In the developing tracheal system of Drosophila melanogaster, six major branches arise by guided cell migration from a sac-like structure. The chemoattractant Branchless/FGF (Bnl) appears to guide cell migration and is essential for the formation of all tracheal branches, while Decapentaplegic (Dpp) signaling is strictly required for the formation of a subset of branches, the dorsal and ventral branches. Using in vivo confocal video microscopy, we find that the two signaling systems affect different cellular functions required for branching morphogenesis. Bnl/FGF signaling affects the formation of dynamic filopodia, possibly controlling cytoskeletal activity and motility as such, and Dpp controls cellular functions allowing branch morphogenesis and outgrowth.

The measuring tip of a force microscope can be converted into a monomolecular sensorif one or few "ligand" molecules are attached to the apex of the tip while maintainingligand function. Functionalized tips are used to study fine details of receptor-ligand interactionby force spectroscopy or to map cognate "receptor" molecules on the sample surface. Thereceptor (or target) molecules can be present on the surface of a biological specimen; alternatively,soluble target molecules must be immobilized on ultraflat supports. This review describes the methodsof tip functionalization, as well as target molecule immobilization. Silicon nitride tips, siliconchips, and mica have usually been functionalized in three steps: (1) aminofunctionalization,(2) crosslinker attachment, and (3) ligand/receptor coupling, whereby numerous crosslinkersare available to couple widely different ligand molecules. Gold-covered tips and/or supports haveusually been coated with a self-assembled monolayer, on top of which the ligand/receptor moleculehas been coupled either directly or via a crosslinker molecule. Apart from these general strategies,many simplified methods have been used for tip and/or support functionalization, even single-stepmethods such as adsorption or chemisorption being very efficient under suitable circumstances. Allmethods are described with the same explicitness and critical parameters are discussed. In conclusion,this review should help to find suitable methods for specific problems of tip and support functionalization.

The 5' regulatory region of the chh2 gene, encoding cellobiohydrolase II, of the filamentous fungus Hypocrea jecorina contains the cbh2 activating element (CAE) which is essential for cbh2 expression. The CAE consists of two separate, adjacent motifs, a CCAAT box on the template strand (ATTGG) and a GTAATA box on the coding strand, which co-operate in the induction of the gene by cellulose or sophorose. EMSA supershift experiments using an antibody against Aspergillus nidulans HAPC suggested that the complex which binds to the H. jecorina CCAAT box contains a HAPC homolog. To obtain direct evidence for this, we have cloned the hap2, hap3 and hap5 genes from H. jecorina. They encode proteins whose core regions display great similarity to Aspergillus HAPB, HAPC and HAPE and to known HAP homologs from other organisms. All three genes are transcribed in a carbon source-independent manner. A. nidulans deltahap strains were functionally complemented in vitro by the overexpressed H. jecorina HAP2, HAP3 and HAP5 proteins, and they thus represent subunits of the CCAAT-binding complex. Furthermore, all three proteins (HAP2, HAP3 and HAP5) were needed to bind to the CAE in the H. jecorina cbh2 gene promoter in vitro. We conclude that the CCAAT box on the template strand in CAE is bound by the H. jecorina equivalent of the HAP protein complex.

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