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Zygotically transcribed genes

Proneural and Neurogenic Genes

What is the Notch pathway?

Developmental biology of the Notch pathway

The achaete-scute complex

Embryonic central nervous system lineages

Aging neural progenitors lose competence to respond to mitogenic Notch signaling

BLOS2 negatively regulates Notch signaling during neural and hematopoietic stem and progenitor cell development

Neurogenic genes functioning in the Notch pathway

Proneural genes of the achaete-scute complex

Additional proneural genes

What is the Notch pathway?

To understand the Notch pathway one has first to know the function of receptors and ligands. A receptor is a protein molecule located partially outside the cell and partially inside. The receptor interacts with outside signals and transmits them to the inside. Ligands are extracellular proteins or other molecules that interact with a receptor to cause it to transmit a signal to the inside of the cell. Notch is a receptor that receives signals from its ligands and transmits these signals to the inside of the cell. The Notch pathway consists of Notch and its ligands, as well as intracellular proteins that transmit the Notch signal to the nucleus. Included in the pathway are transcription factors, the proteins that bear the effector function of the pathway.

The effect of Notch signaling is the phenomenon of lateral inhibition, the singling out of one cell from a cell cluster for a given fate (neurogenesis for example), and the inhibition of those cells not elected to differentiate. Lateral inhibition occurs repeatedly in Drosophila development. Central to this process is the interaction of Notch receptor with a group of ligands including Delta, Scabrous and Serrate. Some of these ligands float free around the outside of the cell and others are bound to the surface of cells.

The interaction of Notch with its ligand triggers a chain of intracellular events resulting in lateral inhibition, the prevention of a specified fate committment on the part of most of the cells of a cluster. The first link in the chain of events is a reduction of the affinity of Suppressor of Hairless protein for the cytoplasmic tail of the Notch receptor. Following this, Su(H) is free to enter the nucleus where it assumes its role as a transcription factor. Su(H) mediates transcriptional activation of Enhancer of split complex of genes. This complex will then inhibit the proneural genes achaete, scute, lethal of scute and asense. The transcriptional inhibitory capacity of Enhancer of split complex genes results in lateral inhibition.

The process of lateral inhibition often involves cell migration. In addition to the signaling capacity of Notch receptor, a second important function of Notch and its ligand Delta is that of cell adhesion. Delta is bound to the surface of cells. The Delta-Notch interaction results in an intimate adhesive contact between fated cell, which often migrates, and inhibited cell, bearing Notch, which remains in its germ layer or cluster of origin. Thus the interaction of Notch with its ligand causes the lateral inhibition of the cell bearing Notch, and the migration and triggering of a differentiated state for the cell bearing the Notch ligand. The ligand most often turns out to be Delta.

How can the Notch pathway components be called neurogenic if the function of Notch is to repress the adoption of a differentiated state in the cell that carries Notch protein? There are two answers to this question. The first is that the term neurogenic is a misnomer, arising because the phenotype of Notch pathway mutants manifests the result of an over production of neurons. The pathway is named neurogenic because of this mutant phenotype. The second answer lies in considering the principle ligand of Notch and its function. Delta is not secreted but is cell bound. The Delta-Notch interaction serves a cell adhesive function between ligand and receptor bearing cells. While the receptor bearing cell is inhibited in assuming a differentiated state, the ligand bearing cell is free to do so. In neurogenesis this cell delaminates, and then migrates out of the epithelial cell layer. The ligand bearing cell assumes the differentiated state of neuroblast and physically resides in the developing nervous system. Thus Notch is neurogenic with respect to the cell that bears the Notch ligand.

Developmental biology of the Notch pathway

The Notch ligand receptor system functions in all three germ layers during embryonic development as well as in the germ cell line and in imaginal discs during the larval and pupal stages. The outline below lists developmental processes in which the Notch pathway functions. All are affected by Notch and where tested, all were found to be affected by Delta (Hartenstein, 1992).

The achaete-scute complex

The achaete-scute complex (AS-C) contains four related genes, achaete, scute, lethal of scute and asense. These are spread over 90 kilobases. The proteins coded for by these genes serve as transcription factors. Each has a DNA binding domain, termed a basic helix-loop-helix domain, and three of the four proteins (the exception is Asense) have a C-terminal acidic domain.

achaete and scute genes are more than 25 kb apart. lethal of scute is more than 12 kb from scute. asense is another 45 kb removed from lethal of scute.

These genes are termed proneural, since they promote neuroblast differentiation and are thus essential for the differentiation of the central and peripheral nervous system and the brain. Other functions are also served by these genes: regulation of sex determination (scute); participation in specification of muscle progenitors (lethal of scute), and regulation of sequential fates in Malpighian tubule development (achaete).

How are achaete-scute complex genes activated and inactivated? Genes like hairy, extramachrochaete and pannier help to establish patterns of achaete expression by acting as repressors. The first two genes received their names because of their interaction with achaete: mutation in either produces extra sensory elements such as hairs and machrochaete. As achaete is responsible for activating the sensory cell fate, Hairy and EMC act to restrict achaete transcription to a limited region where sensory elements will form.

NK2/Ventral nervous system defective regulates the expression of achaete and scute to the medial column of the ventral nervous system (Skeath, 1994). Specifically, first NK2 is essential for AS-C gene expression in the medial column of every other neuroblast row through regulatory elements located 3' to achaete, and second, through a 5' regulatory region, NK2 functions to increase or maintain proneural gene expression within the proneural cluster that normally gives rise to the neuroblast (Skeath, 1994).

The above information about the three repressors and NK2 provides only a nearsighted view of activation and repression by the AS-C. There is a more global answer to the question of how AS-C genes are activated. AS-C genes are activated in proneural clusters in the ventral portion of the fly. The anterior-posterior arrangement of neuroepithelial clusters is intimately connected to the process of segmentation. The proneural genes are expressed in each of the 14 segments. Pair rule genes define these segments, and therefore the placement of proneural AS-C clusters. In fact embryos that are mutant in each of the pair rule genes show an alteration of early cluster pattern.

It is not just the presence or absence of a cluster that is controlled by pair rule genes, but the arrangement of neuroblasts within each cluster, and consequently the arrangement of cells expressing AS-C genes. Each cluster is subdivided into subdomains, and each subdomain has its particular arrangement of neuroblasts. Segment polarity genes act here too, especially gooseberry and naked. They help maintain the AP borders of the cells within each cluster (Skeath, 1992).

See Chris Doe's Hyper-Neuroblast map site for information on the origin and lineage of specific neuroblasts in the CNS and for information about the expression of specific genes regulating neuroblast origin.

achaete and scute are controlled by combinations of axis-patterning genes through a common enhancer or locus control region. The pair rule genes act through this region, situated in the intervening chromosomal segment between achaete and scute. Thus this region is under the initial control of products of the pair rule genes and is later maintained by selected segment polarity genes.

Consider the amazing patterning present in insect epidermis. Flies have hairs that appear as reproducable patterns from one individual to another. These patterns are species specific and therefore distinguish one species from another. How are these patterns made?

Further analysis of the 90 kD region between achaete and asense has defined nine site-specific enhancer-like elements that function to regulate achaete and scute expression in proneural clusters of imaginal discs. Ultimately it is the capacity of site specific enhancers to kindle AS-C transcription at very localized regions in the proneural clusters of imaginal discs that is responsible for the patterning of sensory elements seen in adults.

Each of these enhancers, regulated by a specific set of transcription factors defines a prepattern for achaete and scute expression. The enhancers are named after the sensilla in which they direct achaete and scute expression. For example the L3/TSM enhancer, just upstream of scute directs achaete and scute to third vein sensilla campaniforma (L3) and twin sensilla of the anterior wing margin (TSM) (Gomez-Skarmeta, 1995). It is rather remarkable that each sensory organ or group is specificied by a special enhancer selected through evolution for just that purpose. For a historical view of this incredible work see Ghysen, 1988.

Embryonic central nervous system lineages

In Drosophila, central nervous system (CNS) formation starts with the delamination from the neuroectoderm of about 30 neuroblasts (NBs) per hemisegment. These give rise to approximately 350 neurons and 30 glial cells during embryonic development. Understanding the mechanisms leading to cell fate specification and differentiation in the CNS requires the identification of the NB lineages. Each segment is also subdivided along its dorsal-ventral axis. The delamination of the neuroblasts (NBs) from the neuroectoderm occurs between embryonic stages 8 and 11 and is divided into five phases (S1-S5). On each side of the ventral midline there are three longitudinal columns of cells: medial, intermediate and lateral. S1-S3 NBs form these three longitudinal columns. The positions of these three columns is regulated by genes defining the dorsal-ventral axis: twist, snail decapentaplegic and tolloid. These genes likewise act through the enhancer located between achaete and scute (Skeath, 1992). S4 and S5 NBs become interspersed between the existing columns of NBs.

Perpendicular to the columns, NBs are numbered in each segment from anterior to posterior as rows 1-7. Thus NB 1-2 is in the medial column (1) and represents the 2nd neuroblast in each segment. There are a total of thirty identified neuroblasts in each (right or left) half of a segment (hemisegment), and they give rise to a total of 350 progeny cells. For instance, NB 1-1 delaminates preferentially as a S1 NB. Around mid stage 11 the anterior-posterior position of NB1-1 is in line with, or slightly anterior to, the tracheal pits. At this stage it has given rise to a cluster of six to eight daughter cells on its dorsal side. At the beginning of germ band shortening (end stage 11) two of the progeny, the aCC and pCC leave the cluster and move anteriorly. They are the first progeny of NB 1-1. The aCC neuron innervates a dorsal muscle. Each thoracic NB 1-1 gives rise to 8 - 14 cells including ipsilaterally projecting interneurons and 1 or 2 motoneurons. In the abdomen NB 1-1 gives rise to a smaller cluster comprising four to six ipsilaterally projecting interneurons and no motoneurons; in addition the abdominal clone comprises 3 subperineural glial cells (Bossing, 1996). Different segmental fates of each neuroblast is determined by homeotic genes.

Thirteen lineages derived from the dorsal (lateral) part of the neuroectoderm are described here and 12 of them are assigned to identified NBs. Together, the 13 lineages comprise approximately 120 neurons and 22 to 27 glial cells which have been included in a systematic terminology. Therefore, NBs from the dorsal neuroectoderm produce about 90% of the glial cells in the embryonic ventral ganglion. Two of the NBs give rise to glial progeny exclusively (NB 6-4A, GP); five NBs give rise to glia as well as neurons (NBs 1-3, 2-5, 5-6, 6-4T, 7-4). These seven NBs are arranged as a group in the most lateral region of the NB layer. The other lineages (NBs 2-4, 3-3, 3-5, 4-3, 4-4, 5-4, clone y) are composed exclusively of neurons (interneurons, motoneurons, or both). It has been possible to link the lateral cluster of even-skipped expressing cells (EL) to the lineage of NB 3-3. Along with the previously described clones, the vast majority (more than 90%) of cell lineages in the embryonic ventral nerve cord (in the thorax and abdomen) are now known. Previously identified neurons and most glial cells are now linked to certain lineages and, thus, to particular NBs. This complete set of data provides a foundation for the interpretation of mutant phenotypes and for future investigations on cell fate specification and differentiation (Schmidt, 1997).

Aging neural progenitors lose competence to respond to mitogenic Notch signaling

Drosophila neural stem cells (neuroblasts) are a powerful model system for investigating stem cell self-renewal, specification of temporal identity, and progressive restriction in competence. Notch signaling is a conserved cue that is an important determinant of cell fate in many contexts across animal development; for example, mammalian T cell differentiation in the thymus and neuroblast specification in Drosophila are both regulated by Notch signaling. However, Notch also functions as a mitogen, and constitutive Notch signaling potentiates T cell leukemia as well as Drosophila neuroblast tumors. While the role of Notch signaling has been studied in these and other cell types, it remains unclear how stem cells and progenitors change competence to respond to Notch over time. Notch is required in type II neuroblasts for normal development of their transit amplifying progeny, intermediate neural progenitors (INPs). This study finds that aging INPs lose competence to respond to constitutively active Notch signaling. Moreover, reducing the levels of the old INP temporal transcription factor Eyeless/Pax6 allows Notch signaling to promote the de-differentiation of INP progeny into ectopic INPs, thereby creating a proliferative mass of ectopic progenitors in the brain. These findings provide a new system for studying progenitor competence and identify a novel role for the conserved transcription factor Eyeless/Pax6 in blocking Notch signaling during development (Farnsworth, 2015).

BLOS2 negatively regulates Notch signaling during neural and hematopoietic stem and progenitor cell development

Notch signaling plays a crucial role in the control of proliferation and differentiation of stem and progenitor cells during embryogenesis or organogenesis, but its regulation is incompletely understood. BLOS2, encoded by the Bloc1s2 gene (see Drosophila Blos2), is a shared subunit of two lysosomal trafficking complexes, biogenesis of lysosome-related organelles complex-1 (BLOC-1) and BLOC-1 related complex. Bloc1s2-/- mice are embryonic lethal and exhibit defects in cortical development and hematopoiesis. Loss of BLOS2 results in elevated Notch signaling, which consequently increases the proliferation of neural progenitor cells and inhibits neuronal differentiation in cortices. Likewise, ablation of bloc1s2 in zebrafish or mice leads to increased hematopoietic stem and progenitor cell production in the aorta-gonad-mesonephros region. BLOS2 physically interacts with Notch1 in endo-lysosomal trafficking of Notch1. These findings suggest that BLOS2 is a novel negative player in regulating Notch signaling through lysosomal trafficking by controlling multiple stem and progenitor cell homeostasis in vertebrates (Zhou, 2016).

list of proneural and neurogenic genes


Bossing, T., et al. (1996). The embryonic central nervous system lineages of Drosophila melanogaster. I. Neuroblast lineages derived from the ventral half of the neuroectoderm. Dev. Biol 179: 41-64. 8873753

Cummings, C.A. and Cronmiller, C. (1994). The daughterless gene functions together with Notch and Delta in the control of ovarian follicle development in Drosophila. Development 120: 381-94. 8149916

Farnsworth, D. R., Bayraktar, O. A. and Doe, C. Q. (2015). Aging neural progenitors lose competence to respond to mitogenic Notch signaling. Curr Biol 25: 3058-3068. PubMed ID: 26585279

Ghysen, A. and Dambly-Chaudiere, C. (1988). From DNA to form: The achaete-scute complex. Genes Dev. 2: 495-501

Gomez-Skarmeta, J. L., et al. (1995). Cis-regulation of achaete and scute: shared enhancer-like elements drive their coexpression in proneural clusters of the imaginal discs. Genes Dev. 9: 1869-1882. 7649474

Gonzalez-Gaitan, M. and Jackle, H. (1995). Invagination centers within the Drosophila stomatogastric nervous system anlage are positioned by Notch-mediated signaling which is spatially controlled through wingless. Development 121: 2313-25. 7671798

Hartenstein, A.Y., Rugendorff, A., Tepass, U. and Hartenstein, V. (1992). The function of the neurogenic genes during epithelial development in the Drosophila embryo. Development 116(4): 1203-1220. 1295737

Schmidt, H., et al. (1997). The embryonic central nervous system lineages of Drosophila melanogaster. II. Neuroblast lineages derived from the dorsal part of the neuroectoderm. Dev. Biol. 189(2): 186-204

Schweisguth, F. (1995). Suppressor of Hairless is required for signal reception during lateral inibition in the Drosophila pupal notum. Development 121: 1875-1884. 7601001

Skeath, J. B. et al. (1992). 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. Genes Dev 6: 2606-19. 7601001

Skeath, J. B., Panganiban, J. F. and Carroll, S. B. (1994). The ventral nervous system defective gene controls proneural gene expression at two distinct steps during neuroblast formation in Drosophila. Development 120: 1517-1524. 8050360

Zhou, W., He, Q., Zhang, C., He, X., Cui, Z., Liu, F. and Li, W. (2016). BLOS2 negatively regulates Notch signaling during neural and hematopoietic stem and progenitor cell development. Elife [Epub ahead of print]. PubMed ID: 27719760

Zygotically transcribed genes

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