Fine scale analysis of gene expression in Drosophila melanogaster gonads reveals Programmed cell death 4 promotes the differentiation of female germline stem cells.

BACKGROUND: Germline stem cells (GSCs) are present in the gonads of Drosophila females and males, and their proper maintenance, as well as their correct differentiation, is essential for fertility and fecundity. The molecular characterization of factors involved in maintenance and differentiation is a major goal both in Drosophila and stem cell research. While genetic studies have identified many of these key factors, the use of genome-wide expression studies holds the potential to greatly increase our knowledge of these pathways. RESULTS: Here we report a genome-wide expression study that uses laser cutting microdissection to isolate germline stem cells, somatic niche cells, and early differentiating germ cells from female and male gonads. Analysis of this data, in association with two previously published genome-wide GSC data sets, revealed sets of candidate genes as putatively expressed in specific cell populations. Investigation of one of these genes, CG10990 the Drosophila ortholog of mammalian Programmed cell death 4 (Pdcd4), reveals expression in female and male germline stem cells and early differentiating daughter cells. Functional analysis demonstrates that while it is not essential for oogenesis or spermatogenesis, it does function to promote the differentiation of GSCs in females. Furthermore, in females, Pdcd4 genetically interacts with the key differentiation gene bag of marbles (bam) and the stem cell renewal factor eIF4A, suggesting a possible pathway for its function in differentiation. CONCLUSIONS: We propose that Pdcd4 promotes the differentiation of GSC daughter cells by relieving the eIF4A-mediated inhibition of Bam.

Developmental decoupling of alternative phenotypes: insights from the transcriptomes of horn-polyphenic beetles.

Developmental mechanisms play an important role in determining the costs, limits, and evolutionary consequences of phenotypic plasticity. One issue central to these claims is the hypothesis of developmental decoupling, where alternate morphs result from evolutionarily independent developmental pathways. We address this assumption through a microarray study that tests whether differences in gene expression between alternate morphs are as divergent as those between sexes, a classic example of developmental decoupling. We then examine whether genes with morph-biased expression are less conserved than genes with shared expression between morphs, as predicted if developmental decoupling relaxes pleiotropic constraints on divergence. We focus on the developing horns and brains of two species of horned beetles with impressive sexual- and morph-dimorphism in the expression of horns and fighting behavior. We find that patterns of gene expression were as divergent between morphs as they were between sexes. However, overall patterns of gene expression were also highly correlated across morphs and sexes. Morph-biased genes were more evolutionarily divergent, suggesting a role of relaxed pleiotropic constraints or relaxed selection. Together these results suggest that alternate morphs are to some extent developmentally decoupled, and that this decoupling has significant evolutionary consequences. However, alternative morphs may not be as developmentally decoupled as sometimes assumed and such hypotheses of development should be revisited and refined.

Data pushing: a fly-centric guide to bioinformatics tools.

Bioinformatics tools can be invaluable resources to Drosophila researchers; however, the sheer number of applications and databases can be overwhelming. We present a broad overview of common bioinformatics tasks and the resources used to do them, with a specific focus on resources for Drosophila. The topics covered include: Genome Databases, Sequence Analysis, Comparative Genomics, Gene Expression Databases and Analysis Tools, Function-Based Data and Analysis, Pathways, Networks, and Interactions; and finally, tools to stay current with resources and literature. We also present a compilation of URLs and short descriptions that correspond to the topics and resources mentioned in this review.

Troubleshooting microarray hybridizations.

Microarray experiments are being performed more widely than ever before, but even seasoned investigators can experience technical problems with hybridizations. This chapter provides guidelines for recognizing, rectifying, and avoiding common trouble areas. Specifically, it addresses frequent complications related to artifacts of printing, RNA sample preparation and quality, fluorophore labeling, hybridization conditions, and posthybridization washes. Emphasis is placed on investigating problems though a combination of appropriate controls and image analysis, where diagnostic plots of data quality are used to illustrate characteristics of acceptable and unsatisfactory hybridizations. This chapter also discusses resources available to microarray users hoping to improve the sensitivity and specificity of their experiments.