u-shaped: Biological Overview | Evolutionary Homologs | Regulation | Developmental Biology | Effects of Mutation | References

Gene name - u-shaped

Synonyms -

Cytological map position - 21C6--21C6

Function - transcription factor

Keywords - amnioserosa, peripheral nervous system

Symbol - ush

FlyBase ID: FBgn0003963

Genetic map position - 2-0.1

Classification - zinc finger protein

Cellular location - nuclear

NCBI links: Precomputed BLAST | Entrez Gene

Recent literature
Gao, H., Baldeosingh, R., Wu, X. and Fossett, N. (2016). The Friend of GATA transcriptional co-regulator, U-Shaped, is a downstream antagonist of Dorsal-driven prohemocyte differentiation in Drosophila. PLoS One 11: e0155372. PubMed ID: 27163255
Recent studies suggest that mammalian hematopoietic stem and progenitor cells (HSPCs) respond directly to infection and inflammatory signaling. These signaling pathways also regulate HSPCs during steady-state conditions (absence of infection), and dysregulation may lead to cancer or age-related loss of progenitor repopulation capacity. Toll-like receptors (TLRs) are a major class of pathogen recognition receptors, and are expressed on the surface of immune effector cells and HSPCs. TLR/NF-κB activation promotes HSPCs differentiation; however, the mechanisms by which this signaling pathway alters the intrinsic transcriptional landscape are not well understood. Although Drosophila prohemocytes are the functional equivalent of mammalian HSPCs, a prohemocyte-specific function for Toll signaling has not been reported. Using Drosophila transgenics, this study identified prohemocyte-specific roles for Toll pathway members, Dorsal and Cactus. It was shown that Dorsal is required to limit the size of the progenitor pool. Additionally, activation of Toll signaling in prohemocytes drives differentiation in a manner that is analogous to TLR/NF-κB-driven HSPC differentiation. This was accomplished by showing that over-expression of Dorsal, or knockdown of Cactus, promotes differentiation. The study also investigated whether Dorsal and Cactus control prohemocyte differentiation by regulating a key intrinsic prohemocyte factor, U-shaped (Ush), which is known to promote multipotency and block differentiation. It was found that Dorsal represses Ush expression levels to promote differentiation, whereas Cactus maintains Ush levels to block differentiation. Additionally, another Toll antagonist, Lesswright, also maintains the level of Ush to block differentiation and promote proliferative quiescence. Collectively, these results identify a novel role for Ush as a downstream target of Toll signaling. 

Mutations in the gene u-shaped and the associated embryonic recessive lethal phenotype were first described by Nüsslein-Volhard in a characterization of mutations that affect the pattern of larval cuticle in Drosophila. In ush null embryos, the germ band fails to retract and lateral fusion occurs between anterior and posterior ectoderm. A large number of ushmutant alleles have been isolated; these include hypomorphic mutants that affect the pattern of bristles on the head and thorax (Cubadda, 1997 and references).

A number of dominant alleles for another gene, pannier, result in the development of extra thoracic bristles as well as an overexpression of achaete and scute. Mutations of ush have been isolated in a screen for second-site modifiers of the pannierD phenotype. Alleles of ush, causing a loss of function, act as dominant enhancers of pannierD heterozygotes, resulting in an increase in the number of ectopic thoracic bristles. This suggests that pannier and u-shaped might act in the same developmental pathway to regulate the achaete and scute genes. Such evidence would be of great interest, particularly because pannier's mammalian GATA homologs have been associated with neural development in only a very few studies. The discovery of a mammalian homolog of u-shaped, FOG, that acts as a co-factor for transcription factor GATA-1 in erythroid and magakaryotic differentiation, suggests a conservation of GATA protein interactions in both flies and humans (Cubadda, 1997 and references).

Evidence has been provided for a direct physical interaction between Pannier and U-shaped, explaining the observed genetic interaction. Usp antagonizes Pnr by physically interacting with the Pnr DNA-binding domain. The number of ectopic bristles in pannierD/+ flies increases in flies bearing only a single copy of u-shaped+ but decreases when three copies are present. Activation of a chicken alpha-globin promoter sequence by Pannier (via Pannier's GATA type DNA binding domain) in cultured cells is inhibited by Ush. When both Ush and wild-type Pnr are expressed simultaneously, promoter activation is abolished. Because Pnr and vertebrate globin regulating protein GATA-1 have no homology outside their GATA DNA-binding domain, and because Ush alone has no effect on globin promoter activity, these observations suggest that the function of Ush in inhibiting Panier's activation of globin is mediated through the GATA DNA-binding domain of Pnr (Haenlin, 1997).

Pnr and Ush heterodimerize through the amino-terminal zinc finger of Pnr; when associated with Ush, the transcriptional activity of Pnr is lost. In contrast, the mutant pnr protein with lesions in this finger associates only poorly with Ush and activates transcription even when cotransfected with Ush. In summary, these results suggest an antagonistic effect of Ush on Pnr function and reveal a new mode of regulation of GATA factors during development (Haenlin, 1997).

In the developing Drosophila notum, wingless expression is regulated by positive and negative Decapentaplegic signaling so that only notal cells receiving optimal levels of Decapentaplegic signal express wingless. This Decapentaplegic-dependent regulation of notal wingless expression includes multiple mechanisms involving pannier and u-shaped. In the medial notum, Pannier and U-shaped form a complex. The expression of pannier and u-shaped is positively regulated by Decapentaplegic signals emanating from the dorsal-most region. The Pannier/U-shaped complex serves as a repressor and a transcriptional activator, respectively, for wingless and u-shaped expression. In the more lateral region, wingless expression is up-regulated by U-shaped-unbound Pannier. wingless expression is also weakly regulated by its own signaling (Sato, 2000).

achaete-scute (ac-sc) expression in the notum is affected in several allelic combinations of pnr, whose function is prevented by Ush as a result of direct binding to Pnr. Since pnr appears involved in notal wg expression, and pnr and ush are expressed in the future medial notum in a graded fashion with peaks within the dpp expressing dorsal-most region, the late third instar notum was examined by staining for wg protein and pnr or USH RNA. ush and wg expression areas were found to abut on each other except for the future scutellum, while almost all wg expressing cells were situated in a ventral-most region of the pnr expression domain. Although somewhat ambiguous, a similar relationship among wg, pnr and ush expression areas was detected in small discs at an early third instar stage, the earliest stage of notal wg expression. It may thus follow that wg expression occurs in lateral-notal cells expressing pnr but not ush throughout third instar larval notal development. wg-LacZ expression occurs in a region much broader than the authentic wg expression domain; wg-LacZ expression was always observed to be expanded medially or dorsally, suggesting that the authentic wg expression domain shifts ventrally as a disc grows (Sato, 2000).

To determine the role of pnr in wg expression, examination was made of wg expression on various pnr mutant backgrounds. Strong wg misexpression occurs in medial pnr-null-mutant (pnrVX6) clones. wg-LacZ or Wg signals are detected in the entire medial notum transheterozygous for pnrVX6 and pnrVI, from which most, if not all, pnr activity is absent. In contrast, a significant reduction of wg expression occurs in pnr-null-mutant (pnrVX6) clones generated within the authentic wg expression domain. These findings indicate that pnr is involved in both negative and positive regulation of notal wg expression; Pnr serves as a positive regulator of wg expression in the future lateral notum including the authentic wg expression domain, while it is a negative factor of wg expression in the medial notum. Consistent with this, ubiquitous or clonal expression of wild-type pnr induces wg misexpression in the notum ventral to the authentic wg expression domain, while no or little wg misexpression occurs in the future medial notum (Sato, 2000).

As in the case of ac-sc expression, ush appears to serve as a negative factor for notal wg expression, since (1) wg is misexpressed in ush-null-mutant (ush1) clones in the future medial notum and (2) virtually all endogenous wg expression is abolished when wild-type ush is overexpressed throughout the notal region. In contrast to medial ush1clones, no appreciable change in wg expression is detected in ush1clones generated within the authentic wg expression domain. That the authentic wg expression domain is demarcated by medial ush expression may indicate that medial ush expression is involved in the establishment of the dorsal boundary of the authentic wg expression domain. Based on the fact that Pnr mutants such as PnrD1 and PnrD4, lacking ability to bind to Ush, are still capable of activating ac-sc in the presence of ush activity, wild-type Pnr has been proposed to be inactivated by ush through direct interactions of Ush with Pnr. However, the results presented here show that this may not be the case in notal wg expression and ac-sc expression for the DC macrochaetae formation. If Ush serves only as the inhibitor of Pnr as predicted, a wild-type copy of pnr added in trans to PnrD1 would not decrease the area of wg expression, since wild-type Pnr is considered to either activate wg expression or neutralize the negative function of Ush or both. The results presented here are apparently at variance with this consideration. Both wg and ac-sc misexpression found in the future medial notum of Pnr14/PnrVI discs are abolished in PnrD1/+ discs with no loss of wg and ac-sc expression in the authentic wg expression domain. This negative effect of wild-type pnr is reversed by halving the copy number of endogenous ush. It is concluded that, in the medial notum, Pnr forms a complex with Ush and the resultant Pnr/Ush complex represses wg and ac-sc expression directly or indirectly to establish the dorsal boundaries of the authentic wg expression domain and the ac-sc expression area for the DC macrochaetae formation (Sato, 2000).

Notal wg expression is regulated not only by dpp signaling but also by Pnr and Ush. Thus, pnr and ush expression may be under the control of Dpp signaling or conversely, Dpp signaling is regulated by pnr and ush. The second possibility, however, seems to be unlikely, since neither pnr nor ush mutant clones exhibit any appreciable change in brinker (brk)-LacZ expression. brk is a general Dpp target gene whose expression is negatively regulated by Dpp signaling. Loss of Dpp signaling causes cell-autonomous brk misexpression in the wing pouch and notum of wing imaginal discs. To determine the feasibility of the first possibility, pnr and ush expression was examined in tkva12, Mad1-2 or tkvQ253D(tkvQD) clones; tkvQD is a constitutively active form of tkv. pnr and ush are misexpressed in lateral UAS-tkvQD clones generated in late second instar, an observation indicating that pnr and ush expression is under the control of Dpp signaling. Unlike wg expression, pnr and ush expression are abolished not only in early tkva12 clones but also in late tkva12 and early Mad1-2 clones, both expressing wg, suggesting that pnr and ush expression requires higher levels of Dpp-signaling activity than those required for wg expression. Loss of ush expression in tkva12 and Mad1-2 clones might be a secondary effect due to the loss of pnr expression, since the maintenance of ush expression requires both pnr and ush activities. pnr and ush expression may be independently initiated by Dpp signaling, since pnr expression normally occurs in ush mutant clones and no ush misexpression is induced by ubiquitous pnr expression. It is concluded that the graded expression of pnr and ush is determined by Dpp signaling and hence, Pnr and Ush act downstream of Dpp (Sato, 2000).

In the larval notal region, dpp expression is not continuous but is broken by the authentic wg expression domain, thus suggesting that notal development could be regulated by Dpp signals emanating separately from dorsal and ventral sources up to the wg expression domain. As anticipated, the expression of dad (dad-LacZ), a downstream component of Dpp signaling whose expression is positively regulated by Dpp signaling, is detected not only in medial but also in lateral notum. However, double-staining of dad-LacZ and either PNR or USH RNA expression shows that unlike dad-LacZ, pnr and ush are not induced in the postero-lateral notum in spite of the presence of active Dpp signals. In addition, ectopic wg expression induced by tkvQD is restricted to the antero-lateral notum. It may thus follow that an unidentified factor represses the expression of a fraction of Dpp target genes, which include pnr, ush and wg but not dad, in the postero-lateral notum (Sato, 2000).

Wg signaling represses wg transcription for refinement of its own expression domain in the wing margin. Thus, an examination has been made of notal wg expression on Wg-signaling mutant backgrounds. In contrast to wing-margin, wg expression in the notum is activated by its own signaling though much less effectively. armadillo (arm) and disheveled (dsh) encode Wg signal transducers and wgts is a temperature-sensitive Wg secretion mutant. Weak partial wg misexpression is noted in about 50% of lateral clones (19 of 40 clones) expressing Deltaarm, which constitutively activates Wg signaling. Ectopic wg expression was also detected in a cell non-autonomous fashion when wg misexpressing clones were induced in the lateral notum. In contrast, wg transcription is considerably reduced in dsh null mutant clones. When wgts mutant discs are incubated at a non-permissive temperature, for 48 h, an appreciable reduction of wg expression is detected in the authentic wg domain without any significant change in pnr and ush expression. Taken together, these results indicate that Wg signaling weakly activates wg transcription in the future lateral notum. The failure of induction of wg misexpression in Deltaarm and wg clones in future medial notum may indicate that wg expression due to auto-activation is repressed by Pnr/Ush complexes in the medial notum. One unexpected finding is that, in the hinge region, strong wg misexpression occurred only in cells surrounding wg expressing cells, suggesting possibly a new type of Wg-dependent wg expression (Sato, 2000).

The entire notal ush expression area is included in the notal pnr expression domain and hence, notal ush expression might be positively regulated by pnr. This possibility using a pnr hypomorphic mutant and a significant reduction of notal ush expression was in pnr hypomorphic mutant flies. Thus, it is concluded that Pnr is involved in the up-regulation of notal ush expression. In the case of wg expression, Ush free of Pnr serves as an activator, while a Pnr/Ush complex serves as a repressor. To determine which forms of Pnr are involved in ush expression, examination was made of USH RNA expression in the notum expressing pnr ubiquitously and the notum transheterozygous for pnrD1 and pnrVl. Neither wild-type Pnr free of Ush nor PnrD1, incapable of binding to Ush but capable of activating wg expression, could induce ush expression. It may thus follow that a Pnr/Ush complex (but not Pnr free of Ush) is required for ush expression as a positive transcriptional regulator (Sato, 2000).

A summary is presented of wg regulation in the notum. In both future medial and lateral notal regions, dpp is expressed and Dpp signaling is active. However, ventral Dpp signals are neutralized by an unknown mechanism as far as pnr, ush and wg expression is concerned. Notal wg expression, except for that in the scutellum, is regulated through four different pathways, three under the control of Dpp signals emanating from the dorsal-most region. pnr and ush expression is up-regulated by Dpp signaling, but ush expression is much narrower than that of pnr, possibly because of the requirement of higher Dpp-signaling activity for ush expression than that for pnr expression. In the future medial notum, Pnr and Ush form a complex repressing wg expression, while Ush-unbound Pnr activates lateral wg expression. The authentic wg domain and the medial notum abut one another. Unlike wg expression, ush expression in the future medial notum is positively regulated by the Pnr/Ush complex. This regulation appears required for the maintenance of medial ush expression. Dpp signaling is also capable of activating notal wg expression through an unidentified factor X. This route includes neither Pnr nor Ush. In addition, wg expression is weakly up-regulated by its own signaling in the lateral notum (Sato, 2000).

The Friend of GATA protein U-shaped functions as a hematopoietic tumor suppressor in Drosophila

Drosophila has emerged as an important model system to discover and analyze genes controlling hematopoiesis. One regulatory network known to control hemocyte differentiation is the Janus kinase (JAK)/Signal Transducer and Activator of Transcription (STAT) signal-transduction pathway. A constitutive activation mutation of the Janus kinase Hopscotch (hopscotchTumorous-lethal; hopTum-l) results in a leukemia-like over-proliferation of hemocytes and copious differentiation of lamellocytes during larval stages. Friend of GATA (FOG) protein U-shaped (Ush) is expressed in circulating and lymph gland hemocytes, where it plays a critical role in controlling blood cell proliferation and differentiation. These findings demonstrate that a reduction in ush function results in hematopoietic phenotypes strikingly similar to those observed in hopTum-l animals. These include lymph gland hypertrophy, increased circulating hemocyte concentration, and abundant production of lamellocytes. Forced expression of N-terminal truncated versions of Ush likewise leads to larvae with severe hematopoietic anomalies. In contrast, expression of wild-type Ush results in a strong suppression of hopTum-l phenotypes. Taken together, these findings demonstrate that U-shaped acts to control larval hemocyte proliferation and suppress lamellocyte differentiation, likely regulating hematopoietic events downstream of Hop kinase activity. Such functions appear to be facilitated through Ush interaction with the hematopoietic GATA factor Serpent (Srp) (Sorrentino, 2007).

In wild-type lymph glands, Ush is not detectable in the second instar larva (L2) but is expressed in L3, beginning in the cortical zone and eventually spreading to the entire lymph gland. It stands to reason that during the normal dispersal of the hematopoietic organs in late L3, those lymph gland hemocytes in the cortical zone will be the first to enter circulation. In such a model, Ush-expressing lymph gland hemocytes enter the circulating hemocyte population, which (with the exception of crystal cells) already express Ush. Thus Ush can be viewed as a hemocyte maturation marker. This begs the question of why Ush is expressed in what are apparently the most mature hemocytes. The current observations strongly implicate Ush as being present in order to suppress proliferation and lamellocyte differentiation among mature plasmatocytes (Sorrentino, 2007).

The strongest evidence for a mechanism in which Ush suppresses hemocyte proliferation and lamellocyte differentiation comes from analyses of ush mutants. Reducing Ush function causes lymph gland hypertrophy, which is a direct result of an increase in the number of lymph gland hemocytes. Furthermore, in a manner similar in quality (but somewhat less in intensity) to those of hopTum-l larvae, ush mutant lymph glands disperse precociously, and cortical zone hemocytes appear to be morphologically consistent with lamellocytes. Additionally, total circulating hemocyte concentration (CHC) of ush mutants is over four-fold greater than that of wild-type larvae, and two different alleles of ush when heterozygous, induce a less severe but nonetheless significant hematopoietic phenotype. High CHC is consistent with the mechanism of a large number of hemocytes, including lamellocytes, leaving the lymph gland and entering circulation. The possibility cannot be ruled out that circulating hemocytes, which are of a different embryonic origin than lymph gland hemocytes, can also over-proliferate and/or differentiate into lamellocytes in ush mutants. Since cortical zone hemocytes (predominantly plasmatocytes) express Ush and can differentiate into lamellocytes, the fact that circulating plasmatocytes also express Ush would support the notion of circulating plasmatocytes also being able to differentiate into lamellocytes (Sorrentino, 2007).

Importantly, it was observed that wild-type L2 lymph glands do not express Ush, while L3 organs do (in the cortical zone first, then throughout the lymph gland). Clearly, a mechanism that represses a developmental decision is not necessary unless a cell has the potential to actually make the choice. Thus the existence of a Ush-regulated mechanism for suppression of hemocyte proliferation and lamellocyte differentiation in L3 cortical zone hemocytes is interpreted as supportive of the hypothesis that Ush+ cells are in a different genetic state in which they can, given the proper cues, hyperproliferate and differentiate into lamellocytes. It follows that wild-type L2 lymph gland hemocytes cannot hyperproliferate and become lamellocytes. Such a putative mechanism is consistent with previous findings; Jung (2005) observed that L3 lymph gland hemocyte proliferation takes place primarily within the cortical zone, while Sorrentino (2002) observed that, in larvae parasitized by the wasp Leptopilina boulardi, L2 lymph glands are immune-unresponsive (as indicated by mitotic index, crystal cell population size, and a lamellocyte marker) whereas L3 lymph glands do respond (Sorrentino, 2007).

Additional strong evidence for the role of Ush is provided by transgene expression data. Expression of wild-type Ush in hopTum-l/Y larvae produced an effect opposite in quality to that of ushVX22/ushr24, that being a significant 90% reduction in the hopTum-l-induced circulating lamellocyte population. The CgGAL4 driver is active in hopTum-l/Y L2 hemocytes, thus transgenic Ush has an opportunity to act on hemocytes prior to lymph gland dispersal. The significant reduction could be explained by the suppression of lamellocyte differentiation and/or the suppression of the proliferation of pro-lamellocytes. An apoptotic mechanism may also be partially involved. The reason for the observation that the hopTum-l non-lamellocyte population was not significantly affected by transgenic Ush is unknown, but one explanation would be a dosage-dependent mechanism in which experimental expression of just one copy of a UASush transgene is insufficient to suppress hopTum-l over-proliferation. It is also possible that hemocyte over-proliferation is a secondary effect of lamellocyte differentiation, and thus not under the direct control of Ush (Sorrentino, 2007).

If Ush normally suppresses crystal cell differentiation why do ush mutants not exhibit a severe overabundance of crystal cells? Using the strong amorphic ush1 background, an approximately 30% increase in mean crystal cell counts has been observed in stage-16 embryos. However, since the wild-type mean number of crystal cells in stage-16 embryos is about 24-25 per embryo, a 30% increase amounts to about 8 additional crystal cells. Since there are hundreds of plasmatocytes in an embryo, the overwhelming majority of plasmatocytes do not become crystal cells in the absence of Ush (Sorrentino, 2007).

Such findings can be explained by a model in which Ush suppresses crystal cell differentiation in a small subset of hemocytes, with the primary role of Ush in hematopoiesis being to control lamellocyte differentiation and hemocyte proliferation. In this model, the down-regulation of ush expression in embryonic crystal cells occurs because hemocytes committed to the crystal cell lineage cannot become lamellocytes, and thus require no mechanism to suppress lamellocyte differentiation (Sorrentino, 2007).

Srp is expressed in all hopTum-l/Y hemocytes, both lamellocytes and non-lamellocytes, in circulation and in the lymph gland. Thus, all Drosophila hemocyte classes studied thus far express this hematopoietic GATA factor, and the role of Srp in the differentiation of all hemocyte types is worthy of investigation. The fact that the lamellocyte population observed in ushVX22/+ larvae is strongly reduced by srpneo45/+ and completely reduced to wild-type levels by srp3/+ suggests Srp plays an active role in lamellocyte differentiation. In humans, GATA-3 determines the differentiation of Th2 cells, which like lamellocytes are the primary effectors in a cellular immune response against metazoan endoparasites. FOG-1 inhibits Th2 differentiation by inhibiting GATA-3 activity via physical interaction. However, if Srp is indeed necessary for lamellocyte differentiation, the finding that Srp is expressed in all hemocytes likely means it is not the sole determinant of lamellocyte differentiation. Thus it is considered likely that an additional transcriptional regulator, either another GATA factor (e.g., Grain, dGATAd, dGATAe) or a non-GATA factor, works in conjunction with Srp to specify the lamellocyte differentiation program (Sorrentino, 2007).

Ush232-1191, Ush302-1191, and Ush365-1191 driven by CgGAL4 are dominant inducers of hematopoietic tumor phenotypes, measurably stronger than that of the ushVX22/ushr24 loss-of-function condition. This finding is not interpreted as coincidental. In an important parallel all three of these constructs, when activated by the mesodermal twiGAL4 driver, exhibit a failure to suppress the expression of a cardiac-active Dmef2-lacZ reporter gene. Additionally, it was observed that Ush232-1191, though missing zinc finger 1, is still able to bind to Srp in vitro just as it is able to bind to Pnr. Such observations are consistent with the possibility that endogenous and transgenic Ush may compete in their binding to Srp. In such a situation, transgenic Ush232-1191, even if bound to Srp, might fail to suppress Srp-induced hemocyte proliferation and lamellocyte differentiation. Alternative explanations include: (1) Ush232-1191 bound to Srp may actually enhance normal Srp activity; (2) the Srp:Ush232-1191 complex may behave neomorphically; (3) Ush may normally dimerize while not bound to Srp, if so a Ush:Ush232-1191 complex may not be able to separate into active Ush monomers. The transgenic Ush proteins are assumed to be sufficiently stable and functional as to validate these observations, since the UASush constructs used have been shown to generate stable proteins in other cell types and also to induce measurable phenotypes (Sorrentino, 2007).

There is significantly more Ush present in the nuclei of hopTum-l/Y hemocytes than in nuclei of wild-type blood cells. Interestingly, Ush appears to be exclusively nuclear in hopTum-l/Y hemocytes, whereas there appears to be some cytoplasmic anti-Ush staining in wild-type hemocytes. It is possible that Ush function is in part determined by its cytoplasmic/nuclear ratio. Perhaps the qualitatively higher Ush concentration in hopTum-l hemocytes is the result of nuclear translocation of all cellular Ush. Based on work with human 293T and mouse erythroleukemia cell cultures, Garriga-Canut (2004) proposed a model in which TACC-3 and GATA-1 compete in binding to FOG-1, with FOG-1 bound to TACC-3 retained in the cytoplasm. Such a mechanism may also be at work in Drosophila hemocytes and the possibility of a Ush cytoplasmic sequestration phenomenon remains to be investigated (Sorrentino, 2007).

This study also found that there exist high concentrations of exclusively nuclear Ush in other tumorous backgrounds, those being in Tl10b/+ and CgGAL4>UAScol animals. Larvae carrying the dominant Tl10b allele exhibit a hematopoietic tumor phenotype similar to that of hopTum-l/Y larvae. In addition to srpDGAL4>UAScol, CgGAL4>UAScol is sufficient to induce lamellocyte differentiation (although it also induces L2 developmental arrest). All hemocytes, including lamellocytes, in both of these backgrounds also exhibit high concentrations of nuclear Ush. These observations are consistent with a model in which three different signaling pathways (Hop-Stat, Toll-Dorsal, and the early B-cell related factor Collier) all make use of a single common downstream lamellocyte induction program that involves Ush. Examination of hemocytes from additional tumorous backgrounds will reveal whether such a model is truly universal. An important question remains as to how Ush suppresses hemocyte proliferation and lamellocyte differentiation, yet is expressed so strongly in tumorous hemocytes. Taken together, the findings are supportive of Ush having an early function in repressing lamellocyte differentiation. Up-regulation of the protein in lamellocytes would be suggestive of a second, separate function for Ush within this differentiated hemocyte. Comparable multi-functional properties have been reported for FOG-1 in vertebrate hematopoiesis (Sorrentino, 2007).

Therefore, reduction of Ush function results in a classic hematopoietic tumor phenotype: lymph gland hypertrophy and early dispersal, a significant increase in total circulating hemocyte concentration, large-scale lamellocyte differentiation, and melanotic tumors. These anomalies can be partially induced by the loss-of-function of a single copy of ush. The identification of this FOG class protein as a tumor suppressor raises questions about the roles of other FOG proteins in mammalian leukemias. While mutations in murine fog1 have been associated with hematopoietic dysfunction such as the failure of megakaryopoiesis and the arrest of erythropoiesis, FOG proteins have not been implicated as a causal factor in any human leukemia. While there is no guarantee that the observations in Drosophila will directly translate to specific human hematopoietic pathologies, it may now be worthwhile to examine the state of fog gene expression and function in human leukemias (Sorrentino, 2007).


Genomic length - 19 kb

Transcript length - 4.7 kb

Exons - 8


Amino Acids - 1191

Structural Domains

The Ush open reading frame displays two types of zinc finger repeated motifs, CCHH and CCHC, found in transcription factors of the zinc finger family. In the N-terminal half are two CCHC Zn fingers surrounding a single CCHH finger. In the C-terminal half, there are two CCHC finger motifs followed by three CCHH motifs and a single C-terminal CCHC motif. The first CCHH motif shows a similarity to the second zinc finger motif of the ZFY transcription factor family. The Ush protein also contains an acidic domain in the amino-terminal part of the protein (amino acids 9-104), and several stretches of alanine residues (amino acids 421-431, 674-683 and 1107-1114).


The hematopoietic transcription factor GATA-1 is essential for development of the erythroid and megakaryocytic lineages. Using the conserved zinc finger DNA-binding domain of GATA-1 in the yeast two-hybrid system, a novel, multitype zinc finger protein, Friend of GATA-1 (FOG) has been identified, which binds GATA-1 but not a functionally inactive mutant lacking the amino (N) finger. FOG is coexpressed with GATA-1 during embryonic development and in erythroid and megakaryocytic cells. Furthermore, FOG and GATA-1 synergistically activate transcription from a hematopoietic-specific regulatory region and cooperate during both erythroid and megakaryocytic cell differentiation. These findings indicate that FOG acts as a cofactor for GATA-1 and provide a paradigm for the regulation of cell type-specific gene expression by GATA transcription factors (Tsang, 1997).

GATA transcription factors are required for the differentiation of diverse cell types in several species. Recent evidence suggests that their biologic activities may be modulated through interaction with multitype zinc finger proteins, such as Friend of GATA-1 (FOG) and U-shaped (Ush). In cell culture, FOG cooperates with the hematopoietic transcription factor GATA-1 to promote erythroid and megakaryocytic differentiation. Mice lacking FOG die during mid-embryonic development with severe anemia. FOG-/- erythroid cells display a marked, but partial, blockage of maturation, reminiscent of GATA-1- erythroid precursors. In contrast to GATA-1 deficiency, however, megakaryocytes fail to develop in the absence of FOG. Although the FOG-/- erythroid phenotype supports the proposed role of FOG as a GATA-1 cofactor in vivo, the latter finding points to a pivotal, GATA-1-independent requirement for FOG in megakaryocyte development from the bipotential erythroid/megakaryocytic progenitor. It is speculated that FOG and other FOG-like proteins serve as complex cofactors that act through both GATA-dependent and GATA-independent mechanisms (Tsang, 1998).

Protein-protein interactions play significant roles in the control of gene expression. These interactions often occur between small, discrete domains within different transcription factors. In particular, zinc fingers, usually regarded as DNA-binding domains, are now also known to be involved in mediating contacts between proteins. The interaction between the erythroid transcription factor GATA-1 and its partner, the 9 zinc finger protein, FOG (Friend Of GATA), has been investigated. This interaction represents a genuine finger-finger contact, which is dependent on zinc-coordinating residues within each protein. The contact domains have been mapped to the core of the N-terminal zinc finger of GATA-1 and the 6th zinc finger of FOG. Using a scanning substitution strategy, key residues within the GATA-1 N-finger that are required for FOG binding have been identified. These residues are conserved in the N-fingers of all GATA proteins known to bind FOG, but are not found in the respective C-fingers. This observation may, therefore, account for the particular specificity of FOG for N-fingers. Interestingly, the key N-finger residues are seen to form a contiguous surface, when mapped onto the structure of the N-finger of GATA-1 (Fox, 1998).

Friend of GATA-1 (FOG-1) is a zinc finger protein that has been shown to interact physically with the erythroid DNA-binding protein GATA-1 and modulate its transcriptional activity. Recently, two new members of the FOG family have been identified: a mammalian protein, FOG-2 (which also associates with GATA-1 and other mammalian GATA factors), and U-shaped, a Drosophila protein that interacts with the Drosophila GATA protein Pannier. FOG proteins contain multiple zinc fingers and the sixth finger of FOG-1 is known to interact specifically with the N-finger but not the C-finger of GATA-1. Fingers 1, 5 and 9 of FOG-1, all atypical Cys-Cys:His-Cys fingers, also interact with the N-finger of GATA-1; FOG-2 and U-shaped also contain multiple GATA-interacting fingers and both FOG-2 and U-shaped contain several Cys-Cys:His-Cys zinc fingers. The key contact residues are defined and these residues are shown to be highly conserved in GATA-interacting fingers. The effects of selectively mutating the four interacting fingers of FOG-1 were examined and each is shown to contribute to FOG-1's ability to modulate GATA-1 activity. FOG-1 can repress GATA-1-mediated activation: evidence is presented that this ability involves the recently described CtBP co-repressor proteins that recognize all known FOG proteins (Fox, 1999).

The mechanism by which FOG-1 acts to repress GATA-mediated transcription was investigated. FOG-1 contains a motif that is bound by the CtBP family of co-repressors. This site PIDLSKR occurs immediately N-terminal to finger 7. Yeast two-hybrid and GST pull-down assays were used to test whether a small region of FOG-1 (residues 724-834, spanning the CtBP-binding motif) could interact with one family member, mCtBP2. GST-FOG-1(724-834) can retain in vitro-translated mCtBP2 efficiently, whereas a mutant FOG-1 containing a mutation in the core region (PIDLSKR to AIAASKR) is unable to retain mCtBP2. Similarly, in the yeast two-hybrid system, FOG-1(724-834) is able to interact with mCtBP2, whereas the mutant cannot. To test if this region of FOG-1 can act as a repression domain in vivo, fusions of FOG-1(724-834) (both wild-type and mutant) with the Gal4 DNA-binding domain were prepared and these were co-transfected with a construct harbouring a Gal4-dependent promoter upstream of the human growth hormone reporter gene. Gal4DBD-FOG-1(724-834) represses the basal reporter activity 20-fold. However, the mutant is unable to significantly repress transcription. This result indicates that FOG-1 contains a repression domain that can mediate repression by associating with CtBP family proteins (Fox, 1999).

The FOG-2 gene was disrupted in mice to define its requirement in vivo. FOG-2(-/-) embryos die at midgestation with a cardiac defect characterized by a thin ventricular myocardium, common atrioventricular canal, and the tetralogy of Fallot malformation. Remarkably, coronary vasculature is absent in FOG-2(-/-) hearts. Despite formation of an intact epicardial layer and expression of epicardium-specific genes, markers of cardiac vessel development (ICAM-2 and FLK-1) are not detected, indicative of failure to activate their expression and/or to initiate the epithelial to mesenchymal transformation of epicardial cells. Transgenic reexpression of FOG-2 in cardiomyocytes rescues the FOG-2(-/-) vascular phenotype, demonstrating that FOG-2 function in myocardium is required and sufficient for coronary vessel development. These findings provide the molecular inroad into the induction of coronary vasculature by myocardium in the developing heart (Tevosian, 2000).

Sox is a large family of genes related to the sex-determining region Y gene (designated as the SRY gene), In mammals, Sry expression in the bipotential, undifferentiated gonad directs the support cell precursors to differentiate as Sertoli cells, thus initiating the testis differentiation pathway. In the absence of Sry, or if Sry is expressed at insufficient levels, the support cell precursors differentiate as granulosa cells, thus initiating the ovarian pathway. The molecular mechanisms upstream and downstream of Sry are not well understood. The transcription factor GATA4 and its co-factor FOG2 are required for gonadal differentiation. Mouse fetuses homozygous for a null allele of Fog2 or homozygous for a targeted mutation in Gata4 (Gata4ki) that abrogates the interaction of GATA4 with FOG co-factors exhibit abnormalities in gonadogenesis. Sry transcript levels are significantly reduced in XY Fog2–/– gonads at E11.5, which is the time when Sry expression normally reaches its peak. In addition, three genes crucial for normal Sertoli cell function (Sox9, Mis and Dhh) and three Leydig cell steroid biosynthetic enzymes (p450scc, 3ßHSD and p450c17) are not expressed in XY Fog2–/– and Gataki/ki gonads, whereas Wnt4, a gene required for normal ovarian development, is expressed ectopically. By contrast, Wt1 and Sf1, which are expressed prior to Sry and necessary for gonad development in both sexes, are expressed normally in both types of mutant XY gonads. These results indicate that GATA4 and FOG2 and their physical interaction are required for normal gonadal development (Tevosian, 2002).

GATA transcription factors are important regulators of both hematopoiesis (GATA-1/2/3) and cardiogenesis (GATA-4) in mammals. The transcriptional activities of the GATA proteins are modulated by their interactions with other transcription factors and with transcriptional coactivators and repressors. Recently, two related zinc finger proteins, U-shaped (Ush) and Friend of GATA-1 (FOG) have been reported to interact with the GATA proteins Pannier and GATA-1, respectively, and to modulate their transcriptional activities in vitro and in vivo. In this report, the molecular cloning and characterization of a third FOG-related protein, FOG-2 is described. FOG-2 is an 1,151 amino acid nuclear protein that contains eight zinc finger motifs that are structurally related to those of both FOG and Ush. FOG-2 is first expressed in the mouse embryonic heart and septum transversum at embryonic day 8.5 and is subsequently expressed in the developing neuroepithelium and urogenital ridge. In the adult, FOG-2 is expressed predominately in the heart, brain, and testis. FOG-2 associates physically with the N-terminal zinc finger of GATA-4 both in vitro and in vivo. This interaction appears to modulate specifically the transcriptional activity of GATA-4 because overexpression of FOG-2 in both NIH 3T3 cells and primary rat cardiomyocytes represses GATA-4-dependent transcription from multiple cardiac-restricted promoters. Taken together, these results implicate FOG-2 as a novel modulator of GATA-4 function during cardiac development and suggest a paradigm in which tissue-specific interactions between different FOG and GATA proteins regulate the differentiation of distinct mesodermal cell lineages (Svensson, 1999).

Tricuspid atresia (TA) is a common form of congenital heart disease, accounting for 1%-3% of congenital cardiac disorders. TA is characterized by the congenital agenesis of the tricuspid valve connecting the right atrium to the right ventricle and both an atrial septal defect (ASD) and a ventricular septal defect (VSD). Some patients also have pulmonic stenosis, persistence of a left-sided superior vena cava or transposition of the great arteries. Most cases of TA are sporadic, but familial occurrences with disease in multiple siblings have been reported. Gata4 is a zinc-finger transcription factor with a role in early cardiac development. Gata4-deficient mice fail to form a ventral heart tube and die of circulatory failure at embryonic day (E) 8.5. Zfpm2 (also known as Fog-2) is a multi-zinc-finger protein that is co-expressed with Gata4 in the developing heart beginning at E8.5. Zfpm2 interacts specifically with the N-terminal zinc finger of Gata4 and represses Gata4-dependent transcription. Targeted mutagenesis was used to explore the role of Zfpm2 in normal cardiac development. Zfpm2-deficient mice die of congestive heart failure at E13 with a syndrome of tricuspid atresia that includes an absent tricuspid valve, a large ASD, a VSD, an elongated left ventricular outflow tract, rightward displacement of the aortic valve and pulmonic stenosis. These mice also display hypoplasia of the compact zone of the left ventricle. These findings indicate the importance of Zfpm2 in the normal looping and septation of the heart and suggest a genetic basis for the syndrome of tricuspid atresia (Svensson, 2000a).

GATA4 is a transcriptional activator of cardiac-restricted promoters and is required for normal cardiac morphogenesis. Friend of GATA-2 (FOG-2) is a multizinc finger protein that associates with GATA4 and represses GATA4-dependent transcription. To better understand the transcriptional repressor activity of FOG-2 a functional analysis of the FOG-2 protein was performed. The results demonstrate that (1) zinc fingers 1 and 6 of FOG-2 are each capable of interacting with evolutionarily conserved motifs within the N-terminal zinc finger of mammalian GATA proteins; (2) a nuclear localization signal (RKRRK) (amino acids 736-740) is required to program nuclear targeting of FOG-2, and (3) FOG-2 can interact with the transcriptional co-repressor, C-terminal-binding protein-2 via a conserved sequence motif in FOG-2 (PIDLS). Surprisingly, however, this interaction with C-terminal-binding protein-2 is not required for FOG-2-mediated repression of GATA4-dependent transcription. Instead, a novel N-terminal domain of FOG-2 (amino acids 1-247) has been identifed that is both necessary and sufficient to repress GATA4-dependent transcription. This N-terminal repressor domain is functionally conserved in the related protein, Friend of GATA1. Taken together, these results define a set of evolutionarily conserved mechanisms by which FOG proteins repress GATA-dependent transcription and thereby form the foundation for genetic studies designed to elucidate the role of FOG-2 in cardiac development (Svensson, 2000b).

Members of the GATA family of zinc-finger transcription factors have critical roles in a variety of cell types. GATA-1, GATA-2 and GATA-3 are required for proliferation and differentiation of several hematopoietic lineages, whereas GATA-4, GATA-5 and GATA-6 activate cardiac and endoderm gene expression programs. Two GATA cofactors have recently been identified. Friend of GATA-1 (FOG-1) interacts with GATA-1 and is expressed principally in hematopoietic lineages, whereas FOG-2 is expressed predominantly in heart and brain. Although gene targeting experiments are consistent with an essential role for FOG-1 as an activator of GATA-1 function, reporter assays in transfected cells indicate that FOG-1 and FOG-2 can act as repressors. A Xenopus laevis homolog of FOG has been cloned that is structurally most similar to FOG-1, but is expressed predominantly in heart and brain, as well as the ventral blood island and adult spleen. Ectopic expression and explant assays demonstrate that FOG proteins can act as repressors in vivo, in part through interaction with the transcriptional co-repressor, C-terminal Binding Protein (CtBP). FOG may regulate the differentiation of red blood cells by modulating expression and activity of GATA-1 and GATA-2. It is proposed that the FOG proteins participate in the switch from progenitor proliferation to red blood cell maturation and differentiation (Deconinck, 2000).

The commitment of multipotent cells to particular developmental pathways requires specific changes in their transcription factor complement to generate the patterns of gene expression characteristic of specialized cell types. The role of the GATA cofactor Friend of GATA in the differentiation of avian multipotent hematopoietic progenitors has been examined. Multipotent cells express high levels of FOG mRNA, which are rapidly down-regulated upon their C/EBP-mediated commitment to the eosinophil lineage. Expression of FOG in eosinophils leads to a loss of eosinophil markers and the acquisition of a multipotent phenotype, and constitutive expression of FOG in multipotent progenitors blocks activation of eosinophil-specific gene expression by C/EBPbeta. These results show that FOG is a repressor of the eosinophil lineage, and that C/EBPbeta-mediated down-regulation of FOG is a critical step in eosinophil lineage commitment. Furthermore, the results indicate that maintenance of a multipotent state in hematopoiesis is achieved through cooperation between FOG and GATA-1. A model is presented in which C/EBPbeta induces eosinophil differentiation by the coordinate direct activation of eosinophil-specific promoters and the removal of FOG, a promoter of multipotency as well as a repressor of eosinophil gene expression (Querfurth, 2000).

GATA-4 has been implicated in formation of the vertebrate heart. Since the mouse Gata-4 knock-out is early embryonic lethal because of a defect in ventral morphogenesis, the in vivo function of this factor in heart development remains unresolved. To search for a requirement for Gata4 in heart development, mice were created harboring a single amino acid replacement in GATA-4 that impairs its physical interaction with its presumptive cardiac cofactor FOG-2, a homolog of Drosophila U-shaped. Gata4ki/ki mice die just after embryonic day 12.5, exhibiting features in common with Fog2-/- embryos as well as additional semilunar cardiac valve defects and a double-outlet right ventricle. These findings establish an intrinsic requirement for GATA-4 in heart development. It is inferred that GATA-4 function is dependent on interaction with FOG-2 and, very likely, an additional FOG protein for distinct aspects of heart formation (Crispino, 2001).

The power of this analysis rests on the exquisite specificity of the knock-in mutation within the N finger of GATA-4. The residue that was modified is required for physical interaction with FOG-like proteins and does not influence the DNA-binding specificity of the GATA-protein. Although displaying many similar features, Gata4ki/ki hearts are distinguished from Fog2-/- hearts, however, by the presence of a double-outlet right ventricle and defects in the semilunar valves and outflow tracts. Since immunostaining confirms that GATA-4 is expressed at wild-type levels in the semilunar valves of the Gata4 mutant heart, it is likely that another FOG, or FOG-like protein, that functions as a cofactor for GATA-4 in transcription, is expressed in these valve cells. Though high-level expression of the only other known vertebrate FOG-like factor, FOG-1, has not previously been observed by in situ RNA hybridization, FOG-1 transcripts are present at low levels in Northern blots of total heart RNA. Thus, it is quite possible that disruption of the physical interaction between GATA-4 and FOG-1, or a novel, undefined FOG protein, is responsible for impaired development of the semilunar valves and the appearance of a double-outlet right ventricle (Crispino, 2001).

Given the profound effects of mutation of either GATA-4 or FOG-2 proteins on heart morphogenesis in mice, it is worth considering their potential relevance to human congenital heart defects, such as the Tetralogy of Fallot or the double-outlet right ventricle. Whereas mutation of several genes, such as Jmj (Jumanji), Sox4, and Egfr/Shp2, give rise to the double-outlet right ventricle defect in mice, and defects in other genes for transcription factors, such as FOG-2, NF1, neurotrophin 3, and RXR, result in all or a subset of the Tetralogy of Fallot, the consistent and combined phenotype seen in the Gata4ki/ki mice is unique. Although it is sometimes difficult to distinguish between double-outlet right ventricle with associated pulmonary stenosis and the Tetralogy of Fallot, it is clear that the defects in the Gata4ki/ki hearts are different in the outflow tracts than those observed in Fog2-deficient embryos (Crispino, 2001).

Through the use of an altered specificity mutant, it has been demonstrated that GATA-4 very likely requires both FOG-2 and an additional FOG, or FOG-like protein, as cofactors for distinct aspects of heart development. Interaction with FOG-2 is essential for the initiation of coronary vasculature and for some morphogenetic events, whereas interaction with a distinct FOG protein appears to be required for formation of cardiac valves. The results are surprising in that two other GATA-factors, GATA-5 and GATA-6, are also expressed in myocardium, and indirect data have suggested that they might compensate for the absence of GATA-4. For example, previous studies show that GATA-4 is dispensable for terminal differentiation of cardiomyocytes and that Gata4-/- ES cells contribute to all layers of the heart. In these experiments, it has been suggested that GATA-5 or GATA-6 functionally replace GATA-4. It is possible that proper expression of the GATA-4ki/ki protein, as distinguished from the absence of GATA-4 in the knock-out situation, precludes compensation by other GATA factors. Indeed, immunostaining with an alpha-GATA-6 antibody has demonstrated that GATA-6 expression, though similar to that of GATA-4, is not up-regulated in the Gata4ki/ki hearts. In addition, staining with alpha-GATA-5 antibody reveals a normal pattern in Gata4ki/ki hearts. Since GATA-5 is no longer expressed within the ventricles of the heart at E12.5, it is unlikely that it would compensate for the absence of functional GATA-4 (Crispino, 2001).

These findings implicate GATA-4 as the principal GATA factor relevant to heart morphogenesis and coronary vasculature development and as the primary partner for FOG proteins in the heart. This represents the second example of transcriptional regulation involving GATA-FOG protein complexes and argues for their broad involvement as key regulators of multiple developmental pathways (Crispino, 2001).

The transcription factor GATA-1 and its cofactor FOG-1 are essential for the normal development of erythroid cells and megakaryocytes. FOG-1 can stimulate or inhibit GATA-1 activity depending on cell and promoter context. How the GATA-1-FOG-1 complex controls the expression of distinct sets of gene in megakaryocytes and erythroid cells is not understood. The molecular basis for the megakaryocyte-restricted activation of the aIIb gene has been examined. FOG-1 stimulates GATA-1-dependent aIIb gene expression in a manner that requires their direct physical interaction. Transcriptional output by the GATA-1-FOG-1 complex is determined by the hematopoietic Ets protein Fli-1 that binds to an adjacent Ets element. Chromatin immunoprecipitation experiments show that GATA-1, FOG-1 and Fli-1 co-occupy the aIIb promoter in vivo. Expression of several additional megakaryocyte-specific genes that bear tandem GATA and Ets elements in their promoters also depends on the physical interaction between GATA-1 and FOG-1. These studies define a molecular context for transcriptional activation by GATA-1 and FOG-1, and may explain the occurrence of tandem GATA and Ets elements in the promoters of numerous megakaryocyte-expressed genes (Wang, 2002).

GATA-1 and friend of GATA (FOG) are zinc-finger transcription factors that physically interact to play essential roles in erythroid and megakaryocytic development. Several naturally occurring mutations in the GATA-1 gene that alter the FOG-binding domain have been reported. The mutations are associated with familial anemias and thrombocytopenias of differing severity. To elucidate the molecular basis for the GATA-1/FOG interaction, the three-dimensional structure of a complex comprising the interaction domains of these proteins has been determined. The structure reveals how zinc fingers can act as protein recognition motifs. Notably, none of the FOG ZnFs that contact GATA-1 are part of tandem arrays of ZnFs. Thousands of such 'isolated' ZnFs exist, and it is likely that many serve as protein recognition motifs. The surface used by FOG ZnFs to recognize GATA-1 overlaps with the surface normally used by classical ZnFs to bind to DNA, indicating that the classical ZnF has acted throughout evolution as a versatile structural scaffold, onto which different binding functions have been 'grafted'. In line with this idea, the third classical ZnF from FOG has been shown to mediates a specific interaction with the coiled-coil protein TACC3. Indeed, given that a single classical ZnF is capable of mediating protein-protein interactions, and that an array of such domains is necessary for high affinity DNA binding, it is likely that the latter function arose later as a consequence of gene duplication events (Liew, 2004).

Inderstanding of the maternal factors that initiate early chordate development, and of their direct zygotic targets, is still fragmentary. A molecular cascade is emerging for the ascidian mesendoderm, but less is known about the ectoderm, giving rise to epidermis and nervous tissue. Cis-regulatory analysis surprisingly places the maternal transcription factor Ci-GATAa (GATA4/5/6) at the top of the ectodermal regulatory network in ascidians. Initially distributed throughout the embryo, Ci-GATAa activity is progressively repressed in vegetal territories by accumulating maternal β-catenin. Once restricted to the animal hemisphere, Ci-GATAa directly activates two types of zygotic ectodermal genes. First, Ciona friend of GATA gene (Ci-fog) is activated from the 8-cell stage throughout the ectoderm, then Ci-otx is turned on from the 32-cell stage in neural precursors only. Whereas the enhancers of both genes contain critical and interchangeable GATA sites, their distinct patterns of activation stem from the additional presence of two Ets sites in the Ci-otx enhancer. Initially characterized as activating elements in the neural lineages, these Ets sites additionally act as repressors in non-neural lineages, and restrict GATA-mediated activation of Ci-otx. This study has identified a precise combinatorial code of maternal factors responsible for zygotic onset of a chordate ectodermal genetic program (Rothbacher, 2007).

Role of LDB1 in the transition from chromatin looping to transcription activation

Many questions remain about how close association of genes and distant enhancers occurs and how this is linked to transcription activation. In erythroid cells, lim domain binding 1 (LDB1; see Drosophila Chip) protein is recruited to the beta-globin locus via LMO2 (see Drosophila Beadex) and is required for looping of the beta-globin locus control region (LCR) to the active beta-globin promoter. This study shows that the LDB1 dimerization domain (DD) is necessary and, when fused to LMO2, sufficient to completely restore LCR-promoter looping and transcription in LDB1-depleted cells. The looping function of the DD is unique and irreplaceable by heterologous DDs. Dissection of the DD revealed distinct functional properties of conserved subdomains. Notably, a conserved helical region (DD4/5) is dispensable for LDB1 dimerization and chromatin looping but essential for transcriptional activation. DD4/5 is required for the recruitment of the coregulators FOG1 (U-shaped in Drosophila) and the nucleosome remodeling and deacetylating (NuRD) complex. Lack of DD4/5 alters histone acetylation and RNA polymerase II recruitment and results in failure of the locus to migrate to the nuclear interior, as normally occurs during erythroid maturation. These results uncouple enhancer-promoter looping from nuclear migration and transcription activation and reveal new roles for LDB1 in these processes (Krivega, 2014).

u-shaped: Biological Overview | Regulation | Developmental Biology | Effects of Mutation | References

date revised: 10 July 2000 

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