Genetic evidence for interaction between eta- and beta-tubulins.

The thermosensitive allelic mutations sm19-1 and sm19-2 of Paramecium tetraurelia cause defective basal body duplication: growth at the nonpermissive temperature yields smaller and smaller cells with fewer and fewer basal bodies. Complementation cloning of the SM19 gene identified a new tubulin, eta-tubulin, showing low homology with each of the other five tubulins, alpha to epsilon, characterized in P. tetraurelia. In order to analyze eta-tubulin functions, we used a genetic approach to identify interacting molecules. Among a series of extragenic suppressors of the sm19-1 mutation, the su3-1 mutation was characterized as an E288K substitution in the beta-PT2 gene coding for a beta-tubulin, while the mutation nocr1 conferring nocodazole resistance and localized in another beta-tubulin gene, beta-PT3, was shown to enhance the mutant phenotype. The interaction between eta-tubulin and microtubules, revealed by genetic data, is supported by two further types of evidence: first, the mutant phenotype is rescued by taxol, which stabilizes microtubules; second, molecular modeling suggests that eta-tubulin, like gamma- and delta-tubulins, might be a microtubule minus-end capping molecule. The likely function of eta-tubulin as part of a complex specifically involved in basal body biogenesis is discussed.

Needs and targets for the multi sex combs gene product in Drosophila melanogaster.

We have analyzed the requirements for the multi sex combs (mxc) gene during development to gain further insight into the mechanisms and developmental processes that depend on the important trans-regulators forming the Polycomb group (PcG) in Drosophila melanogaster. mxc is allelic with the tumor suppressor locus lethal (1) malignant blood neoplasm (l(1)mbn). We show that the mxc product is dramatically needed in most tissues because its loss leads to cell death after a few divisions. mxc has also a strong maternal effect. We find that hypomorphic mxc mutations enhance other PcG gene mutant phenotypes and cause ectopic expression of homeotic genes, confirming that PcG products are cooperatively involved in repression of selector genes outside their normal expression domains. We also demonstrate that the mxc product is needed for imaginal head specification, through regulation of the ANT-C gene Deformed. Our analysis reveals that mxc is involved in the maternal control of early zygotic gap gene expression previously reported for some PcG genes and suggests that the mechanism of this early PcG function could be different from the PcG-mediated regulation of homeotic selector genes later in development. We discuss these data in view of the numerous functions of PcG genes during development.

pen repeat sequences are GGN clusters and encode a glycine-rich domain in a Drosophila cDNA homologous to the rat helix destabilizing protein.

Several cDNA clones that contain the pen repeat have been isolated and sequenced; pen consists of clusters of GGN triplets, where N can be any nucleotide. Some of the pen repeat sequences are found within long open reading frames in which they encode oligoglycine stretches. For one of the clones, the deduced amino acid sequence of the entire open reading frame, especially in the region preceding the glycine-rich domain, shows strong homology to the rat helix destabilizing protein [Cobianchi, F., SenGupta, D. N., Zmudzka, B. Z. & Wilson, S. H. (1986) J. Biol. Chem. 261, 3536-3543]. The rat protein and homologs in other organisms are single-stranded nucleic acid binding proteins, some of which are major components of heterogeneous nuclear ribonucleoprotein particles. We suggest that we have cloned a cDNA encoding a Drosophila single-stranded nucleic acid binding protein.

Genetic and molecular analysis of fs(1)h, a maternal effect homeotic gene in Drosophila.

Mutations at the Drosophila melanogaster locus female sterile (1) homeotic (fs(1)h) result in segmental abnormalities including missing organs and homeotic transformations in the progeny of mutant mothers. Homeotic transformations are enhanced when the zygotes carry one of several third chromosome mutations, specifically alleles or deficiencies of the trithorax (trx) locus, also called Regulator-of-bithorax, and some alleles of bithorax complex (BX-C) genes. These observations suggest that maternally derived fs(1)h+ product is required, in interaction with trx and BX-C genes, for normal segment specification. The fs(1)h gene and an adjacent gene, lethal (1) myospheroid (l(1)mys), have been cloned by chromosomal walking. Mutations of fs(1)h were found within a 13-kb stretch of DNA. Poly(A)+ RNAs migrating as a doublet at 7.6 kb and a single band at 5.9 kb, which are homologous to the fs(1)h+ chromosomal region, are found in ovaries and early embryos. The largest RNAs are derived from a 20-kb chromosomal region encompassing the sites of all mapped fs(1)h alleles.

The role of dosage of the region 7D1-7D5-6 of the X chromosome in the production of homeotic transformations in Drosophila melanogaster.

A high frequency of homeotic transformation appears in Df(3)red/+ progeny of Df(1)snC128/+ females. Generally, the metathoracic appendages are partially transformed into mesothoracic ones. Df(1)snC128 includes a small region of the X chromosome: 7D1 to 7D5-6. Hypodosage of this region is mainly effective at the level of the maternal genotype, and the effect is probably due to hypodosage of the wild-type allele of the gene fs(1)h. Df(3)red has an effect that is mainly, if not exclusively, zygotic, probably due to hypodosage of the wild-type allele of Rg-bx. The frequencies of transformed flies resulting from the interaction between Df(1)snC128 and Df(3)red are not very sensitive to external conditions and genetic background. Studies of the interactions between Df(1)snC128 and other mutations or deficiencies of chromosome 3 [Rg-pbx, bx, pbx, Ubx1, Ubx130, Ubx80, Df(3)P9] reveal an analogy between the hypodosage effect of region 7D1-7D5-6 and the effects of ether treatment of blastoderm stage eggs. The role of the gene fs(1)h in the process of segment determination is discussed in the light of these results.