Biochemical and molecular characterization of two cytidine deaminases in the nematode Caenorhabditis elegans

Abstract

Two cytidine deaminases (CDDs) from the free-living nematode Caenorhabditis elegans have been cloned and characterized. Both Ce-CDD-1 and Ce-CDD-2 are authentic deaminases and both exhibit RNA-binding activity towards AU-rich templates. In order to study their temporal and spatial expression patterns in the worm, reporter gene constructs were made using approx. 2kb of upstream sequence. Transfection of C. elegans revealed that both genes localized to the cells of the intestine, although their temporal expression patterns were different. Expression of Ce-cdd-1 peaked in the early larval stages, whereas Ce-cdd-2 was expressed in all life cycle stages examined. RNA-interference (RNAi) assays were performed for both genes, either alone or in combination, but only cdd-2 RNAi produced a consistent visible phenotype. A proportion of eggs laid from these worms were swollen and distorted in shape.

Nucleotide levels regulate germline proliferation through modulating GLP-1/Notch signaling inC. elegans

“…We found that dietary supplementation of uridine, thymidine, or cytidine, but not other nucleosides, fully restored the fertility of cdd-1/2 DKO worms ( Table 2). The rescue effect of uridine (product of CDD enzymes) or thymidine (derivative of uridine) (Lee et al 2009) is consistent with the previous finding that both CDD-1 and CDD-2 are authentic CDDs (Thompson et al 2002).…”

“…Our conclusion that CDD-1 and CDD-2 act on free nucleotides for the germline proliferation function is based on the following data: (1) A previous study determined that CDD-1/2 deaminate free nucleosides (Thompson et al 2002). (2) Our HPLC-UV measurements indicated a very low U/T:C ratio in the cdd-1/2 DKO worms (Fig.…”

“…cdd-1 and cdd-2 act redundantly to support fertility in C. elegans fed OP50 bacteria cdd-1 and cdd-2, two of the nine putative CDD genes in the C. elegans genome, have been shown to have CDD function, yet RNAi knockdown of each alone displayed only mild phenotypes (Thompson et al 2002;Wang et al 2004). We obtained loss-of-function (lf) deletion alleles of each gene, cdd-1(ok390) and cdd-2(tm742) [cdd-1(−) and cdd-2(−) hereafter] for this study.…”

Tissue-specific modification of gld-2 mRNA in C. elegans: Likely C-to-U editing

“…These proteins appear to be positioned evolutionarily between the cytidine and adenosine deaminase families, and deaminate an adenosine in the anticodon of certain yeast tRNAs (Gerber and Keller 1999). RNAi against each of the nine genes depicted in Figure 3 yielded no mutant phenotypes (Gönczy et al 2000;Kamath et al 2003), suggesting functional redundancy; embryonic defects may be associated with cdd-2(RNAi), however (Thompson et al 2002). Identification of the cytidine deaminase that edits gld-2 mRNA is now essential, but may be complicated by redundancy.…”

“…3), using the Superfamily program (Gough et al 2001). Two of the putative deaminases, CDD-1 and CDD-2, possess RNAbinding activity, and deaminate deoxycytidine in vitro (Thompson et al 2002). Another two candidate proteins, JC8.4 and Y47D3A.14, appear to be orthologs of budding FIGURE 1.…”