BarH2 NCBI links: Precomputed BLAST | Entrez Gene | UniGene

BIOLOGICAL OVERVIEW

BarH1 and BarH2 are a pair of homeobox proteins involved in the development of the external sensilli (parts of the peripheral nervous system). These sense organs are responsive to chemical stimulation. BarH1 and BarH2 proteins are also expressed in subsets of neurons in several head segments and the developing brain. In the eye, BarH1/BarH2 proteins are expressed in a subset of photoreceptors.

In the peripheral nervous system BarH1/BarH2 expression is highest in precursors of the chemically sensitive campaniform sensilla and almost absent in precurors of the mechanically sensitive trichord sensilla. Deletion of BarH1/BarH2 in the PNS leads to a homeotic change in these organs with consequent conversion from campaniform-like sensilla to trichord sensilla. Overexpression yields the opposite result, that is, the conversion from chemical to mechanical sensilla. The strongest expression of BarH1/Bar H2 is in the thecogen cells, glial support cells for external sensory neurons. Do these cells participate in the change of fate? Current understanding is not complete. Knockout mutations of BarH2 have been generated. Such flies are perfectly viable. A double mutation in both loci is necessary to affect the transformation of sensory organs described above (Higashijima, 1992b).

Regulation of expression is likely to be complex. Since cut is vital to the develoment of the peripheral nervous system, a question arises: are the Bar genes downstream of cut in external sense organs? They are not under cut control in the brain or eye, since cut is not expressed there. Fuller understanding of basic BarH1/BarH2 expression will provide insight into the process that determines alternative fates in neurons.

The Bar homeobox genes also function as latitudinal prepattern genes in the developing Drosophila notum. In Drosophila notum, the expression of achaete-scute proneural genes and bristle formation have been shown to be regulated by putative prepattern genes expressed longitudinally. The two Bar locus genes may belong to a different class of prepattern genes expressed latitudinally: it is suggested that the developing notum consists of subdomans patterned like checkerboard squares, each subdomain governed by a different combination of prepattern genes. BarH1 and BarH2 are coexpressed in the anterior-most notal region and regulate the formation of microchaetae within the region of BarH1/BarH2 expression through activation of achaete-scute. Presutural macrochaetae formation also requires Bar gene activity. Bar gene expression is restricted in dorsal and posterior regions by Decapentaplegic signaling, while the ventral limit of the expression domain of Bar genes is determined by wingless, whose expression is under the control of Decapentaplegic signaling (M. Sato, 1999).

The Drosophila notum is considered genetically divided into several longitudinal, side by side, domains whose boundaries are determined by pannier, wingless and iroquois expression (listed respectively from medial to lateral). To further clarify relative locations of pnr, wg and iro expression areas, third-instar larval and pupal future notum were stained with various combinations of molecular markers. In larval and pupal future notum, pnr-Gal4 is expressed medially and iro-lacZ laterally. pnr-Gal4 and iro-lacZ domains partially overlap one another, and wg-lacZ (or Wg) expression is noted in the pnr-iro overlapping region and its immediate neighbors. Bar homeobox genes may belong to an additional class of notal subdivision genes. Staining for BarH1 indicates that BarH1 is expressed latitudinally (anterior vs. posterior) in the anterior-most region of future notum and postnotum. BarH1 expression begins at early to mid third instar. Anti-Ac antibody staining and neur-lacZ expression indicates PS macrochaetae are situated in the vicinity of posterior-ventral corners of the anterior BarH1 expression domain. BarH1 and BarH2 are referred to as Bar collectively and the anterior portion of the prescutum or its precursor expressing Bar is referred to as Bar prescutum. The Bar expression domain overlaps that of pnr, wg and iro. Bar expression similar to that in wing discs is observed in haltere discs (M. Sato, 1999).

For clarification of Bar's possible roles in notal development, FLP/FRT-mediated mosaic analysis was undertaken. Few microchaetae are generated in mutant clones within the prescutum. PS macrochaetae formation takes place only when PS macrochaetae formation sites are not included within mjtant clones. In flies hemizygous or homozygous for a deficency in the chromosomal region of Bar, the expression of BarH1 in the medial Bar prescutum is totally absent, implying that this particular deficiency uncovers a Bar enhancer specific to the medial Bar prescutum. In these deficiency flies, microchaetae are lost medially in the anterior three quarters of the prescutum. PS macrochaetae are lost 60% of the time. Loss of BarH1 expression and the microchaetae-less phenotype in the medial Bar prescutum are also detected in another deficiency, uncovering only BarH2 and its 3' Bar enhancer sequences. In these deficiency lines, there is no PS macrochaetae formation. Taken together, these results suggest that Bar genes are essential for bristle formation in the Bar prescutum. Should Bar homeobox genes be redundant in function and involved in bristle formation in the Bar prescutum, bristle defects would certainly be rescued by Bar targeted expression. These considerations were confirmed through the use of the GAL4/UAS system. Loss of microchaetae but not PS macrochaetae is virtually restored by the targeted expression of Bar genes. BarH1 and BarH2 would appear functionally redundant to each other and essential for microchaetae formation in the medial Bar prescutum and PS macrochaetae formation. While searching for Bar enhancers, two notum enhancers have been identified. S8 is found responsible for Bar expression in the medial Bar prescutum. A second enhancer, B4.5, is capable of driving reporter gene (lacZ) expression in the lateral Bar prescutum in wild-type background (M. Sato, 1999).

To assess the capability of Bar for inducing microchaetae formation, hs-BarH1 or hs-BarH2 transgenes were heat-induced during larval or pupal development and the same results were obtained for each. Ectopic microchaetae are generated in the scutellum, wing blades and head capsule. In the scutellum (normally possessing no microchaetae) 30- 50 ectopic microchaetae are generated by a heat-shock at 6 hours APF (after puparium formation), but few ectopic microchaetae are formed by heat shock before 2 hours APF or after 12 hours APF. In contrast to ectopic microchaetae formation in neurogenic mutations, extra microchaetae induced by hs-Bar are unclustered, suggesting that Bar acts as an activator of proneural genes. Staining for Ac shows that pupal notum-specific Ac expression begins in characteristic regions at 6 hours APF, peaking at 8 hours APF and eventually disappearing at 12 hours APF. These Ac-positive regions appear to correspond to microchaetae proneural regions since microchaetae SOP formation starts at 8 hours APF. Macrochaetae SOP are formed during third instar. In the Bar prescutum, the area of Ac expression is seen to overlap that of Bar. Sc expression is quite similar, if not identical, to Ac expression. Thus, a study was made to find whether Bar is capable of acting as an activator of ac-sc to control microchaetae formation. Ac expression is almost entirely absent from the Bar enhancer deficency medial Bar prescutum, where no microchaetae formation takes place. Following induction of hs-Bar, not only ectopic microchaetae formation but strong Ac (and Sc) expression is apparent in the scutellum. hs-Bar dependent microchaetae formation is significantly suppressed in ac-sc hypomorphic mutant backgrounds, while bristle defects including PS macrochaetae loss in Bar enhance deficiency are eliminated by a gain-of-function allele of ac-sc. Bar may thus be considered to be an activator of ac-sc essential for producing proneural clusters for microchaetae and possibly PS macrochaetae as well (M. Sato, 1999).

During late third instar, the expression domain of Bar in the prescutum is immediately adjacent to dpp and wg expression domains. dpp likely regulates Bar expression negatively. When dpp is expressed throughout the notum using UAS-dpp driven by ap-Gal4, Bar expression is totally abolished. Conversely, reduction in dpp activity ectopically induces Bar expression in the medial region of the future notum. Similar medial expansion of Bar expression is observed subsequent to reduction in the activity of hedgehog (hh), an inducer of dpp. Possible effects of wg on Bar expression were sought using wg^ts, in which Wg secretion but not production is temperature-sensitive. Bar expression in the lateral Bar prescutum is abolished after 48 hours (but not 24 hours) incubation at the restrictive temperature. armadillo (arm), a beta-catenin homolog, is a signal transducer of Wg signaling. Bar expression is lost in clones mutant for arm when generated in the lateral Bar prescutum, while Bar misexpression is present in lateral prescutum clones expressing a constitutively active form of arm. It is thus concluded that Bar expression in lateral prescutum is requires Wg signals, whose levels determine the ventral border of the Bar prescutum. During late third instar, the expression domain of Bar in the prescutum overlaps with those of pnr and iro. Since no appreciable change in Bar expression is detected in flies mutant for iro or pnr, Bar expression may be regulated independent of pnr and iro. Bar may be partially involved in wg repression (M. Sato, 1999).

It is concluded that a checker-board-like subdivision of future notum is regulated by putative prepattern gene expression. Future notum may be divided into square subdomains in a checker-board-like manner, each with its own unique combinations of prepattern gene expression. Putative prepattern genes, iro and pnr, form longitudinal domains. Bar homeobox genes form the anterior-most domain. This is the first demonstration of the presence of latitudinal, front to back, prepattern genes in the notum. Bristle formation in each subdomain may be positively regulated by a region-specific combination of prepattern genes. Consistent with this, microchaetae formation in the anterolateral prescutum (the lateral Bar prescutum), where Bar and iro are coexpressed, requires the concerted action of Bar and iro (M. Sato, 1999).

GENE STRUCTURE

BarH1

cDNA clone length - 3.1 kb

Bases in 5' UTR - 633

Exons - three

Bases in 3' UTR - 767

BarH2

cDNA clone length - 3.75 Kb

Bases in 5' UTR - 968

Exons - three

Bases in 3' UTR - 783

PROTEIN STRUCTURE

Amino Acids - 543 BarH1; 604 BarH2

Structural Domains

Except for one amino acid substitution, BarH1 and BarH2 homeodomains are identical in sequence. The phenylalanine residue in helix 3, conserved in all metazoan homeodomains examined thus far, is replaced by a tyrosine residue (Kojima, 1991).

date revised: 3 April 99

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