A plasmid library of full-length zebrafish rab proteins for in vivo cell biology

Abstract

The zebrafish is an emerging model for highly sophisticated medium-throughput experiments such as genetic and chemical screens. However, studies of entire protein families within this context are often hampered by poor genetic resources such as clone libraries. Here we describe a complete collection of 76 full-length open reading frame clones for the zebrafish rab protein family. While the mouse genome contains 60 rab genes and the human genome 63, we find that 18 zebrafish rab genes have 2, and in the case of rab38, 3 paralogues. In contrast, we were unable to identify zebrafish orthologues of the mammalian Rab2b, Rab17 or Rab29. We make this resource available through the Addgene repository to facilitate cell biologic approaches using this model.

Keywords: in vivo; library; rab; screening; zebrafish.

Figure 1.

Homology relationships between zebrafish, mouse and human rab proteins. A) Orthology between rab proteins of different species is shown with solid lines. Green shading represents paralogous genes within a species which are not duplicated within the other 2 species. Red shading represents lack of an ortholog present in the other 2 species. B) Neighbor joining tree for all zebrafish rab protein sequences. Numbers at each node represent the percentage of 10,000 bootstrap replicates. Nodes with less than 50% of bootstrap replicates are collapsed. Green shading represents paralogous genes which are not duplicated in mouse or human. All paralogues cluster together.

Figure 2.

Expression of canonical zebrafish rabs in dermal cells. A) Expression of FusionRed-Rab1ab on a membrane localized EGFPcaax background marks the endoplasmic reticulum. B) Expression of TagBFP2-Rab4a on a membrane localized EGFPcaax background marks the early/recycling endosome compartment. C) Expression of mVenus-Rab9a on a endoplasmic reticulum localized mKate2-KDEL background marks the late endosome/golgi compartment.