Disease relevance of RNH1

• Functional complementation of an Escherichia coli ribonuclease H mutation by a cloned genomic fragment from the trypanosomatid Crithidia fasciculata [1].
• The hallmark of the RNase H/caulimovirus nucleic acid binding motif is a stretch of 40 amino acids with 11 highly conserved residues, seven of which are aromatic [2].
• We have cloned genes encoding RNase H from Escherichia coli rnh mutants, Salmonella typhimurium and Saccharomyces cerevisiae [3].
• RNase H is involved in selection of the origins of replication in E. coli and in DNA synthesis of the positive strand of retroviruses [4].
• Amino acids essential for RNase H activity of hepadnaviruses are also required for efficient elongation of minus-strand viral DNA [5].

High impact information on RNH1

• Interestingly, we find that PRP16 promotes a conformational change in the spliceosome which results in the protection of the 3' splice site against oligo-directed RNase H cleavage [6].
• Differences in the RNase H sensitivity of pre-mRNA found in the two spliceosome forms indicate an increased association of splicing factors with the 5' splice site during spliceosome assembly [7].
• A gene designated Cfa RNH1 has been cloned by complementation of an RNase H deficiency in an Escherichia coli rnhA mutant by using a genomic DNA library from the trypanosomatid Crithidia fasciculata [1].
• RNase H degradation in the presence of oligo(dT) demonstrated that the wild-type and mutant PGK1 mRNAs are deadenylated prior to endonucleolytic cleavage and that the half-life of the poly(A) tail is three- to sixfold lower than that of the remainder of the mRNA [8].
• We have combined oligonucleotide-directed RNase H degradation and immunoprecipitation in a study of the association of the Saccharomyces cerevisiae PRP4 protein with the U4-U6 complex [9].

Chemical compound and disease context of RNH1

• Surprisingly, NEM strongly inhibits the same bacterial RNase H in the presence of a recombinant form of HIV RT devoid of nuclease activity [10].

Biological context of RNH1

• Deletion of the gene (called RNH35) from the yeast genome leads to an about 75% decrease of RNase H activity in preparations from the mutated, still viable cells [11].
• Crystallographic studies of E. coli RNase H indicate that several amino acids, conserved in both cellular and retroviral RNases H, form an active site for hydrolysis of the RNA of RNA-DNA hybrids [4].
• The C-terminal portion of Ty3 RT encodes a functional RNase H domain, although the hydrolysis profile suggests an increased spatial separation between the catalytic centers [12].
• The second 4395-base pair open reading frame, TYB4, encodes a polyprotein that has domains with significant homology to retroviral protease, integrase, reverse transcriptase, and RNase H, structurally arranged in that order [13].
• Reverse transcriptase and RNase H amino acid sequences from elements in the gypsy group--including the recently described SURL elements, TED, Cft1, and Ulysses,--were aligned and analyzed by using parsimony and bootstrapping methods, with plant caulimoviruses and/or retroviruses as outgroups [14].

Anatomical context of RNH1

• Most of the uncleaved pre-mRNA remaining after RNase H challenge was found associated with two forms of the yeast spliceosome [7].
• Ribonuclease H IIb, which seems to play a physiological role during transcription, was purified from calf thymus tissue [15].

Associations of RNH1 with chemical compounds

• Although all mutations had minimal impact on DNA polymerase function, amidation of Asp-358, Glu-401, and Asp-426 eliminated Mg(2+)- and Mn(2+)-dependent RNase H function [16].
• Deletion of HBV reverse transcriptase (RT) or RNase H domains resulted in a dramatic drop in histidine prototrophs [17].
• Replacing His-427 and Tyr-459 with Ala and Asp-469 with Asn resulted in reduced RNase H activity in the presence of Mg(2+), whereas in the presence of Mn(2+) these mutants displayed a lack of turnover [16].
• Two ribonuclease H activities have been found in yeast RNA polymerase A. The nuclease activities comigrated with subunits A49 (Mr = 49,000) and A40 (Mr = 40,000), after electrophoresis in a sodium dodecyl sulfate polyacrylamide gel containing [32P](rG)n . (dC)n as substrate [18].
• Despite conservation of catalytically important residues in the RNase H domain, Fe(2+) fails to replace Mg(2+) in the RNase H catalytic center for localized generation of hydroxyl radicals, again suggesting this domain may be structurally distinct from its retroviral counterparts [12].

Other interactions of RNH1

• Two of these genes have recently been further characterized: ORF YGR255w, renamed RTT102, encodes a regulator of the Ty1-element transposition, whereas ORF YGR276c was found to encode the 70 kDa RNase H activity and was renamed RNH70 (Frank et al., 1999) [19].
• Several lines of evidence suggest that the chromatin RNase H of 49,000 daltons (RNase H49) is the same protein as subunit A49 [18].
• All retroelements use an RNase H-resistant oligoribonucleotide spanning a purine-rich sequence (the polypurine tract or PPT) to prime plus-strand DNA synthesis [20].
• Accumulation of Ty1-integrase and Ty1-reverse transcriptase/ribonuclease H is defective in an hsx1 mutant [21].
• By probing the precatalytic spliceosome formed in temperature-sensitive prp2 mutant extracts with oligonucleotides complementary to snRNAs, we found that the 5' end of U1 was sensitive to RNase H digestion whereas the 5' splice site-interacting region of U6 became resistant [22].

Analytical, diagnostic and therapeutic context of RNH1

• Targeting a cellular RNase H to HIV may help define the site(s) of RNA-DNA hybrids that are susceptible to nonretroviral RNase H and may be useful for gene therapy to inhibit retroviral replication [23].
• The native molecular mass for the enzyme of around 45 kDa, as determined by gel filtration, suggests that calf thymus ribonuclease H IIb is most probably monomeric [15].

References