Substrate and enzyme determinants for recognition by human mitochondrial RNase P

TL;DR

This study explores how human mitochondrial RNase P recognizes and processes tRNA precursors, revealing unique substrate and enzyme determinants.

Contribution

The study identifies a rigid measuring mechanism and specific interactions in human mitochondrial RNase P for tRNA processing.

Findings

MtRNase P processes tRNA precursors more efficiently with longer 5′ extensions.

The enzyme uses a rigid measuring mechanism for cleavage-site selection.

TRMT10C–SDR5C1 interactions with the anticodon loop are not crucial for efficient processing.

Abstract

RNase P enzymes of widely varying architectures recognize the 5′-leader/acceptor-stem junction and the D/T loop-interaction region of precursor tRNAs to direct cleavage to the 5′ end of tRNAs. In contrast, human mitochondrial RNase P (mtRNase P) encases the entire tRNA with the aid of the methyltransferase subcomplex TRMT10C–SDR5C1. Here, we performed a kinetic analysis of substrate recognition by mtRNase P using substrate and protein variants. Surprisingly, processing by mtRNase P was found to be more efficient for tRNA precursors with longer 5′ extensions and decreased sharply at a leader length of 1 nt. MtRNase P also employs a more rigid “measuring mechanism” for cleavage-site selection than the related single-subunit enzymes, so that even substrates with a G:C base-pair extension of the acceptor stem are cleaved predominantly at the canonical site. The specific contacts of TRMT10C–SDR5C1 with the anticodon loop are not crucial for efficient processing, but without interactions with the pre-tRNA, TRMT10C–SDR5C1 is unable to stimulate cleavage by the nuclease subunit PRORP, also explaining why mtRNase P reaches its limits with the D-armless mitochondrial tRNASer(AGY). Our findings set human mtRNase P apart in terms of substrate recognition from all other known forms of RNase P, including the related single-polypeptide PRORPs.

Graphical Abstract

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