GSE130155
Effect of adat2 silencing in Neurospora crassa on translation kinetics and transcriptome
The silencing of tRNA-specific adenosine deaminase adat2 in Neurospora crassa abolished most of the I34 modification, resulting in major tRNA profile changes. The adat2 silencing caused genome-wide codon usage-biased ribosome pausing on mRNAs corresponding to changes in the tRNA profile, which reprograms translation elongation kinetics on ADAT-related codons. The ribosome profiling in this study was used to measure the effect of defective I34 modification of the eight tRNAs on translation kinetics during elongation stage. The accompanying mRNA-seq was used to normalize the ribosome profiling data using the same library construction protocol (reference to ARTseq™ Ribosome Profiling Kit (Catalog Number: RPYSC12116)) under the same growth condition. The independent mRNA-seq was used to measure the effect of I34 abolishment on transcriptome change using standard Illumina protocols. Meanwhile, to isolate the role of codon usage in translation elongation and protein level changes when adat2 is silenced, the independent mRNA-seq was also used to exclude the effect of mRNA level changes. Overall design: The transcription profile and ribosome protected fragment (RPF) profiles of wild-type strain and adat2-silenced mutant of Neurospora crassa were generated by deep sequencing using Illumina sequencing platform.