Gene name - runt
Cytological map position - 19E1-2
Function - transcription factor
Symbol - run
Genetic map position - 1-65
Classification - novel
Cellular location - nuclear
|Recent literature||Hang, S. and Gergen, J.P. (2017). Different modes of enhancer-specific regulation by Runt and Even-skipped during Drosophila segmentation. Mol Biol Cell [Epub ahead of print]. PubMed ID: 28077616
The initial metameric expression of the Drosophila sloppy paired 1 (slp1) gene is controlled by two distinct cis-regulatory DNA elements that interact in a non-additive manner to integrate inputs from transcription factors encoded by the pair-rule segmentation genes. This study performed Chromatin Immuno-Precipitation (ChIP) on reporter genes containing these elements in different embryonic genotypes to investigate the mechanism of their regulation. The Distal Early Stripe Element (DESE) mediates both activation and repression by Runt. The differential response of DESE to Runt was found to be due to an inhibitory effect of Fushi tarazu (Ftz) on P-TEFb recruitment and the regulation of RNA Polymerase II (Pol II) pausing. The Proximal Early Stripe Element (PESE) is also repressed by Runt, but in this case Runt prevents PESE-dependent Pol II recruitment and pre-initiation complex (PIC) assembly. PESE is also repressed by Even-skipped (Eve) but interestingly this repression involves regulation of P-TEFb recruitment and promoter-proximal Pol II pausing. These results demonstrate that the mode of slp1 repression by Runt is enhancer-specific whereas the mode of repression of the slp1 PESE enhancer is transcription factor-specific. The study proposes a model based on these differential regulatory interactions that accounts for the non-additive interactions between the PESE and DESE enhancers during Drosophila segmentation.
|Koromila, T. and Stathopoulos, A. (2017). Broadly expressed repressors integrate patterning across orthogonal axes in embryos. Proc Natl Acad Sci U S A. PubMed ID: 28720706
The role of spatially localized repressors in supporting embryonic patterning is well appreciated, but, alternatively, the role ubiquitously expressed repressors play in this process is not well understood. This study investigated the function of two broadly expressed repressors, Runt (Run) and Suppressor of Hairless [Su(H)], in patterning the Drosophila embryo. Previous studies have shown that Run and Su(H) regulate gene expression along anterior-posterior (AP) or dorsal-ventral (DV) axes, respectively, by spatially limiting activator action, but this study characterizes a different role. The data show that broadly expressed repressors silence particular enhancers within cis-regulatory systems, blocking their expression throughout the embryo fully but transiently, and, in this manner, regulate spatiotemporal outputs along both axes. These results suggest that Run and Su(H) regulate the temporal action of enhancers and are not dedicated regulators of one axis but, instead, act coordinately to pattern both axes, AP and DV.
|Koromila, T. and Stathopoulos, A. (2019). Distinct roles of broadly expressed repressors support dynamic enhancer action and change in time. Cell Rep 28(4): 855-863. PubMed ID: 31340149
How broadly expressed repressors regulate gene expression is incompletely understood. To gain insight, this study investigated how Suppressor of Hairless-Su(H)-and Runt regulate expression of bone morphogenetic protein (BMP) antagonist short-gastrulation via the sog_Distal enhancer. A live imaging protocol was optimized to capture this enhancer's spatiotemporal output throughout the early Drosophila embryo, finding in this context that Runt regulates transcription initiation, Su(H) regulates transcription rate, and both factors control spatial expression. Furthermore, whereas Su(H) functions as a dedicated repressor, Runt temporally switches from repressor to activator. These results demonstrate that broad repressors play temporally distinct roles and contribute to dynamic gene expression. Both Run and Su(H)'s ability to influence the spatiotemporal domains of gene expression may serve to counterbalance activators and function in this manner as important regulators of the maternal-to-zygotic transition in early embryos.
|Mahadeveraju, S., Jung, Y. H. and Erickson, J. W. (2020). Evidence that Runt acts as a counter-repressor of Groucho during Drosophila melanogaster primary sex determination. G3 (Bethesda). PubMed ID: 32457096
Runx proteins are bifunctional transcription factors that both repress and activate transcription in animal cells. Typically, Runx proteins work in concert with other transcriptional regulators, including co-activators and co-repressors to mediate their biological effects. In Drosophila melanogaster the archetypal Runx protein, Runt, functions in numerous processes including segmentation, neurogenesis and sex determination. During primary sex determination Runt acts as one of four X-linked signal element (XSE) proteins that direct female-specific activation of the establishment promoter (Pe) of the master regulatory gene Sex-lethal (Sxl). Successful activation of SxlPe requires that the XSE proteins overcome the repressive effects of maternally deposited Groucho (Gro), a potent co-repressor of the Gro/TLE family. Runx proteins, including Runt, contain a C-terminal peptide, VWRPY, known to bind to Gro/TLE proteins to mediate transcriptional repression. This study shows that Runt's VWRPY co-repressor-interaction domain is needed for Runt to activate SxlPe Deletion of the Gro-interaction domain eliminates Runt-ability to activate SxlPe, whereas replacement with a higher affinity, VWRPW, sequence promotes Runt-mediated transcription. This suggests that Runt may activate SxlPe by antagonizing Gro function, a conclusion consistent with earlier findings that Runt is needed for Sxl expression only in embryonic regions with high Gro activity. Surprisingly it was found that Runt is not required for the initial activation of SxlPe Instead, Runt is needed to keep SxlPe active during the subsequent period of high-level Sxl transcription suggesting that Runt helps amplify the difference between female and male XSE signals by counter-repressing Gro in female, but not in male, embryos.
|Miller, A. C., Urban, E. A., Lyons, E. L., Herman, T. G. and Johnston, R. J., Jr. (2020). Interdependent regulation of stereotyped and stochastic photoreceptor fates in the fly eye. Dev Biol 471: 89-96. PubMed ID: 33333066
Diversification of neuronal subtypes often requires stochastic gene regulatory mechanisms. How stochastically expressed transcription factors interact with other regulators in gene networks to specify cell fates is poorly understood. The random mosaic of color-detecting R7 photoreceptor subtypes in Drosophila is controlled by the stochastic on/off expression of the transcription factor Spineless (Ss). In Ss(ON) R7s, Ss induces expression of Rhodopsin 4 (Rh4), whereas in Ss(OFF) R7s, the absence of Ss allows expression of Rhodopsin 3 (Rh3). This study finds that the transcription factor Runt, which is initially expressed in all R7s, is sufficient to promote stochastic Ss expression. Later, as R7s develop, Ss negatively feeds back onto Runt to prevent repression of Rh4 and ensure proper fate specification. Together, stereotyped and stochastic regulatory inputs are integrated into feedforward and feedback mechanisms to control cell fate.
|Prazak, L., Iwasaki, Y., Kim, A. R., Kozlov, K., King, K. and Gergen, J. P. (2021). A dual role for DNA-binding by Runt in activation and repression of sloppy paired transcription. Mol Biol Cell: mbcE20080509. PubMed ID: 34432496
This work investigates the role of DNA-binding by Runt in regulating the sloppy-paired-1 (slp1) gene, and in particular two distinct cis-regulatory elements that mediate regulation by Runt and other pair-rule transcription factors during Drosophila segmentation. A DNA-binding defective form of Runt was found to br ineffective at repressing both the distal (DESE) and proximal (PESE) early stripe elements of slp1 and is also compromised for DESE-dependent activation. The function of Runt-binding sites in DESE is further investigated using site-specific transgenesis and quantitative imaging techniques. When DESE is tested as an autonomous enhancer, mutagenesis of the Runt sites results in a clear loss of Runt-dependent repression but has little to no effect on Runt-dependent activation. Notably, mutagenesis of these same sites in the context of a reporter gene construct that also contains the PESE enhancer results in a significant reduction of DESE-dependent activation as well as the loss of repression observed for the autonomous mutant DESE enhancer. These results provide strong evidence that DNA-binding by Runt directly contributes to the regulatory interplay of interactions between these two enhancers in the early embryo.
Runt is a novel protein, unrelated to homeodomain, zinc finger or other transcription factors. It has a mammalian homolog that binds to enhancers of retroviruses and polyoma virus, and is involved in T-cell maturation. Like hairy and even-skipped, runt is termed a primary pair rule gene, as opposed to a secondary pair rule gene. Transcription of primary pair rule genes is regulated directly by maternal genes and gap genes, while secondary pair rule genes are regulated by the primary pair rule genes. Arguments can be made as to the validity and ultimate usefulness of primary/secondary pair rule distinctions; the notion is discussed in more detail the fushi tarazu overview.
Runt's effects are felt throughout the developmental hierarchy. Runt can modulate the activity of other pair rule genes, including hairy, even-skipped and ftz. runt possesses gap gene properties as well, altering the expression of giant and hunchback when transiently overexpressed (Tsai, 1994). Runt acts early in sex determination. Its activity is necessary to activate Sex lethal in the soma, but not in the germ line (Garandino, 1993). Runt also interacts with bicoid, restricting bicoid expression in the trunk, although the mechanism for this regulation is not completely understood (Tsai,1994). runt is also involved in neurogenesis, in the specification of neuroblasts (Kania, 1990).
runt is expressed by a subset of the 30 neuroblasts that give rise to each neuromere of the Drosophila embryo. Runt is also expressed in a subset of ganglion mother cells and neurons and its activity has been shown to be necessary for the formation of a subset of even-skipped (eve)-expressing lateral neurons, the EL neurons. There are 8-10 EL neurons per abdominal hemisegment, which originate from neuroblast 3-3. The EL neurons are interneurons that express the zinc-finger transcription factor encoded by eagle. The EL neurons project axons through the anterior commissure across the midline, then turn anteriorly into the longitudinal fascicles. Inactivation of runt during neuroblast delamination, using a temperature-sensitive allele of runt, leads to a loss of eve expression in the EL neurons. Eve expression in the EL neurons is not affected when Runt is inactivated after the neuroblasts have delaminated, suggesting that Runt activity is necessary only at the time of neuroblast delamination for the development of the EL neurons (Dormand, 1998).
To determine which neuroblasts express Runt, embryos were triple labelled with anti-Runt, anti-En and anti-Gsb-d. En is expressed by neuroblasts in row 6 and 7 and neuroblast 1-2; Gsb-d is expressed by neuroblasts in row 5 and 6 and neuroblast 7-1. Contrary to the previously published expression pattern, which showed expression of Runt in the neuroblasts only up to stage 10, it was found that Runt is expressed in neuroblasts throughout neurogenesis. Neuroblasts 2-2, 2-5, 3-1, 3-2, 5-2 and 5-3 express Runt from the time of their delamination (stage 10). By stage 11, Runt is also expressed in neuroblasts 2-3 and 3-3, and expression is lost from neuroblast 2-5. Runt was verified to be expressed in neuroblast 3-3 by double staining for Eagle, which is expressed by neuroblasts 2-4, 3-3, 6-4 and 7-3. A previously published expression pattern of Runt had not shown Runt expression in neuroblast 3-3, which gives rise to the EL neurons. It has been reported that neuroblasts 1-1 and 4-1 also express Runt, but this is not seen in the current study. Therefore, Runt is expressed in five neuroblasts in rows 2 to 3 (neuroblast 2-2, 2-3, 3-1, 3-2 and 3-3) and two neuroblasts in row 5 (neuroblast 5-2 and 5-3) (Dormand, 1998).
Runt is expressed by a large number of GMCs and neurons including the EL neurons. Runt is also expressed in a cluster of two to four cells on the midline and in a pair of neurons one on each side of the midline. By double labelling with anti-Odd, which labels MP1 and dMP2, it was found that the neurons on each side of the midline are the MP1 neurons. Double labelling with antibody to Slit, which labels the midline glia, identifies the cluster of Runt-expressing cells on the midline as the midline glia (Dormand, 1998).
runt is a good candidate for a gene that specifies neuroblast identities. To test this, Runt was ectopically expressed in restricted subsets of neuroblasts. Runt is sufficient to activate even-skipped expression in the progeny of specific neuroblasts. Eve is ectopically induced when runt is mis-expressed in all neuroblasts, using the pan neural driver scabrous-GAL4. The average of 9 EL neurons per hemisegment is increased to an average of 16 eve-expressing lateral cells per hemisegment. Ectopic Runt expression causes a severe disruption of the nerve cord, as shown by the abnormal medial eve expression and severe disorganization of the axons. However, Runt is not sufficient to induce eve expression in the progeny of all the neuroblasts. Neuroblast 6-1 and/or neuroblast 6-2 must express another protein that is essential for Runt to activate eve expression. Using the marker Tau-green fluorescent protein to highlight the axons, it was found that the extra Even-skipped-expressing neurons project axons along the same pathway as the EL neurons. Runt is expressed in neuroblast 3-3, supporting an autonomous role for runt during neuroblast specification (Dormand, 1998).
Proteins expressed both by neuroblast 3-3 and by neuroblasts 6-1 or 6-2 are possible candidates for cofactors acting with Runt to induce EL neurons. Neuroblast 6-1 expresses the steroid receptor superfamily member Seven-up and neuroblast 6-2 expresses the zinc-finger transcription factor Ming (Castor) in common with neuroblast 3-3. Although Eve expression is not affected in castor mutants, it would be interesting to investigate whether either Ming or Seven-up contribute to other aspects of the EL neuron fate (Dormand, 1998).
The Drosophila CNS contains a variety of glia, including highly specialized glia that reside at the CNS midline and functionally resemble the midline floor plate glia of the vertebrate spinal cord. Both insect and vertebrate midline glia play important roles in ensheathing axons that cross the midline and secreting signals that control a variety of developmental processes. The Drosophila midline glia consist of two spatially and functionally distinct populations. The anterior midline glia (AMG) are ensheathing glia that migrate, surround and send processes into the axon commissures. By contrast, the posterior midline glia (PMG) are non-ensheathing glia. Together, the Notch and hedgehog signaling pathways generate AMG and PMG from midline neural precursors. Notch signaling is required for midline glial formation and for transcription of a core set of midline glial-expressed genes. The Hedgehog morphogen is secreted from ectodermal cells adjacent to the CNS midline and directs a subset of midline glia to become PMG. Two transcription factor genes, runt and engrailed, play important roles in AMG and PMG development. The runt gene is expressed in AMG, represses engrailed and maintains AMG gene expression. The engrailed gene is expressed in PMG, represses runt and maintains PMG gene expression. In addition, engrailed can direct midline glia to a PMG-like non-ensheathing fate. Thus, two signaling pathways and runt-engrailed mutual repression initiate and maintain two distinct populations of midline glia that differ functionally in gene expression, glial migration, axon ensheathment, process extension and patterns of apoptosis (Watson, 2011).
This paper describes how the Hh morphogen patterns the midline cells to generate two populations of MG with distinct functional properties. The key output of this signaling is the expression of en that imparts PMG cell fate, in part, by repressing runt. In turn, the runt gene maintains AMG fate by repressing en. Thus, morphogenetic signaling and transcriptional regulation lead to AMG and PMG with divergent molecular, morphological and functional differences (Watson, 2011).
At stage 10, the 16 midline cells per segment consist of three equivalence groups of neural precursors, four to six cells each. Notch signaling directs ten of these 16 cells to become MG; the remainder become MPs and the MNB. Thus, Notch represses neuronal development in MG and activates a core set of MG-expressed genes (e.g., wrapper). MG in the anterior of the segment become AMG; those in the posterior of the segment become PMG. Notch signaling by itself is unlikely to influence different MG fates, as expression of activated Suppressor of Hairless in midline cells drives all cells into a MG fate but does not affect their AMG or PMG patterns of gene expression. Thus, additional factors that can direct AMG and PMG cell fates were sought (Watson, 2011).
Previous work demonstrated that hh can pattern midline cells along the A/P axis, and, indeed, this study demonstrates that hh is required for PMG cell fate. The source of Hh is not in the midline, but in the lateral ectoderm in a stripe of cells, collinear with the pair of midline early en+ cells. Hh signals to midline cells posterior to the early en+ cells, inducing en in an additional six to seven cells. These late en+ cells plus the early en+ cells become about four PMG, as well as MP4-6 and the MNB. Misexpression and mutant analyses indicate that hh is required for all PMG gene expression and for repressing AMG expression. hh signaling probably has multiple target genes because hh is required for en and l(1)sc expression, but en does not regulate l(1)sc. Misexpression of hh can activate en expression in anterior MG, and both hh and en misexpression convert these cells functionally into non-ensheathing MG that resemble PMG, results also consistent with observations that ectopic expression of hh and en in midline cells affects AMG differentiation. However, neither hh nor en can activate all PMG gene expression in anterior MG, because neither activates masquerade (mas) expression in anterior MG. The mas gene is expressed transiently at stage 12 in a subset of PMG, suggesting that functionally distinct classes of PMG might exist. Expression of mas might require other signals in addition to hh that are absent in anterior MG (Watson, 2011).
runt is present in AMG and represses en and PMG-specific gene expression. In runt mutant cells that are runt- en+, expression of three genes expressed in only AMG (CG33275, Fhos and nemy) are absent and wrapper is reduced. This could be due to runt repression of en, repression of other genes or activation by runt. In runt mutant cells that are runt- en-, Fhos and nemy are present, wrapper is at high levels, but CG33275 expression is absent. This suggests that runt does not activate expression of Fhos, nemy and wrapper in AMG, but maintains their AMG levels by repressing en. By contrast, runt is required for expression of CG33275, possibly indicating a positive role for runt in AMG differentiation in addition to its repressive role in AMG maintenance. However, CG33275 is most prominently expressed in a subset of AMG closest to the commissures, and this AMG expression could be dependent on additional signals, perhaps from the developing axon commissure. Thus, absence of CG33275 expression in runt mutant embryos could alternatively be due to an effect of runt on developing axons or CNS development (Watson, 2011).
As most AMG gene expression is not dependent on runt, it is proposed that Notch signaling initially induces an AMG pattern of gene expression in all glia and, either simultaneously or soon after, Hh signaling in the posterior of the segment generates PMG. One important downstream target of Notch signaling is likely to be the sim gene, which encodes a bHLH-PAS protein that functions as a DNA-binding heterodimer with the Tango (Tgo) bHLH-PAS protein. During early development, sim is expressed in all midline primordia and is required for midline cell development. However, later in development, sim is restricted to MG and a subset of midline neurons. Genetically, sim expression is absent in embryos mutant for Notch signaling. The sim gene is likely to be an important aspect of MG transcription, because mutation of Sim-Tgo binding sites in the slit and wrapper MG enhancers results in loss of MG expression, and Sim-Tgo binding sites are present in other identified MG enhancers. The Hh morphogen transforms only posterior MG into PMG. It is unknown why hh does not affect anterior MG, but it is likely to be owing to the presence of unknown factors in these cells that inhibit hh signaling. Since Notch signaling, rather than runt, is primarily required for AMG gene expression, the key role of runt is probably to maintain AMG gene expression by repressing en. Similarly, en functions to maintain PMG gene expression by repressing runt, but also contributes positively to PMG cell fate, as en misexpression confers PMG-like function to AMG (Watson, 2011).
The most striking features of AMG are their ability to migrate around the commissures, ensheath them and extend processes into the axons. The function of PMG is unknown, but they are unable to ensheath the commissures, even though they are in close proximity. One of the major factors influencing AMG-axon interactions is Nrx-IV-Wrapper adhesion. Levels of wrapper expression in AMG are higher than in PMG, and this is likely to be a key determinant of why AMG ensheath commissures, and PMG do not, because loss of wrapper expression results in incomplete migration and ensheathment. Recent work has demonstrated that sim directly regulates wrapper expression, and spitz signaling from axons might also form a positive feedback loop to control wrapper levels and strengthen Nrx-IV-Wrapper interactions. As en genetically reduces wrapper levels in PMG, it will be interesting to determine if this regulation is direct or indirect. Although the control of wrapper levels is likely to be a major factor in AMG-PMG differences and the ability of glia to ensheath axons, other genes whose levels differ between AMG and PMG might also contribute. This illustrates why it will be important to identify target genes and understand better the roles that Notch//Suppressor of Hairless, sim, hh, Ci, en, runt and other MG transcription factors play in regulating MG gene expression and function (Watson, 2011).
The homeodomain (HD) protein Bicoid (Bcd) is thought to function as a gradient morphogen that positions boundaries of target genes via threshold-dependent activation mechanisms. This study analyzed 66 Bcd-dependent regulatory elements, and their boundaries were shown to be positioned primarily by repressive gradients that antagonize Bcd-mediated activation. A major repressor is the pair-rule protein Runt (Run), which is expressed in an opposing gradient and is necessary and sufficient for limiting Bcd-dependent activation. Evidence is presented that Run functions with the maternal repressor Capicua and the gap protein Kruppel as the principal components of a repression system that correctly orders boundaries throughout the anterior half of the embryo. These results put conceptual limits on the Bcd morphogen hypothesis and demonstrate how the Bcd gradient functions within the gene network that patterns the embryo (Chen, 2012).
This study identified 32 enhancers that respond to Bcd-dependent activation and form expression boundaries at different positions along the AP axis of fly embryos. Adding these elements to the 34 previously known enhancers constitutes the largest data set of in vivo-tested and -confirmed enhancers regulated by a specific transcription factor in all of biology (Chen, 2012).
The 32 confirmed enhancers were identified among 77 tested genomic fragments, which were selected because they showed in vivo-binding activity, or they conformed to a stringent homotypic-clustering model for predicted Bcd-binding sites, or both. All seven previously unknown fragments showing in vivo binding and a predicted site cluster directed Bcd-dependent transcription in the early embryo. Other fragments from the top 50 ChIP-Chip signals (which do not conform to the clustering model) were also very likely (21 of 26) to test positive in the in vivo test, but this likelihood drops significantly (9 of 25) in a set of fragments from lower on the list of ChIP-Chip fragments. Interestingly, of 19 tested fragments that contain clusters of predicted sites, but no in vivo binding activity, not a single one tested positive in vivo. These results suggest that in ;vivo binding assays are much better predictors of regulatory function than simple site-clustering algorithms alone (Chen, 2012).
One explanation for the failure of these predicted site clusters to bind Bcd in vivo is that they lie in heterochromatic regions of the genome that prevent site access. However, because they fail to function when taken out of their normal context (in reporter genes), whatever is preventing activation must be a property of the fragment itself and not its location in the genome. Interestingly, a number of Bcd site cluster-containing fragments drive expression later in development. It is proposed that these fragments fail to bind Bcd because they lack sites for cofactors that facilitate Bcd binding. In preliminary experiments it was observed that Bcd-activated fragments contain on average more binding sites for the ubiquitous activator protein Zelda (Zld) than those that fail to activate. Zld has been shown to be critical for timing the zygotic expression of hundreds of genes in the maternal to zygotic transition (Chen, 2012).
These results suggest strongly that a gradient of Run protein plays a major role in limiting Bcd-dependent activation. Run seems to work as part of a repression system that also includes Cic and possibly Kr. Expression boundaries in the region anterior to the presumptive cephalic furrow shift toward the posterior in run and cic mutants, and the double mutant causes boundaries that are normally well separated to collapse into a single position (Chen, 2012).
The use of multiple repressors permits flexibility in binding site architecture within enhancers that establish boundaries at similar positions. For example type I enhancers show overrepresentations of both Run and Cic sites, but 27% lack strong matches to the Cic PWM, and 12% lack strong matches to the Run PWM. Importantly, however, all type I enhancers lacking Cic sites contain Run sites, and those lacking Run sites contain Cic sites. Multiple Kr sites were observed in a large number of Bcd-dependent enhancers, which suggests that Kr is also a major component of the repression system that orders Bcd-dependent expression boundaries. Taken together, these data suggest that antagonistic repression of Bcd-mediated activation is a key design principle of the system that organizes the AP body plan. The repressors identified so far (Run, Cic, and Kr) are expressed in overlapping domains with gradients at different positions, consistent with the formation and ordering of a relatively large number of boundaries throughout the anterior half of the embryo (Chen, 2012).
The close linkage between repressor sites and Bcd sites within discrete enhancers suggests that repression occurs via short-range interactions that interfere directly with Bcd binding or activation. Interestingly, Cic also shows repressive effects that seem to be binding site independent. For example some type I enhancers do not contain recognizable Cic sites, but their expression boundaries expand posteriorly in cic mutants. This could be caused by the reduced expression of run and Kr in cic mutants. However, genetically removing both Kr and run causes a less dramatic expansion than that seen in the absence of cic. This suggests that Cic binds these enhancers via suboptimal sites or that it is required for the correct patterning of another unknown repressor. Another possibility is that these expansions are caused indirectly by changing the balance of MAPK phosphorylation events that control terminal patterning (Chen, 2012).
These results do not strictly falsify the Bcd morphogen hypothesis, but they support the idea that the Bcd gradient can establish only a 'rough framework that is elaborated by the interaction of the zygotic segmentation genes'. What is the nature of this framework, and what role does it play in the network that precisely positions target gene boundaries (Chen, 2012)?
One component of the system, the Cic repression gradient, is maternally produced and formed by downregulation at the poles via the terminal patterning system. This gradient is formed independently of Bcd but is critical for establishing boundaries of Bcd-dependent target genes. In contrast, Bcd is involved in activating the expression patterns of run and Kr and in repressing them in anterior regions. Both run and Kr expand anteriorly in bcd mutants. There is no evidence that Bcd functions directly as a transcriptional repressor, so these repressive activities are probably indirect. Previous work showed that the Bcd target gene gt is involved in setting the anterior Kr boundary, and it is hypothesized that another Bcd target gene, slp1, encodes a forkhead domain (FKH) protein that sets the anterior boundary of the early run pattern. slp1 is expressed in a pattern reciprocal to the run pattern and was previously shown to position the anterior boundaries of several pair-rule gene stripes including run stripe 1 (Chen, 2012).
These results suggest that a major function of the Bcd gradient is the differential positioning of two repressors, Slp1 and Gt, which set the positions of the Run and Kr repression gradients, which then feedback to repress Bcd-dependent target genes. How are slp1 and gt differentially positioned? One possibility is that slp1 and gt enhancers respond to specific concentrations within the Bcd gradient, consistent with the original model for morphogen activity. However, the fact that the slp1 and gt expression domains form boundaries at the same positions in embryos lacking the Cic and Run repressors argues against this model for these genes (Chen, 2012).
It was also shown that Bcd target genes normally expressed in cephalic regions form and correctly position posterior boundaries in embryos containing flattened Bcd gradients. Run is still expressed in these embryos, specifically in a domain that consistently abuts the boundaries of the anterior Bcd target genes, regardless of copy number. This suggests that a mutually repressive interaction between Slp1 and Run is maintained in these embryos but does not explain how these boundaries are consistently oriented perpendicularly to the AP axis. The answer might lie in the fact that the flattened Bcd gradients in these embryos are not completely flat but are present as shallow gradients with slightly higher levels in anterior regions. In these embryos the slight changes in concentration along the AP axis might cause a bias that enables the orientation of the mutual repression interaction. In wild-type embryos, Bcd is much more steeply graded, which makes this bias stronger and the boundary between these mutual repressors more robust (Chen, 2012).
These results suggest that antagonistic repression precisely orders Bcd-dependent expression boundaries. However, repression may not be required for the activity of all morphogens. For example the extracellular signal activin has been shown to activate target genes in a threshold-dependent manner in isolated animal caps from frog embryos. Also, a gradient of the transcription factor Dorsal (Dl) is critical for setting boundaries between different tissue types along the dorsal-ventral (DV) axis of the fly embryo. It is thought that the major mechanism in Dl-specific patterning is threshold-dependent activation, which is quite different from the system described in this paper. One major difference between Bcd and Dl is the number of boundaries specified: three for Dl and more than ten for Bcd. It is proposed that the robust ordering of more boundaries simply requires a more complex system (Chen, 2012).
In general, though, it seems that antagonistic mechanisms are involved in controlling the establishment or interpretation of most morphogen activities. For example in the Drosophila wing disc, the TGF-N2 signal Dpp forms an activity gradient that is refined by interactions with multiple extracellular factors. Also, in vertebrates the signaling activity of the extracellular morphogen Sonic hedgehog (Shh) is affected by positive and negative interactions with specific molecules on the surfaces of receiving cells (Chen, 2012).
There is some evidence that transcriptional repression is also used for refining the patterning activities of extracellular molecules. Dpp acts as a long-range morphogen that activates two major target genes (optomotor blind [omb] and spalt [sal]) in nested patterns with boundaries at different positions with respect to the source of Dpp. Although these boundaries could in theory be formed by differential responses to the morphogen, it is clear that the transcriptional repressor Brinker (Brk), which is expressed in an oppositely oriented gradient, also plays an important role. The Brk gradient is itself positioned by Dpp activity in a manner analogous to positioning of the Run and Kr repressor gradients by Bcd. Also, a similar transcriptional network functions in Shh-mediated patterning of the vertebrate neural tube, where a series of spatially oriented repressors feeds back to limit the expression boundaries of Shh-mediated cell fate decisions (Chen, 2012).
Conceptually, these more complex systems are reminiscent of the reaction-diffusion model proposed by Turing, in which a localized activator would activate a repressor, which would diffuse more rapidly than the activator, and feed back on its activity. These systems strongly suggest that the patterning activity of a single monotonic gradient is insufficiently robust for establishing precise orders of closely positioned expression boundaries. By integrating gradients with repressive mechanisms that refine gradient shape or influence outputs, systems are generated that ensure consistency in body plan establishment while still maintaining the flexibility required for complex systems to evolve (Chen, 2012).
Bases in 5' UTR -251
Exons - two
Bases in 3' UTR - 621
The absence of an identifiable transcription factor motif (e.g., homeo box, zinc finger, leucine zipper, or helix-loop-helix) makes Runt different from the other early-acting segmentation proteins. The subcellular location of the protein is in the nucleus (Kania, 1990).
A highly conserved region in the Runt protein is termed the Runt domain. The functional properties of the Runt domain from the D. melanogaster gene and the human AML1 (acute myeloid leukemia 1) gene were compared. The different DNA binding properties of Runt and AML1 are due to differences within their respective Runt domains. Proteins containing the AML1 Runt domain function in Drosophila embryos, but sequences outside of the runt domain are important in vivo (Pepling, 1995).
date revised: 7 October 2021
Home page: The Interactive Fly © 1995, 1996 Thomas B. Brody, Ph.D.
The Interactive Fly resides on the
Society for Developmental Biology's Web server.