Gerold Schubiger

Parameters controlling transcriptional activation during early drosophila development

We studied transcription during the first 14 mitotic cycles of Drosophila development, by gel electrophoresis of RNA pulse-labeled in vivo. Synthesis of rRNA, tRNAs, 5S RNAs, snRNAs, poly(A)+ RNAs, and histone mRNAs is first detectable during cycle 11 or 12. Histone genes are transcribed during S phases, and reach maximal activation in cycle 12, whereas nonhistone genes are transcribed only in G2 periods, and reach maximal activation during late cycle 14. The high transcriptional activity characteristic of cycle 14 can be precociously induced by extending interphase with cycloheximide as early as, but not before, cycle 10. We conclude that all classes of genes become competent for activation during cycle 10, and that subsequent activation is differentially suppressed by functions associated with nuclear division.

Cell cycle control by the nucleo-cytoplasmic ratio in early Drosophila development.

We have studied the role of the nucleo-cytoplasmic ratio in the early development of Drosophila, using mutants and experimental manipulations that alter nuclear density. Haploid embryos produced by either maternal or paternal effect mutations compensate for haploidy by an extra nuclear division during the syncytial blastoderm stage. Decreasing the nucleo-cytoplasmic ratio in wild-type embryos by ligation can cause a similar extra blastoderm division. Conversely, increasing this ratio can cause the omission of a blastoderm division. The duration of mitotic cycles is affected by the nucleo-cytoplasmic ratio four cycles before the terminal blastoderm division. Transcription patterns in haploid embryos indicate that transcriptional activation is not directly controlled by the nucleo-cytoplasmic ratio, but may be an effect of the lengthening of interphase periods.

Regulation of cellular plasticity in Drosophila imaginal disc cells by the Polycomb group, trithorax group and lama genes

Drosophila imaginal disc cells can switch fates by transdetermining from one determined state to another. We analyzed the expression profiles of cells induced by ectopic Wingless expression to transdetermine from leg to wing by dissecting transdetermined cells and hybridizing probes generated by linear RNA amplification to DNA microarrays. Changes in expression levels implicated a number of genes: lamina ancestor, CG12534 (a gene orthologous to mouse augmenter of liver regeneration), Notch pathway members, and the Polycomb and trithorax groups of chromatin regulators. Functional tests revealed that transdetermination was significantly affected in mutants for lama and seven different PcG and trxG genes. These results validate our methods for expression profiling as a way to analyze developmental programs, and show that modifications to chromatin structure are key to changes in cell fate. Our findings are likely to be relevant to the mechanisms that lead to disease when homologs of Wingless are expressed at abnormal levels and to the manifestation of pluripotency of stem cells.

Peripodial Cells Regulate Proliferation and Patterning of Drosophila Imaginal Discs

Cells employ a diverse array of signaling mechanisms to establish spatial patterns during development. Nowhere is this better understood than in Drosophila, where the limbs and eyes arise from discrete epithelial sacs called imaginal discs. Molecular-genetic analyses of pattern formation have generally treated discs as single epithelial sheets. Anatomically, however, discs comprise a columnar cell monolayer covered by a squamous epithelium known as the peripodial membrane. Here we demonstrate that during development, peripodial cells signal to disc columnar cells via microtubule-based apical extensions. Ablation and targeted gene misexpression experiments demonstrate that peripodial cell signaling contributes to growth control and pattern formation in the eye and wing primordia. These findings challenge the traditional view of discs as monolayers and provide foundational evidence for peripodial cell function in Drosophila appendage development.

Temporal regulation in the early embryo: is MBT too good to be true?

The question of how early embryonic events are temporally regulated has traditionally been tied to the mid-blastula transition (MBT). This concept has directed the studies in Xenopus and influenced the studies in other organisms. By examining the weaknesses in the concept of MBT, we hope to refocus the study of temporal regulation on the many developmental transitions that do exist and to clear the way for an alternative viewpoint that emphasizes the similarities between developmental processes in different organisms.

Stem Cell Plasticity in Mammals and Transdetermination in Drosophila: Common Themes?

Stem cells have been identified in a number of mammalian tissues (e.g., bone marrow, muscle, gut, skin, and neural tissues). Until recently, it was generally believed that the differentiation potential of a mammalian somatic stem cell is restricted to one tissue only, as in the case of hematopoietic stem cells differentiating into hematopoietic cells. In this sense, somatic stem cells are limited in their differentiation potential. Several lines of evidence now challenge the idea of unilateral development. New reports show mammalian somatic stem cells can, in the course of regeneration, repopulate heterologous cell systems and therefore possess a surprisingly broad spectrum of differentiation potential. Thus, mammalian stem cells are apparently capable of fate changes between stem cell systems, although the mechanisms leading to such changes are unclear.

A Transient Cell Cycle Shift in Drosophila Imaginal Disc Cells Precedes Multipotency

When Drosophila imaginal discs regenerate, specific groups of cells can switch disc identity so that, for example, cells determined for leg identity switch to wing. Such switches in cell determination are known as transdetermination. We have developed a system by which individual cells are marked and monitored in vivo as they transdetermine so that their proliferation, cell sizes, and differentiation are accurately traced. Here, we document that when cells transdetermine, they do not convert to a younger cell cycle. Instead, cell cycle changes precede transdetermination and are different from those observed at any time in normal development. We propose that it is not a younger but a unique cell cycle progression and a big cell size that conditions the cells for developmental plasticity.

Cell cycle changes during growth and differentiation of imaginal leg discs in Drosophila melanogaster.

Abstract The relative DNA content of Drosophila melanogaster imaginal leg disc nuclei during larval growth and pupal and adult differentiation was measured by microspectrophotometry. During the larval proliferative phase there were twice as many nuclei in the 4C class as nuclei in the 2C class. At the end of the third larval instar, the proportion of nuclei with a 4C DNA value increased. By 3 hr after pupariation, during pupal cuticle secretion, 90% of the nuclei were in this class. After pupal apolysis which occurs at 12 hr after pupariation, the 4C to 2C ratio was reversed. The increase in the proportion of nuclei with a 2C value was observed until 24 hr after pupariation when 90% of the nuclei were in this class. We propose that most cells divide at least once between pupal and adult differentiation. All of these changes in the cell cycle were correlated temporally with changes in the ecdysteroid titers that occur during these periods.

Lumenal transmission of decapentaplegic in Drosophila imaginal discs.

Drosophila imaginal discs are sac-like appendage primordia comprising apposed peripodial and columnar cell layers. Cell survival in disc columnar epithelia requires the secreted signal Decapentaplegic (DPP), which also acts as a gradient morphogen during pattern formation. The distribution mechanism by which secreted DPP mediates global cell survival and graded patterning is poorly understood. Here we report detection of DPP in the lumenal cavity between apposed peripodial and columnar cell layers of both wing and eye discs. We show that peripodial cell survival hinges upon DPP signal reception and implicate DPP-dependent viability of the peripodial epithelium in growth of the entire disc. These results are consistent with lumenal transmission of the DPP survival signal during imaginal disc development.

Repression and turnover pattern fushi tarazu RNA in the early Drosophila embryo

Embryonic expression of transcripts from the Drosophila gene fushi tarazu (ftz) progresses through a series of spatial patterns, culminating in a seven-banded pattern at the cellular blastoderm stage. We studied the generation of this pattern using inhibitors of RNA synthesis (alpha-amanitin) and protein synthesis (cycloheximide). Injections of alpha-amanitin revealed that ftz RNA turns over extremely rapidly in the embryo, and we think that this may be essential to effect rapid changes in ftz RNA patterns. Injections of cycloheximide added to the normal domains of ftz expression, creating novel expression patterns that were dependent on the time of injection. These novel patterns suggest that two superimposed systems of repression establish the normal, seven-banded pattern of ftz expression. One system sets up a banded pattern over the entire length of the embryo, and the other restricts actual expression to the middle portion of the embryo.