ribbon: Biological Overview | Developmental Biology | Effects of Mutation | References
Gene name - ribbon

Synonyms -

Cytological map position - 56C6--9

Function - transcription factor

Keywords - tracheal migration,
dorsal closure, epithelial migration

Symbol - rib

FlyBase ID: FBgn0003254

Genetic map position - 2-88

Classification - BTB/POZ domain

Cellular location - nuclear

NCBI links: Precomputed BLAST | Entrez Gene | UniGene |
Recent literature
Loganathan, R., Lee, J. S., Wells, M. B., Grevengoed, E., Slattery, M. and Andrew, D. J. (2015). Ribbon regulates morphogenesis of the Drosophila embryonic salivary gland through transcriptional activation and repression. Dev Biol. PubMed ID: 26477561
Ribbon (Rib) controls cell shape/volume increases during elongation of the Drosophila salivary gland (SG), without effects on general SG cell attributes such as specification, proliferation and apoptosis and without compromising epithelial-specific morphological attributes. To identify the genes regulated by Rib, ChIP-seq analysis was performed in embryos driving expression of GFP-tagged Rib specifically in the SGs. Microarray analysis compared RNA samples from age-matched wild-type and rib null embryos. From the superposed ChIP-seq and microarray gene expression data, 60 genomic sites bound by Rib were identified that were likely to regulate SG-specific gene expression. Several of the identified Rib targets were identified by qRT-pCR and/or in situ hybridization. The results indicate that Rib regulates cell growth and tissue shape via a diverse array of targets through both transcriptional activation and repression. Furthermore, the results suggest that autoregulation of rib expression may be a key component of the SG morphogenetic gene network.
Silva, D., Olsen, K. W., Bednarz, M. N., Droste, A., Lenkeit, C. P., Chaharbakhshi, E., Temple-Wood, E. R. and Jemc, J. C. (2016). Regulation of gonad morphogenesis in Drosophila melanogaster by BTB family transcription factors. PLoS One 11(11): e0167283. PubMed ID: 27898696
During embryogenesis, primordial germ cells (PGCs) and somatic gonadal precursor cells (SGPs) migrate and coalesce to form the early gonad. A failure of the PGCs and SGPs to form a gonad with the proper architecture not only affects germ cell development, but can also lead to infertility. Therefore, it is critical to identify the molecular mechanisms that function within both the PGCs and SGPs to promote gonad morphogenesis. This study has characterized the phenotypes of two genes, longitudinals lacking (lola) and ribbon (rib), that are required for the coalescence and compaction of the embryonic gonad in Drosophila melanogaster. rib and lola are expressed in the SGPs of the developing gonad, and genetic interaction analysis suggests these proteins cooperate to regulate gonad development. Both genes encode proteins with DNA binding motifs and a conserved protein-protein interaction domain, known as the Broad complex, Tramtrack, Bric-a-brac (BTB) domain. Through molecular modeling and yeast-two hybrid studies, it was demonstrated that Rib and Lola homo- and heterodimerize via their BTB domains. In addition, analysis of the colocalization of Rib and Lola with marks of transcriptional activation and repression on polytene chromosomes reveals that Rib and Lola colocalize with both repressive and activating marks and with each other. While previous studies have identified Rib and Lola targets in other tissues, Rib and Lola are likely to function via different downstream targets in the gonad. These results suggest that Rib and Lola act as dual-function transcription factors to cooperatively regulate embryonic gonad morphogenesis.

During development of the Drosophila tracheal (respiratory) system, the cell bodies and apical and basal surfaces of the tracheal epithelium normally move in concert as new branches bud and grow out to form tubes. Mutations in the ribbon (rib) gene disrupt this coupling: the basal surface continues to extend towards its normal targets, but movement and morphogenesis of the tracheal cell bodies and apical surface is severely impaired, resulting in long basal membrane protrusions but little net movement or branch formation. rib mutant tracheal cells are still responsive to the Branchless fibroblast growth factor (FGF) that guides branch outgrowth, and they express apical membrane markers normally. This suggests that the defect lies either in transmission of the FGF signal from the basal surface to the rest of the cell or in the apical cell migration and tubulogenesis machinery. rib encodes a nuclear protein with a BTB/POZ domain and Pipsqueak DNA-binding motif. It is expressed in the developing tracheal system and other morphogenetically active epithelia. Directed cell migration of the salivary gland and dorsal epidermis are also affected in ribbon mutants. It is proposed that Rib is a key regulator of epithelial morphogenesis that promotes migration and morphogenesis of the tracheal cell bodies and apical surface and other morphogenetic movements (Shim, 2001; Bradley, 2001).

In the wild type migrating tracheal branch, the basal surface of the tracheal epithelium is broad and smooth with an occasional pseudopodium extending from cells at the growing tip, much like those seen at the leading edge of migrating fibroblasts. As the basal surface extends toward the Bnl FGF signaling centers, the cell bodies and apical surface follow, and basal cytoplasmic extensions are never very prominent. In rib mutants, pseudopodia still extend from the basal surface, and are more numerous and pronounced than in wild type, occasionally forming extremely long processes that appear to reach their normal targets. This dissociation of the migration of the apical and basal tracheal surfaces is evident at stage 12 and continues for several hours into stage 14. At this later stage, the apical surface begins to deform in the direction of the basal cytoplasmic extensions, but even at this later stage the net distance traveled by the apical side is far less than in wild type. The apical defect is unlikely to reflect a general defect in apical-basal polarity of the tracheal epithelium because the apical determinant Crumbs, the apical marker TL1, and an apically localized mRNA are expressed and localize properly at the apical tracheal surface in rib mutants. It is concluded that rib mutations selectively affect movement and morphogenesis of the tracheal cell bodies and apical surface (Shim, 2001).

In ribbon mutant tracheae, the dorsal trunk fails to form, and ventral branches are stunted; however, directed migrations of the dorsal and visceral branches are largely unaffected. The dorsal trunk also fails to form when FGF or Wingless/WNT signaling is lost, and ribbon functions downstream of, or parallel to, these pathways to promote anterior-posterior migration. Directed cell migration of the salivary gland and dorsal epidermis are also affected in ribbon mutants, suggesting that conserved mechanisms may be employed to orient cell migrations in multiple tissues during development (Bradley, 2001).

Analysis of Wg signaling in tracheal branching suggests that cells are allocated to branches (cell allocation) independently from cell fate specification. (1) In Wingless signaling mutants the 'pre-dorsal trunk' cells are positioned correctly, but fail to migrate away from the transverse connective. (2) Wg signaling mutants do not express spalt, a dorsal trunk-specific marker. Thus, the cells are allocated to the dorsal trunk (DT), but do not express DT markers or behave like dorsal trunk cells. rib mutants, like Wg signaling mutants, also fail to form the DT, and 'pre-DT' cells are stalled at the transverse connective; however, unlike embryos lacking Wg signaling, rib mutants express sal in DT cells. Thus, rib is not required for cell allocation or cell fate specification (as monitored by sal), but is only required for branch migration. In summary, these observations suggest that, at least for the tracheal DT, cell allocation is independent of cell fate specification, and cell fate can be further subdivided into branch identity (controlled by genes such as sal that specify branch features) and branch migration, which involves rib (Bradley, 2001).

The similarity of the tracheal DT phenotypes in rib mutants and Wg signaling mutants raises the possibility that rib functions with Wg signaling for migration of DT cells. sal is the only known early downstream target of Wg signaling in the DT. Because the DT phenotype is more severe in embryos lacking Wg signaling than in sal mutants, there must be additional downstream targets of Wg signaling. Indeed, it can be predicted that these other genes control migration based on two findings. (1) DT cells are capable of migrating in sal mutants, but move in the wrong direction (dorsally). (2) When both Wg and Decapentaplegic signaling are activated in wild-type embryos (activated armadillo and activated thick veins in all tracheal cells), a complete longitudinal DT forms that does not express sal, suggests that sal may be dispensable for anteroposterior migration in some cases. Loss of rib results in a DT phenotype identical to that observed in loss of Wg signaling and rib functions in parallel to Wg-dependent sal expression. Together these results suggest that rib is working with Wg signaling, either in parallel or potentially as a downstream target, to direct DT migration (Bradley, 2001).

It is hypothesized that rib may respond to signals from multiple pathways based on analysis of a ventral cuticle phenotype. In rib mutants, the defects in ventral cuticle patterning appear most similar to the phenotype reported for the combined loss of late Wg signaling and Egfr signaling. In this tissue, rib could be integrating signaling from Wg and Egfr. In several other tissues requiring rib function, Wg signaling and signaling through a MAPK cascade are also required; however, in these cases, loss of either of the individual pathways results in phenotypes similar to those of rib mutants. For instance, rib is required for the cell shape changes in the leading edge cells during dorsal closure, a process that requires both Wg signaling and Jnk signaling. The second midgut constriction and the morphogenesis of the Malpighian tubules are defective in rib mutants, and both events also require both Wg and Egfr signaling. Similarly, in the trachea, rib could respond to Wg signaling and either of the two pathways (FGF or Egfr) that activate the MAPK cascade in tracheal cells. Since the rib phenotype is distinct from Egfr signaling mutants, a role for rib downstream of FGF signaling is favored. Indeed, the stalled outgrowth of all tracheal branches and stunted ventral branches observed in rib mutants may be linked to FGF signaling. Consistent with the idea that rib responds to MAPK signaling, the Rib protein has seven consensus MAPK phosphorylation sites (Bradley, 2001).

rib identifies a new step in primary branch budding and outgrowth that lies downstream or parallel to bnl signaling. What is this step? During primary branch budding, the basal surface of the tracheal epithelium is exposed to secreted Bnl/FGF, and the cells respond by extending pioneer cytoplasmic processes towards the Bnl source. But for a new branch to form, the cell bodies and apical surface must follow. Unlike isolated cells like Dictyostelium or fibroblasts migrating up a chemoattractant gradient, where the entire cell is exposed to the attractant, the cell bodies and apical surface of the tracheal epithelium are sealed off and do not have direct access to Bnl. These parts of the cell must receive the stimulus to move indirectly. rib might be required for transmission of the Bnl signal from the basal surface to the rest of the cell or to otherwise couple their movement. Alternatively, rib may be needed to propel the cell body forward once it receives the signal to move. Fibroblasts use distinct molecular mechanisms to move their leading and trailing sides forward; perhaps rib is required for a myosin-dependent process, like the one used to propel the trailing side of a migrating fibroblast forward. Rib might also be needed to promote the cell shape and apical surface changes necessary to form or extend a new tube. Although the data do not pinpoint the precise cellular event mediated by Rib, it is clear that the gene identifies a distinct step in tracheal branching – one that may be common to a number of other epithelial morphogenetic events (Shim, 2001).

The genetic analysis indicates that rib functions downstream or parallel to the Bnl FGF pathway. If Rib functions downstream in the FGF pathway, for example, if its transcriptional regulatory activity is regulated by RAS/MAPK signaling like the Yan, DSRF and possibly Pointed transcription complexes, then this would provide a natural way of coupling morphogenetic events in the tracheal cell bodies and apical surface to events at the basal surface, where the Bnl signal is received. When the Bnl pathway is active, the receptor would directly stimulate outgrowth of the basal tracheal surface. But it would also indirectly stimulate migration and morphogenesis of the cell body and apical surface, via activation of Rib and induction of its target genes (Shim, 2001).

Rib, together with the Drosophila proteins Pipsqueak (Psq) and Tyrosine kinase related (Tkr), defines a new subfamily of BTB proteins containing Psq DNA-binding motifs. Although most other BTB domain proteins are also believed to function as transcription factors, this much larger subfamily of BTB domain proteins contains zinc-finger (ZF) DNA-binding motifs. The BTB domains of BTB/Psq proteins are no more similar to each other than they are to those of BTB/ZF proteins, although they are more similar to each other than they are to the BTB domain of Kelch, a cytoplasmic protein. The reason for the common coupling of BTB domains with two structurally unrelated DNA binding motifs, but not other DNA binding motifs, is unclear (Shim, 2001).

ribbon is thought to be required for generating specialized cell shapes. For instance, during dorsal closure, leading edge cells of the lateral epidermis fail to elongate in rib mutants. rib mutants also show abnormal dilation of salivary gland lumina in late embryogenesis, suggesting that either rib is also required at late stages to maintain organ shape or loss of early rib function indirectly causes the late lumenal dilation. rib appears to control cell shapes by regulating the cytoskeleton. During dorsal closure, a band of actin and myosin forms at the dorsal margin of leading edge cells. In rib embryos, the actin band is narrower and myosin heavy chain (MHC) is absent from leading edge cells. Thus, rib may be required for the localization or organization of cytoskeletal components. zip encodes a nonmuscle MHC and is required in many of the same tissues as rib; however, strong loss-of-function mutations in zip suppress the distended lumenal phenotype of rib salivary glands, suggesting that rib does not positively regulate myosin activities. Instead, rib may repress myosin contraction or regulate the direction of contraction, perhaps by providing a balancing force to the direction of basal myosin contractions. These studies reveal a role for rib in coordinating directed cell migration, a process that clearly involves actin/myosin dynamics. Thus, rib may modulate actin/myosin behavior for cell movement and cell shape during both tissue formation and tissue homeostasis. If rib is responding to signaling pathways, rib could be a critical factor linking signaling events to changes in the cytoskeleton (Bradley, 2001).

In addition to the tracheal system, rib is expressed in a number of other developing tissues, in a complex and dynamic pattern that coincides with various morphogenetic movements. Several sites of expression coincide with epithelial invaginations or evaginations to form tubes or sacs, as occurs during tracheal branch budding. These sites include the anterior and posterior midgut invaginations , salivary gland primordia as they invaginate and extend, stomodeum and proctodeum as they invaginate to form the foregut and hindgut, and Malpighian (renal) tubules as they bud. rib is also expressed during spreading of a planar epithelium, the epidermis, during dorsal closure. rib is expressed during or just after a number of mesenchymal-epithelial transitions, including the midgut mesenchyme as it reorganizes into an epithelium and forms the central portion of the gut (Shim, 2001).

Several of these morphogenetic events are defective in rib mutants. Indeed, rib mutants were first identified by their dorsal closure defect (Nusslein-Volhard, 1984). rib mutants also have defects in a number of other tissues including the Malpighian tubules, salivary glands and hindgut (Jack, 1997; Blake, 1998; Blake, 1999). Each of these are epithelia, and there are defects in the cell shape changes that underlie their morphogenesis. For example, rib mutant epidermal cells fail to elongate properly, and salivary gland cells fail to constrict apically (Shim, 2001).

Almost all epithelia undergoing morphogenetic movements face similar challenges as the tracheal epithelium during primary branching -- different parts of the epithelium must move coordinately although they are exposed to different environments. Based on the analysis of the tracheal function of rib, as well as its expression and activity in a variety of other morphogenetic processes, it is proposed that Rib is a key regulator of the movement and morphogenesis of epithelia, particularly events in the cell bodies or apical surface during budding of tubular epithelia. In some migrating epithelia, such as the spreading epidermis during dorsal closure, it is not clear that rib mutations specifically disrupt an apical process, so Rib may also influence epithelial migration and morphogenesis in other ways. Insight into the mechanisms and molecules that execute these morphogenetic events may come through the identification of transcriptional targets of Rib (Shim, 2001).


cDNA clone length - 3939

Bases in 5' UTR - 671

Exons - 2

Bases in 3' UTR - 1282


Amino Acids - 661

Structural Domains

Blast sequence homology searches with the deduced Rib protein sequence identified a region similar to BTB (or POZ for Poxvirus and zinc finger) domains, a protein-protein interaction domain found in zinc finger transcription factors and actin-associated proteins of the Drosophila Kelch family. Like other BTB domains, the BTB domain in Ribbon consists of ~120 residues and is located close to the N terminus. It shares 35%-38% identity (~60% similarity) with three founding members of the BTB family: the Drosophila developmental regulatory proteins Bric à brac, Tramtrack and Broad-complex transcription factor. The BTB domain of Rib is slightly more similar (40% identity, 63% similarity) to the BTB domain of longitudinals lacking (lola), a zinc-finger nuclear factor implicated in axon growth and targeting in the Drosophila nervous system. Other BTB domains show less identity, such as Kelch (25% identity) and human promyelocytic leukemia zinc finger (PLZF) protein (24% identity) (Shim, 2001).

Many BTB domain proteins also contain a zinc-finger DNA-binding domain; no zinc finger motif was identified in Rib. However, Blast/BEAUTY searches with the Rib-coding region revealed a 49 residue region (residues 367 to 415) with similarity to the DNA-binding domain of Pipsqueak, a protein involved in patterning the early Drosophila egg, embryo and adult eye. The Psq DNA binding domain consists of four tandem repeats of an ~50 amino acid motif, each of which is 30%-48% identical to each other and 28%-36% identical to the region in Rib. The Psq repeats also show similarity to the helix-turn-helix DNA-binding domain of a number of prokaryotic recombinases including E. coli Hin Invertase. A single Psq motif is found in Drosophila Tyrosine kinase related protein. Interestingly, both Psq and Tkr also contain N-terminal BTB domains (36% identity between the Psq and Rib BTB domains; 32% between Tkr and Rib). Psq, Rib, and Tkr thus define a new subtype of BTB domain proteins with a Psq DNA-binding motif (Shim, 2001).

rib encodes a 1983 nt ORF corresponding to a 661-residue protein with an amino- (N) terminal BTB/POZ (Bric a brac, Tramtrack, Broad-Complex/Poxvirus, zinc finger) domain. The BTB/POZ domain is an evolutionarily conserved domain that mediates homo- or hetero-dimerization with other BTB/POZ domains and is found in over 400 proteins. A short coiled-coil region in the carboxy (C) terminus is predicted. Rib has four consensus nuclear localization signals (NLS), two of which are bipartite, and Rib is predicted to be nuclear by the PSORT program. Thus, Rib may function in the nucleus. Rib also contains consensus phosphorylation sites for a number of kinases that mediate the formation of tissues in which rib is required (e.g., MAPK). Consensus glycosylation, myristylation and amidation sites are also present in the rib ORF (Bradley, 2001).

The rib gene encodes a novel protein with two protein-protein interaction domains, an N-terminal BTB/POZ domain and a C-terminal coiled-coil region. The BTB/POZ domain mediates dimerization, and BTB/POZ proteins often contain additional domains that define protein function and/or subcellular localization. Many BTB/POZ proteins contain multiple DNA binding zinc fingers and function as transcriptional regulators. For example, the Drosophila Tramtrack protein is required to represses transcription of pair-rule genes in early embryogenesis. BTB/POZ domain proteins can also mediate cytoskeletal organization. For instance, the Drosophila Kelch protein, which oligomerizes via its BTB domain and binds actin through its kelch domains, is required to maintain cytoskeletal organization of ring canals during oogenesis. BTB/POZ proteins can also function outside the cell; the mammalian BTB/POZ protein Mac-2 binding protein (M2BP) localizes to the extracellular matrix (ECM) and forms multivalent ring structures proposed to be important for its interactions with collagens IV, V and VI, fibronectin, and other ECM proteins. One family of Arabidopsis BTB/POZ-containing proteins has a composition very similar to that of RIB: a BTB/POZ domain at the N-terminus and a coiled-coil at the C terminus. One family member, RPT2, appears to respond to signals that promote phototropism. RPT2 also contains an NLS; however, it is not yet known where RPT2 functions. Based on the four putative NLSs, it is speculated that RIB may function in the nucleus, where it would be positioned to regulate the expression of genes required for cytoskeletal changes during morphogenesis. Alternatively, RIB may reside in the cytoplasm and more directly regulate cytoskeletal organization. Since BTB/POZ domains can heterodimerize, RIB may have a partner(s) providing additional functional motifs. rib RNA is expressed in a dynamic pattern during development, including expression in cells that appear phenotypically normal in rib embryos. Thus, rib function is likely to be post-transcriptionally regulated, perhaps through the phosphorylation of its MAPK sites or through limited expression or activation of cofactors (Bradley, 2001).

ribbon: Biological Overview | Developmental Biology | Effects of Mutation | References

date revised: 30 January 2002

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