Stage‐specific expression of Caenorhabditis elegans ribonuclease H1 enzymes with different substrate specificities and bivalent cation requirements

Abstract

An enzyme that specifically degrades the RNA strand of RNA-DNA hybrids was first characterized from extracts of calf thymus tissue [1] and was named ribonuclease H (RNase H; EC 3.1.26.4) [2]. This protein has also been identified in viruses [3], bacteria [4,5], and archaea [6,7], indicating the essential nature of its roles in cellular metabolism. Prokaryotic RNase H enzymes are divided by sequence similarity into three groups: HI, HII, and HIII. On the other hand, there are two types of eukaryotic RNase H: RNase H1 is homologous to prokaryotic RNase HI, and RNase H2 is homologous to prokaryotic RNase HII. There is a distinct difference between the two types of eukaryotic RNase H in terms of the bivalent metal ion cofactor. Ribonuclease H1 (RNase H1) is a widespread enzyme found in a range of organisms from viruses to humans. It is capable of degrading the RNA moiety of DNA-RNA hybrids and requires a bivalent ion for activity. In contrast with most eukaryotes, which have one gene encoding RNase H1, the activity of which depends on Mg 2+ ions, Caenorhabditis elegans has four RNase H1-related genes, and one of them has an isoform produced by alternative splicing. However, little is known about the enzymatic features of the proteins encoded by these genes. To determine the differences between these enzymes, we compared the expression patterns of each RNase H1-related gene throughout the development of the nematode and the RNase H activities of their recombinant proteins. We found gene-specific expression patterns and different enzymatic features. In particular, besides the enzyme that displays the highest activity in the presence of Mg 2+ ions, C. elegans has another enzyme that shows preference for Mn 2+ ion as a cofactor. We characterized this Mn 2+ -dependent RNase H1 for the first time in eukaryotes. These results suggest that there are at least two types of RNase H1 in C. elegans depending on the developmental stage of the organism.Abbreviations dsRHbd, double-stranded RNA and RNA-DNA hybrid-binding domain; Ec-RNHI, Escherichia coli ribonuclease HI; Pf-RNHII, Pyrococcus furiosus ribonuclease HII; PTC, premature termination codon, RNase H, ribonuclease H.

Evolution of ribonuclease H genes in prokaryotes to avoid inheritance of redundant genes

“…Interestingly, human genome contains one RNase H1 with dsRHbd and at least three pseudogenes related to RNase H1 with dsRHbd [40]. Although we previously showed that four RNase H1-encoding genes in Caenorhabditis elegans exhibited gene-specific expression patterns during development; one gene encodes RNase H1 with dsRHbd and other three gene encode RNase H1 without dsRHbd [41], it was also found that most of the eukaryotic genomes contained single-copy genes encoding RNase H1 with dsRHbd but RNase H1 without dsRHbd had rarely been identified in the same genome (data not shown). Functional characteristic of RNase H1 with dsRHbd seems to depend on eukaryotic species because disruptions of RNase H1 with dsRHbd resulted in lethal phenotype of fly [42] and mice [13] but showed normal growth in yeast [43] and trypanosoma [44].…”

Ribonuclease H: the enzymes in eukaryotes

“…RNase H1.1 has two potential start codons, the first will initiate a protein with a MTS. Interestingly, the abundance of the mRNA encoding RNase H1.1 increases dramatically at the same stage of development when the amount of mtDNA increases fivefold over the level present initially in the embryo [28]. Perhaps RNase H1.1 plays a role in C. elegans mtDNA replication.…”

An Upstream Open Reading Frame and the Context of the Two AUG Codons Affect the Abundance of Mitochondrial and Nuclear RNase H1

“…Most eukaryotic organisms, including S. cerevisiae, have single genes encoding RNase H1, whereas three related RNase H1 proteins are present in Caenorhabditis elegans (3,18). RNase H1.0, a homolog of mouse RNase H1, does not have a canonical N-terminal MTS (3,18), but another gene encoding a unique form of RNase H1 (RNase H1.1) has no HBD but does have an MTS (5,18). Interestingly, regardless of the presence or absence of the extra N-terminal sequences, an AUG codon is present immediately prior to the HBD.…”