Distinct roles of Central missing and Dispatched in sending the Hedgehog signal.

Secreted Hedgehog (Hh) proteins control many aspects of growth and patterning in animal development. The mechanism by which the Hh signal is sent and transduced is still not well understood. We describe a genetic screen aimed at identifying positive regulators in the hh pathway. We recovered multiple new alleles of hh and dispatched (disp). In addition, we identified a novel component in the hh pathway, which we name central missing (cmn). Loss-of-function mutations in cmn cause similar patterning defects to those caused by hh or dispatched (disp) mutations. Moreover, cmn affects the expression of hh responsive genes but not of hh itself. Like disp, cmn acts upstream of patched (ptc) and its activity is required only in the Hh secreting cells. However, unlike disp, which is required for the release of the cholesterol-modified form of Hh, cmn regulates the activity of Hh in a manner that is independent of cholesterol modification. Finally, we show that cmn mutations bear molecular lesions in CG11495, which encodes a putative membrane bound acyltransferase related to Porcupine, a protein implicated in regulating the secretion of Wingless (Wg) signal.

Interactions with Costal2 and suppressor of fused regulate nuclear translocation and activity of cubitus interruptus.

The Hedgehog (Hh) family of secreted proteins controls many aspects of growth and patterning in animal development. In Drosophila, Hh acts by preventing the formation of a truncated repressor form of Cubitus interruptus (Ci) and stimulating the transcriptional activity of full-length Ci. Here, we provide evidence that Costal2 (Cos2) and Suppressor of Fused [Su(fu)] inhibit Ci by tethering it in the cytoplasm, whereas Hh induces nuclear translocaltion of Ci through Fused (Fu). We have identified a 125 amino acid domain in the C-terminal part of Ci that mediates response to Cos2 inhibition. We show that Cos2 binds Ci, prevents its nuclear import, and inhibits its activity via this domain. We also provide evidence that Su(fu) regulates Ci through two distinct mechanisms: (1) Su(fu) blocks Ci nuclear import through the N-terminal region of Ci, and (2) it inhibits the activity of Ci through a mechanism independent of Ci nuclear translocation. Finally, we show that Cos2 is required for transducing high levels of Hh signaling activity, and it does so by alleviating the blockage of Ci activity imposed by Su(fu).

Double-segment defining role of even-skipped homologs along the evolution of insect pattern formation.

Recent studies on insect patterning suggest that the genetic hierarchy may be roughly conserved in phylogenetically divergent species, but pair-rule genes may not function identically in all insects. In order to understand potential evolutionary changes in the role of the pair-rule genes, a Bombyx even-skipped homolog was cloned and its expression pattern during early embryogenesis studied. Eight stripes of Bombyx even-skipped were progressively expressed in an antero-posterior order. Later, these stripes disappeared anteriorly. Under this detection system, Bombyx even-skipped stripes clearly do not resolve into the corresponding secondary stripes, an obvious difference from Drosophila and Tribolium. These results suggest that Bombyx even-skipped may serve a double-segment defining role and may determine the odd-numbered engrailed stripes.

Involvement of the Bombyx Scr gene in development of the embryonic silk gland.

Homeotic selector genes determine the identity of each segment and induce the differentiation of segment-specific organs. To analyze how the silk glands of the lepidopteran, Bombyx mori, develop, we cloned and identified two genes that encode the homeodomain and its flanking regions identical to the corresponding regions of Deformed and Sex combs reduced. Using in situ hybridization and immunohistochemistry, we analyzed the expression patterns of these genes during Bombyx embryogenesis. Bombyx Deformed is expressed in the mandibular and maxillary segments, whereas expression of Bombyx Sex combs reduced is first limited to the labial segment and at later stages extended to the anterior part of the prothoracic segment. The expression of Bombyx Sex combs reduced then disappears from the invaginating placodes of silk glands where expression of Bombyx fork head/SGF-1 follows. In the Nc/Nc mutant embryos, which lack the 3' end region of Bombyx Antennapedia, in addition to the expression in the labial segment, the Bombyx Sex combs reduced is expressed ectopically in the thoracic and abdominal regions, and Bombyx fork head/SGF-1 is also ectopically expressed in the T1, T2, and T3 segments, resulting the ectopic induction of the silk gland invaginations. These results suggest that Bombyx homeobox genes such as the Bombyx Deformed and Sex combs reduced are associated with determination of the segment identities and Bombyx Sex combs reduced is involved in the induction of silk gland development.