Istvan Török

Down-regulation of RpS21, a putative translation initiation factor interacting with P40, produces viable minute imagos and larval lethality with overgrown hematopoietic organs and imaginal discs.

ABSTRACT Down-regulation of the Drosophila ribosomal protein S21 gene (rpS21) causes a dominant weak Minutephenotype and recessively produces massive hyperplasia of the hematopoietic organs and moderate overgrowth of the imaginal discs during larval development. Here, we show that the S21 protein (RpS21) is bound to native 40S ribosomal subunits in a salt-labile association and is absent from polysomes, indicating that it acts as a translation initiation factor rather than as a core ribosomal protein. RpS21 can interact strongly with P40, a ribosomal peripheral protein encoded by the stubarista(sta) gene. Genetic studies reveal that P40 underexpression drastically enhances imaginal disc overgrowth inrpS21-deficient larvae, whereas viable combinations betweenrpS21 and sta affect the morphology of bristles, antennae, and aristae. These data demonstrate a strong interaction between components of the translation machinery and showed that their underexpression impairs the control of cell proliferation in both hematopoietic organs and imaginal discs.

Patterns of importin-α expression during Drosophila spermatogenesis

Importin-α proteins do not only mediate the nuclear import of karyophilic proteins but also regulate spindle assembly during mitosis and the assembly of ring canals during Drosophila oogenesis. Three importin-α genes are present in the genome of Drosophila. To gain further insights into their function we analysed their expression during spermatogenesis by using antibodies raised against each of the three Importin-α proteins identified in Drosophila, namely, Imp-α1, -α2, and -α3. We found that each Imp-α is expressed during a specific and limited period of spermatogenesis. Strong expression of Imp-α2 takes place in spermatogonial cells, persists in spermatocytes, and lasts up to the completion of meiosis. In growing spermatocytes, the intracellular localisation of Imp-α2 appears to be dependent upon the rate of cell growth. In pupal testes Imp-α2 is essentially present in the spermatocyte nucleus but is localised in the cytoplasm of spermatocytes from adult testes. Both Imp-α1 and -α3 expression initiates at the beginning of meiosis and ends during spermatid differentiation. Imp-α1 expression extends up to the onset of the elongation phase, whereas that of Imp-α3 persists up to the completion of nuclear condensation when the spermatids become individualised. During meiosis Imp-α1 and -α3 are dispersed in the karyoplasm where they are partially associated with the nuclear spindle, albeit not with the asters. At telophase they aggregate around the chromatin. During sperm head differentiation, both Imp-α1 and -α3 are nuclear. These data indicate that each Imp-α protein carries during Drosophila spermatogenesis distinct, albeit overlapping, functions that may involve nuclear import of proteins, microtubule organisation, and other yet unknown processes.

Drosophila spag is the homolog of RNA polymerase II-associated protein 3 (RPAP3) and recruits the heat shock proteins 70 and 90 (Hsp70 and Hsp90) during the assembly of cellular machineries

Background: Mammalian RNA polymerase II-associated protein 3 (RPAP3) recruits heat shock protein 90 (Hsp90) to assemble cellular machineries such as RNA polymerases. Results: Spaghetti encodes the Drosophila homolog of RPAP3. Spaghetti is essential for development. Spag protein binds and stimulates Hsp90 and Hsp70. Conclusion: RPAP3 function is conserved among metazoans. Significance: Our data suggest that Hsp70 assists RPAP3 in complex assembly. The R2TP is a recently identified Hsp90 co-chaperone, composed of four proteins as follows: Pih1D1, RPAP3, and the AAA+-ATPases RUVBL1 and RUVBL2. In mammals, the R2TP is involved in the biogenesis of cellular machineries such as RNA polymerases, small nucleolar ribonucleoparticles and phosphatidylinositol 3-kinase-related kinases. Here, we characterize the spaghetti (spag) gene of Drosophila, the homolog of human RPAP3. This gene plays an essential function during Drosophila development. We show that Spag protein binds Drosophila orthologs of R2TP components and Hsp90, like its yeast counterpart. Unexpectedly, Spag also interacts and stimulates the chaperone activity of Hsp70. Using null mutants and flies with inducible RNAi, we show that spaghetti is necessary for the stabilization of snoRNP core proteins and target of rapamycin activity and likely the assembly of RNA polymerase II. This work highlights the strong conservation of both the HSP90/R2TP system and its clients and further shows that Spag, unlike Saccharomyces cerevisiae Tah1, performs essential functions in metazoans. Interaction of Spag with both Hsp70 and Hsp90 suggests a model whereby R2TP would accompany clients from Hsp70 to Hsp90 to facilitate their assembly into macromolecular complexes.

Differential expression of two scribble isoforms during Drosophila embryogenesis.

The tumour suppressor gene scribble (scrib) is required for epithelial polarity and growth control in Drosophila. Here, we report the identification and embryonic expression pattern of two Scrib protein isoforms resulting from alternative splicing during scrib transcription. Both proteins are first ubiquitously expressed during early embryogenesis. Then, during morphogenesis each Scrib protein displays a specific pattern of expression in the central and peripheral nervous systems, CNS and PNS, respectively. During germ band extension, the expression of the longer form Scrib1 occurs predominantly in the neuroblasts derived from the neuro-ectoderm and becomes later restricted to CNS neurones as well as to the pole cells in the gonads. By contrast, the shorter form Scrib2 is strongly expressed in the PNS and a subset of CNS neurones.

Specific Cooperation Between Imp-α2 and Imp-β/Ketel in Spindle Assembly During Drosophila Early Nuclear Divisions.

The multifunctional factors Imp-α and Imp-β are involved in nuclear protein import, mitotic spindle dynamics, and nuclear membrane formation. Furthermore, each of the three members of the Imp-α family exerts distinct tasks during development. In Drosophila melanogaster, the imp-α2 gene is critical during oogenesis for ring canal assembly; specific mutations, which allow oogenesis to proceed normally, were found to block early embryonic mitosis. Here, we show that imp-α2 and imp-β genetically interact during early embryonic development, and we characterize the pattern of defects affecting mitosis in embryos laid by heterozygous imp-α2D14 and imp-βKetRE34 females. Embryonic development is arrested in these embryos but is unaffected in combinations between imp-βKetRE34 and null mutations in imp-α1 or imp-α3. Furthermore, the imp-α2D14/imp-βKetRE34 interaction could only be rescued by an imp-α2 transgene, albeit not imp-α1 or imp-α3, showing the exclusive imp-α2 function with imp-β. Use of transgenes carrying modifications in the major Imp-α2 domains showed the critical requirement of the nuclear localization signal binding (NLSB) site in this process. In the mutant embryos, we found metaphase-arrested mitoses made of enlarged spindles, suggesting an unrestrained activity of factors promoting spindle assembly. In accordance with this, we found that Imp-βKetRE34 and Imp-βKetD bind a high level of RanGTP/GDP, and a deletion decreasing RanGTP level suppresses the imp-βKetRE34 phenotype. These data suggest that a fine balance among Imp-α2, Imp-β, RanGTP, and the NLS cargos is critical for mitotic progression during early embryonic development.

Krebsentstehung und Differenzierung

Die Konfrontation der Wissenschaftler in der Grundlagenforschung mit dem Krebsproblem hat in den letzten Jahren zu einer Fulle neuer Erkenntnisse und zu neuen Denk- und Forschungsansatzen gefuhrt. Wahrend die Auswahl der erfolgversprechenden Forschungsansatze fruher eher ungerichtet und zufallig erfolgte, das heist von einzelnen Beobachtungen von Unterschieden normaler und maligner Zellen ausging, konnen seit einigen Jahren die Krebserkrankungen nun auch gezielt nach bestimmten grundlegenden zell- und molekularbiologischen Prinzipien erforscht werden.