The distribution of Eld is ubiquitous in early embryos, showing no hint of a striped pattern. It is also ubiquitious in wing discs. Although the protein is present everywhere in eye discs, its strongest expression occurs in a band just anterior to the morphogenetic furrow, in the postion where cells respond to Hedgehog and Decapentaplegic signaling (Treisman, 1997).

Effects of mutation or deletion

Very few eyelid mutant cells are found when clones mutant for eld are analyzed in adult eyes. However, such clones are associated with scars, suggesting that eld mutant cells are present at a certain stage but interfer with normal develpment. The eld mutant clones are relatively small compared with clones mutant for other genes, suggesting that eld is required for cell proliferation and/or survival. Exceptions are clones that include the posterior margin of the disc, which were frequently much larger than internal clones. The difference might lie in the fact that development at the posterior margin is driven by dpp whereas the internal propagation of the furrow depends on hedgehog (Treisman, 1997).

eld also has an affect on neuronal differentiation. Most photoreceptor clusters that form within eld clones contain fewer neuronal cells than normal; therefore, there may be a partial, though not absolute, requirement for eld for neuronal differentiation. The lack of differentiation is not simply attributable to poor cell viability, as all the cells in eld clones can be induced to differentiate as neurons by removing the function of Enhancer of split complex genes. The block to differentiation caused by loss of eld function in the eye resembles the effect of loss of shaggy or ectopic expression of wingless (Treisman, 1997).

Clones of eld mutant cells induced in the wing disc also produce pattern alterations suggestive of antagonism to wingless. One effect of clones produced early in development is the transformation of the posterior notum into a partial second wing. These wings have a reversed anterior-posterior polarity; their most clearly differentiated structure is an alula produced consistently at their anterior margin. This transformation is the reverse of that produced by the wingless1 mutation, which transforms the wing into a duplicated notum, and is similar to that produced by overexpressing wingless, decapentaplegic or optomotor-blind in the notum. Clones induced later in wing development are associated with ectopic wing margin bristles. Many or all of these ectopic bristles are not mutant for eld but they are sometimes seen to form adjacent to eld clones. Ectopic bristle formation is restricted to the dorsal surface of the wing within the anterior compartment, and is observed most commonly near the wing margin in tufts of the bristle type appropriate to their postion along the anterior-posterior axis. Ectopic wing margin bristles are also produced in clones mutant for shaggy. However, shaggy clones show neither the non-autonomy nor the positional restrictions observed for eld clones. These results suggest a cell non-autonomous role for eld in wing patterning (Treisman, 1997).

The activity of the E2F transcription factor is regulated in part by pRB, the protein product of the retinoblastoma tumor suppressor gene. Studies of tumor cells show that the p16ink4a/cdk4/cyclin D/pRB pathway is mutated in most forms of cancer, suggesting that the deregulation of E2F, and hence the cell cycle, is a common event in tumorigenesis. Extragenic mutations that enhance or suppress E2F activity are likely to alter cell-cycle control and may play a role in tumorigenesis. An E2F overexpression phenotype in the Drosophila eye was use to screen for modifiers of E2F activity. Coexpression of dE2F and its heterodimeric partner dDP in the fly eye induces S phases and cell death. Thirty three enhancer mutations of this phenotype were isolated by EMS and X-ray mutagenesis and by screening a deficiency library collection. The majority of these mutations sorted into six complementation groups, five of which have been identified as alleles of brahma (brm), moira (mor), osa, pointed (pnt), and polycephalon (poc). osa, brm, and mor encode proteins with homology to SWI1, SWI2, and SWI3, respectively, suggesting that the activity of a SWI/SNF chromatin-remodeling complex has an important impact on E2F-dependent phenotypes. Mutations in poc also suppress phenotypes caused by p21CIP1 expression, indicating an important role for Polycephalon in cell-cycle control (Staehling-Hampton, 1999).

The molecular basis of the interaction between E2F and a BRM/MOR/OSA chromatin-remodeling complex is not yet clear and a range of possibilities exists. The genetic interaction may result from a direct physical interaction between RBF/E2F complexes and chromatin-remodeling machinery. In support of this idea the human homologs of BRM, hBRM, and BRG1 have been found to physically associate with pRB. This raises the possibility that BRM/MOR/OSA may help E2F/RBF repressor complexes bind to their target sites. This interpretation is supported by experiments from Trouche and co-workers who used transient transfection of mammalian cells to demonstrate that BRG1 can cooperate with pRB to repress E2F-dependent transcription (Trouche, 1997). Consistent with this model, the introduction of two copies of GMR-RBF into a GMR-dE2FdDPp35/+; brm-/+ background suppresses the enhancement by brm. Thus the effect caused by low levels of brm can be overcome by increasing the dosage of RBF. Additional evidence has been sought that would be predicted by this model; to date, however, these experiments have been inconclusive. BRM lacks the LXCXE motifs found in hBRM and BRG1, which have been suggested to mediate the interaction with pRB. To date no physical interaction between BRM and RBF or between BRM and dE2F has been detected. The interaction between endogenous pRB and hBRM or BRG1 proteins is hard to detect even in mammalian cells, and the failure to find BRM/RBF complexes may simply reflect difficulty in extracting chromatin-associated proteins under conditions that maintain the interaction (Staehling-Hampton, 1999).

An alternative possibility is that the BRM/MOR/OSA chromatin-remodeling complex is an important regulator of the expression of some key E2F-target genes, but this complex does not interact directly with either RBF or E2F. In this case the functional interaction occurs because these proteins converge on overlapping sets of promoters. This model is difficult to test because it is not yet clear which, and how many, E2F target genes are functionally significant. RNR2, one example of an E2F-dependent gene, is expressed normally in embryos mutant for brm, osa, or mor; no change in the expression of RNR2 in GMR-dE2FdDPp35 eye disks heterozygous for brm, osa, or mor alleles could be detected. While RNR2 expression is often used to provide an in vivo readout of E2F activity, experiments suggest that it is not a critical E2F target. The effects of brm, mor, and osa may only be evident at a subset of E2F-regulated promoters and an extensive screen of E2F targets will be necessary to find the appropriate gene (Staehling-Hampton, 1999).

It is possible that E2F and brm act in distinct pathways that influence cell-cycle progression. In this model the activity of a BRM/MOR/OSA-containing complex may have a function that influences the ability of E2F or RBF to control S-phase entry. Several observations have linked BRM-related proteins to cell-cycle control. brm null clones in the adult cuticle often show duplications of bristle structures, suggesting a possible role for brm in proliferation, and mice lacking the BRM homolog SNF2alpha show evidence of increased cell proliferation. Although brm, mor, and osa have no effect on the GMR-p21 phenotype, both brm and mor mutations have been isolated as suppressors of a hypomorphic cyclin E eye phenotype, demonstrating that brm and mor can affect other cell-cycle phenotypes in the eye. Other studies have shown that the activity of hSWI/SNF complexes is itself cell-cycle regulated. Transformation by activated Ras decreases the expression of the murine ortholog of hBRM in mouse fibroblasts, whereas growth arrest leads to an accumulation of protein. Recently, BRG1 and BAF155, a human ortholog of Moira, have been shown to associate with cyclin E and are suggested to be targets for cyclin E-dependent kinases during S-phase entry (Staehling-Hampton, 1999 and references).

During this study it was observed that GMR-dE2FdDP p35/+; brm-/+ eyes develop necrotic patches that increase in severity with the age of the adult fly. This raised the possibility that brm mutations might enhance the phenotype by promoting E2F-induced apoptosis. However, further experiments have failed to support this hypothesis. brm mutations fail to enhance the GMR-dE2FdDP phenotype, which has elevated levels of apoptosis, or to modify a GMR-rpr phenotype. In addition, brm mutations have no effect on the phenotype of animals in which GMR-rpr and GMR-hid-induced apoptosis is blocked by GMR-p35. No increase in the number of apoptotic cells is detected when GMR-dE2FdDPp35/+; brm-/+ third instar eye disks are stained with acridine orange (Staehling-Hampton, 1999).

Effects of Mutation

Ectopic expression of DREF induces DNA synthesis, apoptosis, and unusual morphogenesis in the Drosophila eye imaginal disc: possible interaction with Polycomb and trithorax group proteins

The promoters of Drosophila genes encoding DNA replication-related proteins contain transcription regulatory element DRE (5'-TATCGATA) in addition to E2F recognition sites. A specific DRE-binding factor, DNA replication-related element factor or DREF, positively regulates DRE-containing genes. In addition, it has been reported that DREF can bind to a sequence in the hsp70 scs' chromatin boundary element that is also recognized by boundary element-associated factor, and thus DREF may participate in regulating insulator activity. To examine DREF function in vivo, transgenic flies were established in which ectopic expression of DREF was targeted to the eye imaginal discs. Adult flies expressing DREF exhibited a severe rough eye phenotype. Expression of DREF induces ectopic DNA synthesis in the cells behind the morphogenetic furrow that are normally postmitotic, and abolishes photoreceptor specifications of R1, R6, and R7. Furthermore, DREF expression caused apoptosis in the imaginal disc cells in the region where commitment to R1/R6 cells takes place, suggesting that failure of differentiation of R1/R6 photoreceptor cells might cause apoptosis. The DREF-induced rough eye phenotype is suppressed by a half-dose reduction of the E2F gene, one of the genes regulated by DREF, indicating that the DREF overexpression phenotype is useful to screen for modifiers of DREF activity. Among Polycomb/trithorax group genes, it was found that a half-dose reduction of some of the trithorax group genes involved in determining chromatin structure or chromatin remodeling (brahma, moira, and osa) significantly suppresses and that reduction of Distal-less enhances the DREF-induced rough eye phenotype. The results suggest a possibility that DREF activity might be regulated by protein complexes that play a role in modulating chromatin structure. Genetic crosses of transgenic flies expressing DREF to a collection of Drosophila deficiency stocks allowed identification of several genomic regions, deletions of which caused enhancement or suppression of the DREF-induced rough eye phenotype. These deletions should be useful to identify novel targets of DREF and its positive or negative regulators (Hirose, 2001).

Enhancer-promoter communication mediated by Chip during Pannier-driven proneural patterning is regulated by Osa

The GATA factor Pannier activates proneural achaete/scute (ac/sc) expression during development of the sensory organs of Drosophila through enhancer binding. Chip bridges Pannier with the (Ac/Sc)-Daughterless heterodimers bound to the promoter and facilitates the enhancer-promoter communication required for proneural development. This communication is regulated by Osa, which is recruited by Pannier and Chip. Osa belongs to Brahma chromatin remodeling complexes, and this study shows that Osa negatively regulates ac/sc. Consequently, Pannier and Chip also play an essential role during repression of proneural gene expression. This study suggests that altering chromatin structure is essential for regulation of enhancer-promoter communication (Heitzler, 2003).

ChipE is a viable allele of Chip that is associated with a point mutation in the LIM-interacting domain (LID), which specifically reduces interaction with the bHLH proteins Ac, Sc, and Da. As a consequence, the ChipE mutation disrupts the functioning of the proneural complex encompassing Chip, Pnr, Ac/Sc, and Da. A homozygous ChipE mutant shows thoracic cleft and loss of the DC bristles, similar to loss of function pnr alleles (Heitzler, 2003).

To identify new factors that regulate this proneural complex, a screen was performed for second-site modifiers of the ChipE phenotypes. One allele of osa (osaE17) was found among the putative mutants. OsaE17 corresponds to a loss-of-function allele, and homozygous embryos die with normal cuticle patterning. Both osaE17 and null alleles of osa (osa616 or osa14060) enhance the cleft but suppress the loss of DC bristle phenotypes of ChipE flies. Indeed, ChipE flies with only one copy of osa+ (ChipE;osa616/+) are weak and sterile but show wild-type DC bristle pattern (Heitzler, 2003).

These genetic interactions suggest that Osa can antagonize the function of Pnr. Moreover, overexpressed Osa (+/UAS-osa;Gal4-pnrMD237/+) induces a thoracic cleft and the loss of DC bristles similar to the loss-of-function pnr alleles. In contrast, loss-of-function osa alleles display an excess of DC bristles similar to overexpressed Pnr. For example, (osa14060/+), (osa616/+), and (osaE17/+) flies exhibit respectively 2.35 ± 0.12, 2.38 ± 0.12, and 2.43 ± 0.17 DC bristles per heminotum (Oregon wild-type flies have 2.00 DC bristles/heminotum). Furthermore, transallelic combination of osa14060 with the hypomorphic osa4H (osa4H/osa14060) accentuates the excess of DC bristles compared with (osa14060/+). (osa4H/osa14060) flies display 4.17 ± 0.19 DC bristles per heminotum. In contrast, (osa4H/osa4H) flies display 2.50 ± 0.11 DC bristles per hemithorax. The development of the extra DC bristles revealed by phenotypic analysis was compared with the positions of the DC bristle precursors detected with a LacZ insert, A101, in the neuralized gene that exhibits staining in all sensory organs. In (osa14060/osa4H) discs, additional DC precursors are observed that lead to the excess of DC bristles. The pnrD alleles encode Pnr proteins carrying a single amino acid substitution in the DNA binding domain that disrupts interaction with the U-shaped (Ush) antagonist. Consequently, PnrD constitutively activates ac/sc, leading to an excess of DC bristles. This excess is accentuated when osa function is simultaneously reduced (pnrD1/osa616) (Heitzler, 2003).

Since osa shows genetic interactions with trithorax group genes encoding components of the Brm complex like moira (mor) and brm, whether mutations in mor and brm suppress the ChipE phenotype was investigated. Loss of one copy of brm+ in (ChipE; brm2/+) flies suppresses the lack of DC bristles observed in ChipE flies, similar to loss of one copy of osa+. This shows that brm and osa both act during Pnr-dependent patterning, in agreement with the fact that they have been shown to be associated in the Brm complex. In contrast, reducing the amount of Mor by half [(ChipE;mor1/+) flies] is not sufficient to modify the ChipE phenotype. This does not definitely exclude the possibility that mor is directly or indirectly involved, via the Brm complex, in Pnr-dependent patterning (Heitzler, 2003).

The complete osa open reading frame of 2715 amino acids and the intronic splicing signals were PCR amplified from genomic DNA prepared from homozygous embryos (osaE17 and osa14060) and homozygous first instar larvae (osa4H). For osa14060 and osa4H, the sequence analysis revealed a single mutation in the N terminus that causes a glutamine to stop codon substitution. The conceptual translation of osa14060 leads to a truncated Osa protein lacking both functional domains, whereas Osa4H retains the ARID domain but lacks the C-terminal EHD. Wild-type osa function is necessary for patterning of the DC bristles. Although osaE17 behaves as a stronger allele than osa14060 and osa4H, molecular identity of the mutation is unknown. Hence, the osaE17 phenotype may result from a mutation in regulatory sequences that affects osa expression (Heitzler, 2003).

It has been shown that a complex containing Pnr, Chip, and the (Ac/Sc)-Da heterodimer activates proneural expression in the DC proneural cluster and promotes development of the DC macrochaetae. Osa and Pnr/Chip have antagonistic activities during development because loss of osa function (osa4H and osa14060) displays additional DC bristles. However, since the current study reveals that osa genetically interacts with pnr and Chip, it was asked whether Osa physically interacts with the Pnr and Chip proteins. Immunoprecipitations of protein extracts made from Cos cells cotransfected with expression vectors for tagged Osa and either Pnr or tagged Chip were immunoprecipitated. Because Osa is a large protein, several expression vectors encoding contiguous domains of Osa were used. Osa coimmunoprecipitates with Pnr and Chip and can be detected on Western blots with appropriate antibodies. The interactions appear to require the overlapping domains Osa E (His1733/Glu2550) and Osa F (Ala2339/Ala2715) corresponding to the EHD. Enhancer-promoter communication during proneural activation and development of the DC bristles requires regulatory sequences scattered over large distances and appears to be negatively regulated by interaction of Pnr and Chip with Osa through the EHD. Interestingly, the EHD is not conserved in yeast. In yeast, the UAS sequences are generally close to the promoter and there is no requirement for long-distance interactions. This observation could support the idea that the EHD is essential for long-distance enhancer-promoter communication. Alternatively, yeast may just lack proteins like Chip or Pnr (Heitzler, 2003).

The DNA-binding domain and the C-terminal region are essential for the function of Pnr during development of the DC sensory organs. The pnrVX1 and pnrVX4 alleles (collectively pnrVX1/4) are characterized by frameshift deletions that remove two C-terminal alpha-helices and result in reduced proneural expression and loss of DC bristles (Heitzler, 2003).

The molecular interactions between Osa and PnrD1 and between Osa and PnrVX1 were investigated. PnrD1 protein interacts with the EHD as efficiently as wild-type Pnr. In contrast, the physical interaction is disrupted when the C terminus of Pnr encompassing the alpha-helices is removed. Because the C terminus of Pnr is required for the Pnr-Osa interaction in transfected cells extracts, the abilities of in vitro translated 35S-labeled Osa domains to bind to GST-CTPnr attached to glutathione-bearing beads were investigated. Only Osa E and Osa F interact with the C terminus of Pnr. The interaction between Chip and Osa, and it was found that Osa associates with the N-terminal homodimerization domain of Chip, also required for the interaction between Chip and Pnr, was investigated. Furthermore, Osa E and Osa F also bind to immobilized GST-Chip. Deletion of the alpha helix H1 disrupts the interactions between Pnr and Osa. Interestingly, the same deletion also disrupts the interaction with Chip. Therefore, the functional antagonism between Chip and Osa during neural development may result from a competition between these proteins for association with Pnr. Alternatively, the deletion of H1 may affect the overall structure of the C terminus of Pnr and disrupt the physical interactions with Chip and Osa. To discriminate between these hypotheses, immunoprecipitations of protein extracts containing a constant amount of Pnr, a constant amount of the tagged Osa E domain, and increasing concentrations of Chip were performed. Pnr immunoprecipitates with immunoprecipitated tagged Osa E and the amount of Pnr immunoprecipitated increases in the presence of increasing concentrations of Chip. The presence of increasing amounts of Chip does not inhibit the Osa-Pnr interaction as would be expected if Osa and Chip were to compete for binding to Pnr. In contrast, it suggests that Chip and Pnr act together to recruit Osa and to target its activity and possibly the activity of the Brm complex to the ac/sc promoter sequences (Heitzler, 2003).

Using expression vectors encoding contiguous domains of Osa, it was shown that the EHD of Osa mediates interactions with Pnr and Chip. Because the EHD is lacking in the truncated Osa14060 and Osa4H, it is hypothesized that the loss of interaction with Pnr and Chip are responsible for the excess of DC bristles observed in osa4H and osa14060 (Heitzler, 2003).

To investigate whether these interactions between Osa, Pnr, and Chip function in vivo during DC bristle development, the effects of both loss of function and overexpression of osa were examined on the activity of a LacZ reporter whose expression is driven by a minimal promoter sequence of ac fused to the DC enhancer (transgenic line DC:ac-LacZ). It was found that expression of the LacZ transgene is increased in osa14060/osa4H wing discs when compared with the wild-type control. For overexpression experiments, the UAS/GAL4 system was used, using as a driver the pnrMD237 strain that carries a GAL4-containing transposon inserted in the pnr locus (driver: pnr-Gal4). This insert gives an expression pattern of Gal4 indistinguishable from that of pnr. It was found that overexpressed Osa leads to a strong reduction of LacZ staining in the DC area, consistent with the lack of DC bristles. Thus, overexpressed Osa represses activity of the ac promoter sequences required for DC ac/sc expression and development of the DC bristles. It has been previously reported that wingless expression is also required for patterning of the DC bristles. However, the repressing effect of Osa on development of the DC bristles is unlikely to be the result of an effect of Osa on wingless expression because overexpressed Osa driven by pnrMD237 has no effect on the expression of a LacZ reporter inserted into the wingless locus. Thus, Osa acts through the DC enhancer of the ac/sc promoter sequences to repress ac/sc and neural development (Heitzler, 2003).

ChipE disrupts the enhancer-promoter communication and strongly affects expression of the LacZ reporter driven by the ac promoter linked to the DC enhancer. Because null alleles of osa suppress the loss of DC bristles displayed by ChipE, the consequences of reducing the dosage of osa was examined in ChipE flies. The expression of the LacZ reporter is not affected in ChipE flies when Osa concentration is simultaneously reduced (Heitzler, 2003).

In conclusion, Pnr function during proneural patterning is regulated by interaction with several transcription factors. Pnr function is negatively regulated by Ush, which interacts with its DNA-binding domain. Chip associates with the C terminus of Pnr, bridging Pnr at the DC enhancer with the AC/Sc-Da heterodimers bound at the proneural promoters, thus activating proneural gene expression. The current study reveals that Pnr function can also be regulated by interaction with Osa. Thus, Osa activity is specifically targeted to ac/sc promoter sequences and the binding of Osa therefore has a negative effect on Pnr function, leading to reduced expression of the proneural ac/sc genes. Osa belongs to Brm complexes, which are believed to play an essential role during chromatin remodeling necessary for gene expression. For example, in vitro transcription experiments with nucleosome assembled human beta-globin promoters have shown that the BRG1 and BAF155 subunits of the mammalian SWI/SNF homolog are essential to target chromatin remodeling and promote transcription initiation mediated by GATA-1. In contrast to what was observed in vitro, the current results suggest that in vivo the SWI/SNF complexes can also act to remodel chromatin in a way that represses transcription. Alternatively, the observed repression of proneural genes may simply define a novel function of Osa, independent of chromatin remodeling (Heitzler, 2003).

Osa modulates the expression of Apterous target genes in the Drosophila wing

The establishment of the dorsal-ventral axis of the Drosophila wing depends on the activity of the LIM-homeodomain protein Apterous. Apterous activity depends on the formation of a higher order complex with its cofactor Chip to induce the expression of its target genes. Apterous activity levels are modulated during development by dLMO (Beadex). Expression of dLMO in the Drosophila wing is regulated by two distinct Chip dependent mechanisms. Early in development, Chip bridges two molecules of Apterous to induce expression of dLMO in the dorsal compartment. Later in development, Chip, independently of Apterous, is required for expression of dLMO in the wing pouch. A modular P-element based EP (enhancer/promoter) misexpression screen was conducted to look for genes involved in Apterous activity. Osa, a member of the Brahma chromatin-remodeling complex, was found to be a positive modulator of Apterous activity in the Drosophila wing. Osa mediates activation of some Apterous target genes and repression of others, including dLMO. Osa has been shown to bind Chip. It is proposed that Chip recruits Osa to the Apterous target genes, thus mediating activation or repression of their expression (Milan, 2004).

This study presents evidence that Osa, a member of a subset of Brahma chromatin remodeling complexes, behaves overall as a general activator of Apterous activity in the Drosophila wing. Overexpression of Osa rescues and loss of Osa enhances the Beadex1 phenotype. It does so by modulating the expression levels of Apterous target genes, some of them being activated (e.g. Serrate and probably other unknown target genes) and some repressed (e.g. Delta, fringe). Chip has been shown to bind Osa. The fact that Osa has different effects on the transcription of Apterous target genes suggests that Chip recruits Osa to the promoters and in combination with other unknown factors mediates either transcriptional repression or activation. Osa mediates repression of both Apterous dependent and independent expression of fringe, suggesting a direct and probably Chip independent effect of Osa on fringe transcription (Milan, 2004).

Apterous activity is regulated during development by dLMO. Osa is required to mediate repression of dLMO expression. Since both early and late expression of dLMO depend on Chip, it is postulated that Chip forms a transcriptional complex with Apterous in the D compartment and an unknown transcription factor expressed in the wing pouch. Osa may interact with Chip thus recruiting the Brahma complex to the dLMO locus and remodeling chromatin in a way that limits dLMO transcriptional activation. High levels of dLMO protein reduce Apterous activity and the Notch dependent organizer is not properly induced along the DV boundary. Osa mediated repression of dLMO expression may ensure moderate levels of expression of dLMO in the wing, thus allowing proper wing development. Gain of function mutations that cause misexpression of vertebrate LMO proteins have been implicated in cancers of the lymphoid system. Truncating mutations in the human SWI-SNF complex, the human homologues of the Brahma complex, cause various types of human cancers. The SWI-SNF complex may be required to mediate repression of LMO expression in lymphoid tissues. Thus, it would be very interesting to analyze if truncating mutations in members of the human SWI-SNF complex cause higher levels of LMO expression and are associated with lymphoid malignancies (Milan, 2004).

It has been shown that the Brahma complex plays a general role in transcription by RNA Polymerase II. Then, is Osa having a general effect on the expression levels of every gene involved in wing patterning? Several observations indicate this is not the case. (1) Osa is a component of a subset of Brahma (Brm) chromatin complexes. (2) Brahma and Polycomb were shown to have non-overlapping binding patterns in polytenic chromosomes. Those genes involved in wing patterning and regulated by Polycomb (i.e. Hedgehog) may not be affected by overexpression of Osa. (3) Overexpression of Osa has different effects on the expression levels of Serrate, Delta and fringe. (4) Osa has been shown to specifically regulate the expression of Wingless target genes and the Achaete-scute complex genes, interestingly by restricting their expression levels (Milan, 2004).

Gene regulatory networks controlling hematopoietic progenitor niche cell production and differentiation in the Drosophila lymph gland

Hematopoiesis occurs in two phases in Drosophila, with the first completed during embryogenesis and the second accomplished during larval development. The lymph gland serves as the venue for the final hematopoietic program, with this larval tissue well-studied as to its cellular organization and genetic regulation. While the medullary zone contains stem-like hematopoietic progenitors, the posterior signaling center (PSC) functions as a niche microenvironment essential for controlling the decision between progenitor maintenance versus cellular differentiation. This study used PSC-specific GAL4 driver and UAS-gene RNAi strains, to selectively knockdown individual gene functions in PSC cells. The effect of abrogating the function of 820 genes was assessed as to their requirement for niche cell production and differentiation. 100 genes were shown to be essential for normal niche development, with various loci placed into sub-groups based on the functions of their encoded protein products and known genetic interactions. For members of three of these groups, loss- and gain-of-function phenotypes were characterized. Gene function knockdown of members of the BAP chromatin-remodeling complex resulted in niche cells that do not express the hedgehog (hh) gene and fail to differentiate filopodia believed important for Hh signaling from the niche to progenitors. Abrogating gene function of various members of the insulin-like growth factor and TOR signaling pathways resulted in anomalous PSC cell production, leading to a defective niche organization. Further analysis of the Pten, TSC1, and TSC2 tumor suppressor genes demonstrated their loss-of-function condition resulted in severely altered blood cell homeostasis, including the abundant production of lamellocytes, specialized hemocytes involved in innate immune responses. Together, this cell-specific RNAi knockdown survey and mutant phenotype analyses identified multiple genes and their regulatory networks required for the normal organization and function of the hematopoietic progenitor niche within the lymph gland (Tokusumi, 2012).

The discovery of a stem cell-like hematopoietic progenitor niche in Drosophila represents a significant contribution of this model organism to the study of stem cell biology and blood cell development. Extensive findings support the belief that the PSC functions as the niche within the larval lymph gland, with this cellular domain essential to the control of blood cell homeostasis within this hematopoietic organ. Molecular communication between the PSC and prohemocytes present in the lymph gland medullary zone is crucial for controlling the decision as to maintaining a pluri-potent progenitor state versus initiating a hemocyte differentiation program. This lymph gland cellular organization and the signaling pathways controlling hematopoieis therein have prompted several researchers in the field to point out its functional similarity to the HSC niche present in mammalian (Tokusumi, 2012).

As a means to discover new information on genetic and molecular mechanisms at work within a hematopoietic progenitor niche microenvironment, an RNAi-based loss-of-function analysis was carried out to selectively eliminate individual gene functions in PSC cells. The effect of knocking-down the function of 820 lymph gland-expressed genes was assessed as to their requirement for niche cell production and differentiation, and 100 of these genes were shown to be required for one or more aspects of niche development. The distinguishable phenotypes observed in these analyses included change in number of Hh-expressing cells, change in number of Antp-expressing cells, scattered and disorganized niche cells, rounded cells lacking extended filopodia, and lamellocyte induction in the absence of a normal PSC. The genes were placed into sub-groups based on their coding capacity and known genetic interactions, and the phenotypes associated with the functional knockdown of members of three of these gene regulatory networks were characterized (Tokusumi, 2012).

Previous studies have demonstrated that the PSC-specific ablation of srp function resulted in a lack of expression of the crucial Hh signaling molecule in these cells, the inactivity of the hh-GFP transgene in the niche, failure of niche cells to properly differentiate filopodial extensions, and the loss of hematopoietic progenitor maintenance coupled with the abundant production of differentiated hemocytes. Thus it was intriguing when it was observed that RNAi function knockdown of several members of the BAP chromatin-remodeling complex resulted in the identical phenotypes of lack of hh-GFP transgene expression and absence of filopodia formation in PSC cells. A convincing functional interaction was observed between srp encoding the hematopoietic GATA factor and osa encoding the DNA-binding Trithorax group protein in the inability of niche cells to express hh-GFP in double-heterozygous mutant lymph glands. Thus one working model is that the BAP chromatin-remodeling complex establishes a chromatin environment around and within the hh gene that allows access of the Srp transcriptional activator to the PSC-specific enhancer, facilitating Hh expression in these cells. It will be of interest to determine if there exists a direct physical interaction between Osa and Srp in this positive regulation of hh niche transcription and if so, what are the functional domains of the proteins essential for this critical regulatory event in progenitor cell maintenance. It is also likely that these functional interactions are important for Srp's transcriptional regulation of additional genes needed for the formation of niche cell filapodia (Tokusumi, 2012).

In this study, a total of 33 gain- or loss-of-function genetic conditions were analyzed that enhanced or eliminated the function of various positive or negatively-acting components of the insulin-like growth factor and TOR signaling pathways. A conclusion to be drawn from these analyses is that genetic conditions that have an end effect of enhancing translation activity and protein synthesis result in supernumerary PSC cell numbers in disorganized niche domains, while conditions that promote growth suppression lead to substantially reduced populations of niche cells. The same conclusion was obtained from recent studies performed by Benmimoun (2012). The Wg and Dpp signaling pathways have also been shown to be important for the formation of a PSC niche of normal size and function, and it is possible that the insulin-like growth factor and TOR signaling networks regulate the translation of one or more members of the Wg and/or Dpp pathways. These analyses have also shown that mutation of the Pten, TSC1, and TSC2 tumor suppressor genes results in severely altered blood cell homeostasis in lymph glands and in circulation, including the prolific induction of lamellocytes. A recent report demonstrated that in response to larval wasp infestation, the PSC secretes the Spitz cytokine signal, which triggers an EGFR-mediated signal transduction cascade in the generation of dpERK-positive lamellocytes in circulation. As dpERK activity is known to inhibit TSC2 function, inactivation of the TSC complex may be a downstream regulatory event leading to robust lamellocyte production in larvae in response to wasp immune challenge (Tokusumi, 2012).

To summarize, an RNAi-based loss-of-function analysis has been undertaken to identify new genes and their signaling networks vital for normal PSC niche formation and function. While information has been gained on the requirements of three such networks for PSC development and blood cell homeostasis within the lymph gland, numerous other genes have been discovered that likewise play key roles in these hematopoietic events. Their characterization is warranted as well to further enhance knowledge of genetic and molecular mechanisms at work within an accessible and easily manipulated hematopoietic progenitor niche microenvironment (Tokusumi, 2012).

Drosophila as a model for MECP2 gain of function in neurons

Methyl-CpG-binding protein 2 (MECP2) is a multi-functional regulator of gene expression. In humans, loss of MECP2 function causes classic Rett syndrome (RTT: see Rett Syndrome in 'Drosophila as a Model for Human Diseases'), but gain of MECP2 function also causes mental retardation. Although mouse models provide valuable insight into Mecp2 gain and loss of function, the identification of MECP2 genetic targets and interactors remains time intensive and complicated. This study takes a step toward utilizing Drosophila as a model to identify genetic targets and cellular consequences of MECP2 gain-of function mutations in neurons, the principle cell type affected in patients with Rett-related mental retardation. It was shown that heterologous expression of human MECP2 in Drosophila motoneurons causes distinct defects in dendritic structure and motor behavior, as reported with MECP2 gain of function in humans and mice. Multiple lines of evidence suggest that these defects arise from specific MECP2 function. First, neurons with MECP2-induced dendrite loss show normal membrane currents. Second, dendritic phenotypes require an intact methyl-CpG-binding domain. Third, dendritic defects are amended by reducing the dose of the chromatin remodeling protein, osa, indicating that MECP2 may act via chromatin remodeling in Drosophila. MECP2-induced motoneuron dendritic defects cause specific motor behavior defects that are easy to score in genetic screening. In sum, this study shows that some aspects of MECP2 function can be studied in the Drosophila model, thus expanding the repertoire of genetic reagents that can be used to unravel specific neural functions of MECP2. However, additional genes and signaling pathways identified through such approaches in Drosophila will require careful validation in the mouse model (Vonhoff, 2012).

Methyl-CpG-binding protein 2 (MECP2) is a multifunctional transcriptional regulator involved in chromatin remodeling. Loss of MECP2 function mutations cause classic Rett Syndrome (RTT), an X-linked, dominant, progressive, neuro-developmental disorder. Patients with RTT suffer from cognitive, language, motor conditions, and seizures. However, MECP2 duplication is a frequent case of mental retardation and progressive neurological symptoms in males, and overexpression of MECP2 in the developing mouse brain also causes progressive neurological disorder (Vonhoff, 2012).

The MECP2 protein contains at least five distinct functional domains (NTD, ID, MBD, TRD, and CTDα) which either bind DNA autonomously or regulate MBD (methyl-CpG binding) function. Historically, MECP2 is viewed as a transcriptional repressor that localizes to chromatin by binding to CpG dinucleotides to regulate gene expression through interactions with histone deacetylases and other cofactors. However, MECP2 can also activate transcription, associate also with un-methylated DNA, has chromatin compaction and RNA splicing functions, and several MECP2 interacting proteins are known. Therefore, multiple MECP2 functions might be mediated by interactions with diverse co-factors and by binding to both methylated and non-methylated DNA, consistent with the wide range of phenotypes observed in patients with RTT (Vonhoff, 2012).

Although Mecp2 mouse models recapitulate RTT phenotypes and provide valuable mechanistic insight into neuronal defects caused by Mecp2 mis-regulation, such as axon targeting, synaptic, and dendritic defects, the identification of MECP2 functions and target genes in this system is time intensive and complicated (Vonhoff, 2012).

The Drosophila genetic model system is increasingly being used as a tool to analyze specific genetic and cellular aspects of neurodevelopmental disorders. Short generation times, high fecundity, high throughput screening techniques, facile genetic tools, and relatively low costs have provided valuable mechanistic insights into inherited diseases like Fragile-X, Angelman syndrome, and neurofibromatosis. However, despite considerable conservation in fundamental cell biological pathways, the Drosophila genome encodes only about 75 percent of human disease associated genes, and mecp2 is not among these genes. Therefore, Drosophila can not be used to study the pathophysiology resulting from loss of endogenous mecp2. Instead, the Drosophila model relies on heterologous expression of human MECP2 allele and consequential gain of MECP2 function. Although classic Rett is mostly caused by loss-of-function of MECP2, this is likely not an artificial approach since in humans and in mouse models increased levels of MECP2 also cause disease. Genetic and behavioral proof of principle for the use of the Drosophila model to address MECP2 gain-of-function has been provided earlier. In MECP2 transgenic flies, the MECP2 protein associates with chromatin, interacts with homologs of known human MECP2 interactors, modifies the transcription of multiple genes, and is phosphorylated at serine 423, as in mammals. Most importantly, reported consequences are developmental dysfunctions and motor defects, suggesting parallels with RTT phenotypes. However, previous work on MECP2 in the Drosophila CNS has not tested for cellular phenotypes resulting from MECP2 over-expression in neurons, although mouse models demonstrate that disease phenotypes result from Mecp2 mis-regulation in postmitotic neurons. This study presents the first data on cellular defects as resulting from MECP2 gain-of-function in developing postmitotic Drosophila neurons (Vonhoff, 2012).

It was demonstrated that heterologous expression of human MECP2 in Drosophila motoneurons does not affect axonal pathfinding, dendritic territory boundaries, or the neurons' electrophysiology, but it causes a significant reduction in new dendritic branch formation during development. Similarly, in the mouse model Mecp2 mis-regulation results in pyramidal neuron dendritic defects. This study provides four lines of evidence that dendritic defects in Drosophila motoneurons are caused by specific cellular functions that result from MECP2 gain-of-function, and not from non-specific over-expression or sequestering effects. First, MECP2 protein specifically localizes to the nucleus of Drosophila neurons, so that interactions of MECP2 with molecules in the cytoplasm are unlikely. Second, targeted expression of MECP2 in Drosophila motoneurons causes significant dendritic branching defects but does not affect firing responses to current injections, voltage activated potassium current, or firing frequencies during motor behavior, indicating normal regulation of electrophysiological properties. Although it was earlier demonstrated that Drosophila motoneuron dendritic structure may undergo compensatory changes in response to altered neuronal activity, and a link between motoneuron activity and dendritic growth has clearly been established, any evidence for homeostatic changes in motoneuron excitability in response to developmental defects in dendritic structure was not found in this study. Third, MECP2-induced dendritic defects require intact MBD function of the MECP2 protein because dendritic architecture is not affected following expression of MECP2 alleles with non-functional MBD. This indicates that human MECP2 exerts specific action in Drosophila neurons via chromatin remodeling. Fourth, MECP2-induced dendritic phenotypes can be ameliorated by reducing the dose of osa, a member of the SWI/SNF complex. This genetic interaction is consistent with the hypothesis that human MECP2 may exert specific action in Drosophila motoneurons via chromatin remodeling. It also indicates that MECP2 gain-of-function activates specific cell signaling pathways in Drosophila, and may not cause unspecific over-expression effects. Therefore, the study concludes that Drosophila neurons can serve as a valuable model system to identify some cellular mechanisms by which MECP2 gain-of-function affects neuronal development (Vonhoff, 2012).

It was shown that dendritic defects, as induced by heterologous expression of MECP2 in Drosophila motoneurons, require an intact MBD domain, because expression of MECP2 with a point mutated or truncated MBD domain does not affect dendritic structure. However, each UAS-MECP2 transgene is likely inserted into a unique site in the Drosophila genome, and therefore, the possibility that different UAS-MECP2 transgenes may yield different expression levels or other genetic interactions can not be excluded. The finding that dendritic defects as caused by the expression of full length UAS-MECP2, but not by the expression of UAS-MECP2 transgenes with defective MBD domain, are a result of the unique insertion sites of the UAS-MECP2 constructs into the Drosophila genome, is unlikely for two reasons. First, both UAS-transgenes with defective MBD do not cause dendritic defects. Second, similar dendritic defects are observed following the expression of the full length MECP2 construct inserted in the second or in the third chromosome (Vonhoff, 2012). 

MBD domains recognize two key mechanisms of chromatin regulation in eukaryotes, C5 methylations of DNA at cytosines and post-translational histone modifications. Although the existence of DNA methylation was demonstrated earlier in the fly genome, methylation levels are several orders of magnitude lower than in mammals. The fly genome contains only one methylated DNA binding protein (dMBD2/3) and only one DNA methyltransferase (dDNMT2), which shows highest affinity to t-RNA. Consequently, Drosophila DNA is only sparsely methylated, so that MECP2 interactions with modified histone tails seem the more parsimonious scenario. This is consistent with the finding in this study that MECP2-dependent dendritic defects are suppressed in an osa heterozygous mutant background. Osa is a member of the SWI/SNF complex (human homolog is BAF250), a class of trithorax proteins involved in chromatin remodeling which are highly conserved between flies and humans. This indicates that human MECP2 may exert specific action in Drosophila motoneurons via chromatin remodeling. In fact, it was previously suggested that MECP2 associates with human Brahma, a catalytic component of the SWI/SNF chromatin remodeling complex to regulate gene repression, although this finding is disputed. Nevertheless, the Drosophila system provides some unique advantages to study possible interactions of MECP2 and members of the SWI/SNF chromatin remodeling complex with genetic tools (Vonhoff, 2012). 

The finding in this study that flies with MECP2 over-expression in motoneurons show normal take-off likelihoods as well as normal motoneuron firing and wing beat frequencies but can not sustain flight is in accord with specific MECP2 effects on dendrite development in otherwise normal motoneurons. In Drosophila, take-off can be mediated by the escape response neural circuitry. This circuitry bypasses flight motoneuron dendrites by synapsing directly on MN5 axon, but it relies on normal synaptic transmission and flight motoneuron physiology. Therefore, initial take-off and initial motoneuron firing are not affected by dendritic defects. In Drosophila motoneuron, firing frequencies are directly proportional to wing beat frequency, and thus, these are also not affected. By contrast, flight can not be sustained because the significantly reduced dendritic surface likely reduces the excitatory synaptic drive to motoneuron dendrites that is necessary to stay in flight. Therefore, flies with MECP2-caused motoneuron dendritic defects show a 30- to 60-fold reduction in flight duration. This behavioral phenotype is obvious, and thus, useful for screening. Although the quantification of flight durations and take-off likelihoods does not allow for rapid genetic screening, high throughput screening can easily be developed based on the observed reduction in flight duration by more than 30-fold. Moreover, high throughput assays which utilize Drosophila behavior for rapid screening have been developed by others. Such approaches may help the future identification of candidate MECP2 targets or interactors (Vonhoff, 2012).

Identification of genetic interactors and modifiers of MECP2 function in neurons will be imperative toward developing future treatment strategies. MECP2 itself is not a promising treatment target because the X-linked MECP2 gene is mosaic regulated in the human brain. Furthermore, both loss and gain of function cause disease phenotypes. The sparse methylation landscape in Drosophila may offer unique promise of identifying non-methylated DNA-dependent functions of MECP2 in neurons, the cell type that is most relevant to Rett syndrome. Since known binding partners of MECP2 are conserved in flies (e.g. YB-1, mSin3A etc.), it seems plausible that gain-of-function of human MECP2 may affect neural development via a cellular machinery that is partly conserved between flies and humans (Vonhoff, 2012).

MECP2-induced dendritic phenotypes in flight motoneurons cause a severe motor behavioral phenotype in that flight bout duration is reduced approximately 30- to 60-fold. Rapid screening assays for Drosophila behavioral phenotypes are available. Combined with the fast generation times, high fecundity and facile genetic tools available in Drosophila, this offers a powerful tool to identify molecules that interact with MECP2 in neurons. However, potential MECP2 candidate target genes or genetic modifiers of MECP2 function that can readily be identified in the Drosophila system will then have to be further evaluated in the existing mouse models of RTT (Vonhoff, 2012).


Baig, J., et al. (2010). The chromatin-remodeling Protein Osa interacts with CyclinE in Drosophila eye imaginal discs. Genetics 184: 731-744. PubMed Citation: 20008573

Benmimoun, B., Polesello, C., Waltzer, L., Haenlin, M. (2012). Dual role for Insulin/TOR signaling in the control of hematopoietic progenitor maintenance in Drosophila. Development 139(10): 1713-7. PubMed Citation: 22510984

Bonnay, F., Nguyen, X. H., Cohen-Berros, E., Troxler, L., Batsche, E., Camonis, J., Takeuchi, O., Reichhart, J. M. and Matt, N. (2014). Akirin specifies NF-kappaB selectivity of Drosophila innate immune response via chromatin remodeling. EMBO J 33(20):2349-62. PubMed ID: 25180232

Bronstein, R. and Segal, D. (2011). Modularity of CHIP/LDB transcription complexes regulates cell differentiation. Fly (Austin) 5: 200-205. PubMed ID: 21406967

Bronstein, R., Levkovitz, L., Yosef, N., Yanku, M., Ruppin, E., Sharan, R., Westphal, H., Oliver, B. and Segal, D. (2010). Transcriptional regulation by CHIP/LDB complexes. PLoS Genet 6: e1001063. PubMed ID: 20730086

Carrera, P., et al. (1998). A modifier screen in the eye reveals control genes for Kruppel activity in the Drosophila embryo. Proc. Natl. Acad. Sci. 95(18): 10779-84. 9724781

Collins, R. T., et al. (1999). Osa associates with the Brahma chromatin remodeling complex and promotes the activation of some target genes. EMBO J. 18: 7029-7040. PubMed Citation: 10601025

Collins, R. T. and Treisman, J. E. (2000). Osa-containing Brahma chromatin remodeling complexes are required for the repression of Wingless target genes. Genes Dev. 14: 3140-3152. 11124806

Dallas, P. B., Cheney, I. W., Liao, D., Bowrin, V., Byam, W., Pacchione, S., Kobayashi, R., Yaciuk, P. and Moran, E. (1998) p300/CREB binding protein-related protein p270 is a component of mammalian SWI/SNF complexes. Mol. Cell. Biol. 18: 3596-3603. PubMed Citation: 9584200

Fiedler, M., Graeb, M., Mieszczanek, J., Rutherford, T. J., Johnson, C. M. and Bienz, M. (2015). An ancient Pygo-dependent Wnt enhanceosome integrated by Chip/LDB-SSDP. Elife 4:e09073. PubMed ID: 26312500

Gregory, S. L., et al. (1996). Characterization of the dead ringer gene identifies a novel, highly conserved family of sequence-specific DNA-binding proteins. Mol. Cell. Biol. 16(3): 792-799. PubMed Citation: 8622680

Heitzler, P., Vanolst, L., Biryukova, I. and Ramain, P. (2003). Enhancer-promoter communication mediated by Chip during Pannier-driven proneural patterning is regulated by Osa. Genes Dev. 17: 591-596. 12629041

Herrscher, R. F., et al. (1995). The immunoglobulin heavy-chain matrix-associating regions are bound by Bright: a B cell-specific trans-activator that describes a new DNA-binding protein family. Genes Dev. 9(24): 3067-3082. PubMed Citation: 8543152

Hirose, F., Ohshima, N., Shiraki, M., Inoue, Y. H., Taguchi, O., Nishi, Y. Matsukage, A. and Yamaguchi, M. (2001). Ectopic expression of DREF induces DNA synthesis, apoptosis, and unusual morphogenesis in the Drosophila eye imaginal disc: possible interaction with Polycomb and trithorax group proteins. Mol. Cell. Biol. 21(21): 7231-42. 11585906

Kalionis, B. and OÂ’Farrell, P. H. (1993). A universal target sequence is bound in vitro by diverse homeodomains. Mech. Dev. 43: 57-70. PubMed Citation: 7902124

Kennison, J. A. and Tamkun, J. W. (1988). Dosage-dependent modifiers of Polycomb and Antennapedia mutations in Drosophila. Proc. Natl. Acad. Sci. 85: 8136-8140. PubMed Citation: 3141923

Li, X. S., Trojer, P., Matsumura, T., Treisman, J. E. and Tanese, N. (2010). Mammalian SWI/SNF--a subunit BAF250/ARID1 is an E3 ubiquitin ligase that targets histone H2B. Mol. Cell Biol. 30(7): 1673-88. PubMed Citation: 20086098

Milan, M., Pham, T. T. and Cohen, S. M. (2004). Osa modulates the expression of Apterous target genes in the Drosophila wing. Mech. Dev. 121: 491-497. 15147766

Mohrmann, L., et al. (2004). Differential targeting of two distinct SWI/SNF-related Drosophila chromatin-remodeling complexes. Mol. Cell. Biol. 24(8): 3077-88. 15060132

Nakamura, K., Ida, H. and Yamaguchi, M. (2008). Transcriptional regulation of the Drosophila moira and osa genes by the DREF pathway. Nucleic Acids Res. 36: 3905-3915. PubMed Citation: 18511465

Panne, D., Maniatis, T. and Harrison, S. C. (2007). An atomic model of the interferon-beta enhanceosome. Cell 129: 1111-1123. PubMed ID: 17574024

Rajaiya, J., et al. (2005). Bruton's tyrosine kinase regulates immunoglobulin promoter activation in association with the transcription factor Bright. Mol. Cell. Biol. 25(6): 2073-84. 15743806

Staehling-Hampton, K., et al. (1999). A genetic screen for modifiers of E2F in Drosophila melanogaster. Genetics 153: 275-287. PubMed Citation: 10471712

Terriente-Félix, A. and de Celis. J. F. (2009). Osa, a subunit of the BAP chromatin-remodelling complex, participates in the regulation of gene expression in response to EGFR signalling in the Drosophila wing. Dev. Biol. 329(2): 350-61. PubMed Citation: 19306864

Tokusumi, Y., Tokusumi, T., Shoue, D. A., Schulz, R. A. (2012). Gene regulatory networks controlling hematopoietic progenitor niche cell production and differentiation in the Drosophila lymph gland. PLoS One 7(7):e41604. PubMed Citation: 22911822

Treisman, J. E., Luk, A., Rubin, G. M. and Heberlein, U. (1997). eyelid antagonizes wingless signaling during Drosophila development and has homology to the Bright family of DNA-binding proteins. Genes Dev. 11: 1949-1962. PubMed Citation: 9271118

Trouche, D., et al. (1997). RB and hBRM cooperate to repress the activation functions of E2F-1. Proc. Natl. Acad. Sci. 94: 11268-11273. PubMed Citation: 9326598

Vazquez, M., Moore, L. and Kennison, J. A. (1999). The trithorax group gene osa encodes an ARID-domain protein that genetically interacts with the Brahma chromatin-remodeling factor to regulate transcription. Development 126: 733-742. PubMed Citation: 9895321

Vonhoff, F., Williams, A., Ryglewski, S. and Duch, C. (2012). Drosophila as a model for MECP2 gain of function in neurons. PLoS One 7: e31835. PubMed ID: 22363746

osa/eyelid: Biological Overview | Evolutionary Homologs | Regulation | Developmental Biology | Effects of Mutation

date revised: 21 November 2016

Home page: The Interactive Fly © 1997 Thomas B. Brody, Ph.D.

The Interactive Fly resides on the
Society for Developmental Biology's Web server.