A genome-wide survey and systematic RNAi-based characterization of helicase-like genes in Caenorhabditis elegans

Abstract

Helicase-like proteins play a crucial role in nucleic acid- and chromatin-mediated reactions. In this study, we identified 134 helicase-like proteins in the nematode Caenorhabditis elegans and classified the proteins into 10 known subfamilies and a group of orphan genes on the basis of sequence similarity. We characterized loss-of-function phenotypes in RNA interference (RNAi)-treated animals for helicase family members, using the RNAi feeding method, and found several previously unreported phenotypes. Fifty-one (39.5%) of 129 genes tested showed development- or growth-defect phenotypes, and many of these genes were putative nematode homologs of essential genes in a unicellular eukaryote, budding yeast, suggesting conservation of these essential proteins in both species. Comparative analyses between these species identified evolutionarily diverged nematode proteins as well as conserved family members. Chromosome mapping of the nematode genes revealed 10 pairs of putative duplicated genes and clusters of C. elegans-specific SNF2-like genes and Helitrons. Analyses of transcriptional profile data revealed a predominantly oogenesis- and germline-enriched expression of many helicase-like genes. Finally, we identified the D2005.5(drh-3) gene in an RNAi-based screen for genes involved in resistance to X-ray irradiation. Analysis of DRH-3 will clarify the potentially novel mechanism by which it protects against X-ray-induced damage in C. elegans.

Figure 1

Typical phenotypes of F1 progeny from nematodes RNAi-treated for helicase-like genes. Typical images of the F1 progeny from eggs laid by RNAi-treated P0 animals on RNAi plates for control [vector alone (A)], mcm-6(ZK632.1) RNAi (B), W08D2.7 RNAi (C), ZK686.2 RNAi (D), Y50D7A.11 RNAi (E), and cgh-1 (C07H6.5) RNAi [(F) and (G) in a threefold enlarged image] are shown. The progeny were cultured on RNAi plates supplemented with dsRNA-expressing bacteria for 3 days after laying, and images were then captured. The RNAi phenotypes shown are embryonic lethal (Emb in Table 1) (B), larval arrest (Lva) (C), slow growth (Gro) (D), slow growth and sterile progeny (Gro Stp) (E), and protruding vulva (Pvl) (F and G). Arrows indicate protruded vulva (F) and resultant abdominal burst (G). Bar: 1 mm.

Figure 2

Influence of RNAi treatment of helicase family genes on larval growth. The growth of F1 larvae from eggs laid by RNAi-treated P0 animals was monitored by measuring the body length of progeny. The resultant growth curves of progeny of animals (N2 strain) that were RNAi-treated for the indicated 10 genes (T26G10.1 to Y23H5B.6) in the DEAD-box subfamily are shown together with the growth curve of progeny without RNAi-treatment [control (A)]. The growth curves obtained from RNAi experiments for the genes in other subfamilies are shown with their control growth curve [control (B)] as follows: Y54E10A.9(vbh-1) from the DEAD-box subfamily; C06E1.10(rha-2), F56D2.6, and T05E8.3 from the DEAH-box subfamily; W08D2.7 from the SKI2 subfamily; Y80D3A.2(emb-4) in the UPF1 subfamily; F26F12.7(let-418) and Y111B2A.22(ssl-1) from the SWI2/SNF2 subfamily; Y50D7A.11 from the RAD3 subfamily; and F20H11.2(nsh-1) as an orphan member, respectively. Experiments for C08F8.2 (SKI2 subfamily) and F37A4.8(isw-1) (SWI2/SNF2 subfamily) indicated in bold letters were carried out using the rrf-3 mutant as a host because of weak slow-growth phenotypes of the RNAi-treated N2 animals, and the resultant growth curves of progeny of control (open triangle) and RNAi-treated (closed triangle) animals are shown. The calculated growth rate for each population was 17.5 µm/h [control (A)], 0.7 (T26G10.1), 1.5 (Y71H2AM.19), 3.3 (B0511.6), 3.4 (C24H12.4), 4.0 (ZK512.2), 4.5 (Y94H6A.5), 5.1 (F55F8.2), 5.5 (ZK686.2), 8.4 (H20J04.4), 8.7 (Y23H5B.6), 12.5 [control (B)], 6.8 (Y54E10A.9(vbh-1)), 2.7 (C06E1.10(rha-2)), 3.0 (F56D2.6), 7.4 (T05E8.3), 3.2 (W08D2.7), 9.1 (Y80D3A.2(emb-4)), 2.6 (F26F12.7(let-418)), 9.1 (Y111B2A.22(ssl-1)), 7.6 (Y50D7A.11), 6.0 (F20H11.2(nsh-1)), 8.5 (C08F8.2) and 12.5 (rrf-3 control), and 8.3 (F37A4.8(isw-1)) and 11.8 (rrf-3 control).