cis-regulation of achaete and scute: shared enhancer-like elements drive their coexpression in proneural clusters of the imaginal discs.

The pattern of bristles and other sensory organs on the adult cuticle of Drosophila is prefigured in the imaginal discs by the pattern of expression of the proneural achaete (ac) and scute (sc) genes, two members of the ac-sc complex (AS-C). These genes are simultaneously expressed by groups of cells (the proneural clusters) located at constant positions in discs. Their products (transcription factors of the basic-helix-loop-helix family) allow cells to become sensory organ mother cells (SMCs), a fate normally realized by only one or a few cells per cluster. Here we show that the highly complex pattern of proneural clusters is constructed piecemeal, by the action on ac and sc of site-specific, enhancer-like elements distributed along most of the AS-C (approximately 90 kb). Fragments of AS-C DNA containing these enhancers drive reporter lacZ genes in only one or a few proneural clusters. This expression is independent of the ac and sc endogenous genes, indicating that the enhancers respond to local combinations of factors (prepattern). We show further that the cross-activation between ac and sc, discovered by means of transgenes containing either ac or sc promoter fragments linked to lacZ and thought to explain the almost identical patterns of ac and sc expression, does not occur detectably between the endogenous ac and sc genes in most proneural clusters. Our data indicate that coexpression is accomplished by activation of both ac and sc by the same set of position-specific enhancers.

The function of the proneural genes achaete and scute in the spatio-temporal patterning of the adult labellar bristles of Drosophila melanogaster.

The sensory precursors for labellar taste bristles develop from the labial disc in three distinct temporal waves occurring at 0 h, 8 h and 14 h of pupal development. In each temporal wave, transcripts for the achaete (ac) and scute (sc) genes are expressed in overlapping patterns in cells of the disc epithelium prior to the appearance of sensory mother cells (SMCs). No bristles form in mutant flies in which the ac and sc genes are absent. When the sc gene alone is deleted, a set of seven bristles fail to form. Pulses of ubiquitous sc + expression during pupal development, in a strain mutant for both ac and sc, can result in flies with all the labellar bristles at their correct positions. sc + pulses at times corresponding to the initiation of each of the waves of SMC specification in the disc was sufficient to restore bristle pattern. Bristles were not induced at ectopic positions and times as a result of the ubiquitous expression of sc +. These results suggest that the proneural genes ac and sc do not themselves set the pattern of the labellar bristles. Instead, they are required for the elaboration of the pattern set by other gene products. We also show that the formation and positioning of the later waves of bristles can take place even in the absence of bristles normally specified earlier.

Gene regulation in two dimensions: the proneural achaete and scute genes are controlled by combinations of axis-patterning genes through a common intergenic control region.

The mechanisms that generate precise patterns of discrete cell types within developing fields are not well understood. One model for analyzing how cells interpret positional information in two dimensions is the regulation of proneural cluster formation within insect segments. Two adjacent proneural regulatory genes, achaete and scute, are expressed coincidently in cell clusters at reproducible anteroposterior (AP) and dorsoventral (DV) coordinates within the Drosophila embryo from which single neuroblasts later arise. Here, we show that the AP and DV position of these clusters is regulated through a common cis-acting region between the genes under the initial control of the products of the pair-rule and DV polarity genes and is later maintained by selected segment polarity genes. The combination of proneural gene activation/repression in AP stripes and repression within specific DV domains positions each cluster of achaete/scute expressing cells within segments; interactions between these cells then determine individual cell fates.

The extramacrochaetae gene provides information for sensory organ patterning.

The Drosophila adult epidermis displays a stereotyped pattern of bristles and other types of sensory organs (SOs). Its generation requires the proneural achaete (ac) and scute (sc) genes. In the imaginal wing disc, the anlage for most of the thoracic and wing epidermis, their products accumulate in groups of cells, the proneural clusters, whose distribution prefigures the adult pattern of SOs. These proteins then induce the emergence of SO mother cells (SMCs). Here, we show that the extramacrochaetae (emc) gene, an antagonist of the proneural function, is another agent that contributes to SO positioning. In the wing disc, emc is expressed in a complex and evolving pattern. SMCs appear not only within proneural clusters but also within minima of emc expression. When one of these spatial restrictions is eliminated, by ubiquitously expressing ac-sc, SMCs still emerge within minima of emc. When in addition, the other spatial restriction is reduced by decreasing emc expression, many ectopic SMCs emerge in a relatively even spaced and less constant pattern. Thus, the heterogeneous distribution of the emc product is one of the elements that define the positions where SMCs arise. emc probably refines SMC (and SO) positioning by reducing both the size of proneural clusters and the number of cells within clusters that can become SMCs.

Cross-Regulatory Interactions Between the Proneural achaete and scute Genes of Drosophila

The achaete (ac) and scute (sc) genes of Drosophila allow cells to become sensory organ mother cells. Although ac and sc have similar patterns of expression, deletion of either gene removes specific subsets of sensory organs. This specificity was shown to reside in the peculiar regulation of ac and sc expression. These genes are first activated in complementary spatial domains in response to different cis-regulatory sequences. Each gene product then stimulates expression of the other gene, thus generating similar patterns of expression. Therefore, removal of one gene leads to the absence of both proneural gene products and sensory organs in the sites specified by its cis-regulatory sequences.

Competence to develop sensory organs is temporally and spatially regulated in Drosophila epidermal primordia.

The Drosophila adult cuticle displays a stereotyped pattern of sensory organs (SOs). Its deployment requires the expression of the achaete (ac) and scute (sc) genes. Their products confer to cells of epidermal primordia (imaginal discs and histoblasts) the ability to become SO precursors (SOPs). In imaginal discs, ac and sc expression is spatially restricted to cell clusters within which one or a few cells become SOP(s). With the help of ubiquitous sc expression provided at different developmental times by a heat shock-sc (HSSC) chimeric gene, we have analyzed the response of epidermal primordia to the proneural action of the sc product, and have tested whether the patterned distribution of ac/sc products is necessary to position SOs correctly within the epidermis. Each primordium responds to HSSC expression by developing SOs only during a characteristic developmental period. In the absence of the endogenous ac and sc genes, most SOs induced by HSSC are of the correct type and are located in wild type positions. These results indicate that the capacity of primordia to respond to sc is temporally and spatially regulated, that specification of the type of SO does not depend on ac/sc, and that SO positioning utilizes topological information independent of the spatially restricted distribution of ac/sc products.

Extramacrochaetae, a negative regulator of sensory organ development in Drosophila, defines a new class of helix-loop-helix proteins

The function of the extramacrochaetae ( emc) gene is required to establish the normal spatial pattern of adult sensory organs in Drosophila. emc acts to suppress sensory organ development in certain regions of the body surface, apparently by antagonizing the function of the achaete and scute genes of the achaetescute complex ( AS-C). We have found that emc encodes a novel member of the helix-loop-helix (HLH) family of proteins. The emc protein shares the dimerization domain of other HLH proteins but lacks their DNA binding motif. We propose a model in which the emc protein negatively regulates sensory organ determination by forming heterodimers with the HLH proteins encoded by the AS-C and/or daughterless, thereby altering or interfering with their activity.

Expression of achaete and scute genes in Drosophila imaginal discs and their function in sensory organ development.

Several kinds of sensory organs (SOs) appear in stereotyped positions on the adult Drosophila cuticle. The generation of these SOs requires the activity of the achaete (ac) and scute (sc) genes. To investigate whether ac and sc also provide spatial information for the positioning of SOs, we have analyzed the patterns of expression of these genes in the wing imaginal disc around the time that SO precursors are being specified. We find that expression coincides with and is restricted to areas of the disc where these precursors are known to be located. In the loss-of-function sc mutant, sc RNA is depleted in a single area located in the region where the precursor for the suppressed macrochaeta should be found. Moreover, some, and probably all, SOs require expression of these genes to reach the earliest detectable differentiated state. These and other results presented here, together with the finding that expansion of the areas of ac and/or sc expression causes the development of ectopic SOs, indicate that ac and sc promote the determination of SO precursors and delimit the regions of the imaginal discs where they can develop.

Insertion-deletion variation at theyellow-achaete-scuteregion in two natural populations ofDrosophila melanogaster

We have surveyed the region of the X chromosome of Drosophila melanogaster which encodes the yellow, achaete and scute genes for restriction map variation. Two natural populations, one from North Carolina, U.S.A. and the other from southern Spain were screened for variation at about 70 restriction sites and for variation due to DNA insertion or deletion events in 120 kilobases of DNA. Mean heterozygosity per nucleotide was estimated to be 0.0024 and 15 large insertions were found in the 49 chromosomes screened. Extensive disequilibrium between polymorphic sites were found across much of the region in the North Carolina population. The frequency of large insertions, which usually correspond to transposable genetic elements, is significantly lower than has been observed in autosomal regions of the genome. This is predicted for X-linked loci by certain models of transposable element evolution, where copy number is restricted by virtue of the recessive deleterious effects of the insertions. Our results appear to support such models. The deficiency of insertions may in this case be enhanced by hitch-hiking effects arising from the high level of disequilibrium.