Double-stranded RNA interference in the spider Cupiennius salei : the role of Distal-less is evolutionarily conserved in arthropod appendage formation

Abstract

Chelicerates represent a basal arthropod group, which makes them an excellent system for the study of evolutionary processes in arthropods. To enable functional studies in chelicerates, we developed a double-stranded RNA-interference (RNAi) protocol for spiders while studying the function of the Distal-less gene. We isolated the Distal-less gene from the spider Cupiennius salei. Cs-Dll gene expression is first seen in cells of the prosomal segments before the outgrowth of the appendages. After the appendages have formed, Cs-Dll is expressed in the distal portion of the prosomal appendages, and in addition, in the labrum, in two pairs of opisthosmal (abdominal) limb buds, in the head region, and at the posterior-most end of the spider embryo. In embryos, in which Dll was silenced by RNAi, the distal part of the prosomal appendages was missing and the labrum was completely absent. Thus, Dll also plays a crucial role in labrum formation. However, the complete lack of labrum in RNAi embryos may point to a different nature of the labrum from the segmental appendages. Our data show that the expression of Dll in the appendages is conserved among arthropods, and furthermore that the role of Dll is evolutionarily conserved in the formation of segmental appendages in arthropods.

Knockdown of Parhyale Ultrabithorax recapitulates evolutionary changes in crustacean appendage morphology

“…We designed a chemically modified siRNA oligo (Stealth siRNAs, Invitrogen) targeting a nonconserved region of PhDll-e. Injection of that siRNA at the 1-cell stage produced truncated appendages in hatchlings (11 of 80 animals; Fig. 2C, E, and F) consistent with the Dll knockdown phenotype observed in diverse arthropods (8)(9)(10). Control injections of siRNAs designed to target DsRed gave no morphological abnormalities (n ϭ 64).…”

Appendage patterning in the South American bird spider Acanthoscurria geniculata (Araneae: Mygalomorphae)

“…and the function of the Dll gene (e.g. Beermann et al 2001;Schoppmeier and Damen 2001;Khila and Grbic 2007), argue for the homology of the proximal part of the legsthe coxa plus the gnathendite-in all arthropods. Fossil data, however, suggest that the chelicerate coxa is homologous to the crustacean basis; the crustacean coxa is thought to be derived from a proximal endite in an ancestral appendage type (Waloßek 1995).…”

Fossil evidence for the origin of spider spinnerets, and a proposed arachnid order

“…In Limulus these appendages consist of a segmented median branch and a lateral branch with a plate covering lamellate gills. In spider embryos, distalless gene expression shows 4 pairs of spinnerets (anterior and posterior median and lateral pairs) represented by 2 pairs of appendage buds on opisthosomal somites 4 and 5 (15). The appendage buds each later divide in 2 to produce potentially 4 pairs of spinnerets, although in nearly all spiders some of these buds do not develop into functional postembryonic spinnerets.…”