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Whole genome sequencing of the model organisms has created increased demand for efficient tools to facilitate the genome annotation efforts. Accordingly, we report the further implementations and analyses stemming from our publicly available P[wHy] library for Drosophila melanogaster. A two-step regime--large scale transposon mutagenesis followed by hobo-induced nested deletions--allows mutation saturation and provides significant enhancements to existing genomic coverage. We previously showed that, for a given starting insert, deletion saturation is readily obtained over a 60-kb interval; here, we perform a breakdown analysis of efficiency to identify rate-limiting steps in the process. Transrecombination, the hobo-induced recombination between two P[wHy] half molecules, was shown to further expand the P[wHy] mutational range, pointing to a potent, iterative process of transrecombination-reconstitution-transrecombination for alternating between very large and very fine-grained deletions in a self-contained manner. A number of strains also showed partial or complete repression of P[wHy] markers, depending on chromosome location, whereby asymmetric marker silencing allowed continuous phenotypic detection, indicating that P[wHy]-based saturational mutagenesis should be useful for the study of heterochromatin/positional effects. DOI: 10.1534/genetics.109.103762

The spectrin cytoskeleton crosslinks actin to the membrane, and although it has been greatly studied in erythrocytes, much is unknown about its function in epithelia. We have studied the role of spectrins during epithelia morphogenesis using the Drosophila follicular epithelium (FE). As previously described, we show that α-Spectrin and β-Spectrin are essential to maintain a monolayered FE, but, contrary to previous work, spectrins are not required to control proliferation. Furthermore, spectrin mutant cells show differentiation and polarity defects only in the ectopic layers of stratified epithelia, similar to integrin mutants. Our results identify α-Spectrin and integrins as novel regulators of apical constriction-independent cell elongation, as α-Spectrin and integrin mutant cells fail to columnarize. Finally, we show that increasing and reducing the activity of the Rho1- Myosin II pathway enhances and decreases multilayering of a- Spectrin cells, respectively. Similarly, higher Myosin II activity enhances the integrin multilayering phenotype. This work identifies a primary role for α-Spectrin in controlling cell shape, perhaps by modulating actomyosin. In summary, we suggest that a functional spectrin-integrin complex is essential to balance adequate forces, in order to maintain a monolayered epithelium. [ABSTRACT FROM AUTHOR]