Development Genes and Evolution

, Volume 217, Issue 3, pp 189–196 | Cite as

RNAi-induced phenotypes suggest a novel role for a chemosensory protein CSP5 in the development of embryonic integument in the honeybee (Apis mellifera)

Original Article

Abstract

Small chemosensory proteins (CSPs) belong to a conserved, but poorly understood, protein family found in insects and other arthropods. They exhibit both broad and restricted expression patterns during development. In this paper, we used a combination of genome annotation, transcriptional profiling and RNA interference to unravel the functional significance of a honeybee gene (csp5) belonging to the CSP family. We show that csp5 expression resembles the maternal-zygotic pattern that is characterized by the initiation of transcription in the ovary and the replacement of maternal mRNA with embryonic mRNA. Blocking the embryonic expression of csp5 with double-stranded RNA causes abnormalities in all body parts where csp5 is highly expressed. The treated embryos show a “diffuse”, often grotesque morphology, and the head skeleton appears to be severely affected. They are ‘unable-to-hatch’ and cannot progress to the larval stages. Our findings reveal a novel, essential role for this gene family and suggest that csp5 (unable-to-hatch) is an ectodermal gene involved in embryonic integument formation. Our study confirms the utility of an RNAi approach to functional characterization of novel developmental genes uncovered by the honeybee genome project and provides a starting point for further studies on embryonic integument formation in this insect.

Keywords

Insect cuticle Embryonic development Gene silencing CSP family 

Supplementary material

427_2006_127_MOESM1_ESM.gif (13 kb)
Fig. S1RT-PCR amplification of uth and Dnmt in 48-h-old honeybee embryos injected at 10 ± 2 h with dsRNA (+) and in control eggs (−) injected with DNA. The primers for Dnmt are described in Wang et al. (2006). Computer-generated images (MD Phosphor-Imager 400S) of agarose gels were analysed using ImageQuant software (Kucharski and Maleszka 2002a, b). The levels of both genes in dsRNA-treated embryos are shown relatively to controls (100%) (JPG 593 kb).
427_2006_127_MOESM2_ESM.gif (109 kb)
Fig. S2In situ hybridization showing the expression of csp5/uth in 44- to 48-old embryos: a Longitudinal sections plus DAPI staining, b transverse section through thoracic region showing the form and relation of the ectoderm (E) and mesoderm (M). Mesoderm is composed of two layers continuous with one another at the lateral margins (Nelson 1915) (JPG 805 kb).
427_2006_127_MOESM3_ESM.jpg (27 kb)
Fig. S3In situ hybridization of uth showing strong labelling in the cerebral lobes of 66- to 70-h-old embryo. See Fig. 3 for more details (JPG 1.16 kb).
427_2006_127_MOESM4_ESM.gif (140 kb)
Fig. S4aBlack arrows show the cuticular structures maintaining the shape of the control embryo. bd dsRNA-injected embryos lack the cuticular scaffolding. In all embryos, the chorion is indicated by white arrows (JPG 6.20 kb).

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Copyright information

© Springer-Verlag 2007

Authors and Affiliations

  1. 1.ARC Centre for the Molecular Genetics of Development and Visual Sciences, Research School of Biological SciencesThe Australian National UniversityCanberraAustralia

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