A simplified qPCR method revealing tRNAome remodeling upon infection by genotype 3 hepatitis E virus

Abstract

The landscape of tRNA-viral codons regulates viral adaption at the translational level, presumably through adapting to host codon usage or modulating the host tRNAome. We found that the major zoonotic genotype of hepatitis E virus (HEV) has not adapted to host codon usage, prompting exploration of the effects of HEV infection on the host tRNAome. However, tRNAome quantification is largely impeded by the extremely short sequences of tRNAs and redundancy of tRNA genes. Here, we present a length-extension and stepwise simplified qPCR method that utilizes a universal DNA/RNA hybrid tRNA adaptor and degenerate primers. Using this novel methodology, we observe that HEV infection dramatically reprograms the hepatic tRNAome, which is likely to facilitate translation of viral RNAs. This tRNAome quantification method bears broad implications for future tRNA research and possibly tRNA-based diagnostics.

The biological process of lysine‐tRNA charging is therapeutically targetable in liver cancer

“…We have recently developed a novel qPCR method that specifically quantifies mature tRNAs. 6 We think this form of tRNAs is functionally most relevant as these mature tRNAs are directly charged with amino acids and thus facilitate protein synthesis.…”

“…5 From the GtRNA 2.0 database, we have recently retrieved 57 human mature tRNA species constituting as the tRNAome. 6 Dysregulation of gene translation has been well-recognized in cancer development and progression. 7,8 Involvement of tRNAs has been demonstrated in breast cancer, lung cancer and melanoma.…”

“…Based on these unique characteristics, we have developed a simplified qRT-PCR assay to quantify the mature tRNAome. 6 Several enzymes participate in the modification and maturation of tRNAs. ARSs responsible for charging amino acids have evolutionarily conserved enzymatic properties.…”

“…In this study, we aim to profile the tRNAome landscape in tumours of HCC patients using our recently developed mature tRNA quantification assay. 6 Secondly, we aim to assess the therapeutic potential of targeting the interface of amino acid charging to tRNA in experimental models of liver cancer.…”

“…However, the minimally required tRNA species for decoding in real life is usually less than the theoretically calculated number 5 . From the GtRNA 2.0 database, we have recently retrieved 57 human mature tRNA species constituting as the tRNAome 6 …”