Non-AUG Translation Initiation Generates Peroxisomal Isoforms of 6-Phosphogluconate Dehydrogenase in Fungi.

Marco Kremp,Johannes Freitag,Alexander Klingenberger,Michael Bölker,Domenica Martorana,Elena Bittner,Thorsten Stehlik

doi:10.3389/fcell.2020.00251

Abstract

Proteins destined for transport to specific organelles usually contain targeting information, which are embedded in their sequence. Many enzymes are required in more than one cellular compartment and different molecular mechanisms are used to achieve dual localization. Here we report a cryptic type 2 peroxisomal targeting signal encoded in the 5′ untranslated region of fungal genes coding for 6-phosphogluconate dehydrogenase (PGD), a key enzyme of the oxidative pentose phosphate pathway. The conservation of the cryptic PTS2 motif suggests a biological function. We observed that translation from a non-AUG start codon generates an N-terminally extended peroxisomal isoform of Ustilago maydis PGD. Non-canonical initiation occurred at the sequence AGG AUU, consisting of two near-cognate start codons in tandem. Taken together, our data reveal non-AUG translation initiation as an additional mechanism to achieve the dual localization of a protein required both in the cytosol and the peroxisomes.

Highlights

Organelles are specialized reaction chambers of eukaryotic cells
We show that a peroxisomal isoform of U. maydis 6-phosphogluconate dehydrogenase (Pgd1) containing a functional PTS2 signal is translated via initiation at a non-AUG start codon
We have previously identified a network of sugar-metabolizing enzymes, which is dually targeted to the cytosol and peroxisomes via post-transcriptional mechanisms

Summary

INTRODUCTION

Organelles are specialized reaction chambers of eukaryotic cells. Each type of organelle harbors a unique combination of enzymes to catalyze specific metabolic pathways such as aerobic ATP generation in the mitochondria or β-oxidation of long-chain fatty acids inside peroxisomes (Lodish et al, 2000). A more recently described mechanism to generate dually targeted protein variants is programmed translational read-through of stop codons. This mechanism is widely used for the formation of C-terminally extended enzymes containing peroxisomal targeting signals (PTS1) (Freitag et al, 2012; Schueren et al, 2014; Stiebler et al, 2014; Schueren and Thoms, 2016). A closer examination of genes for putative 6-phosphogluconate dehydrogenases in Ustilago maydis and related fungi leads to the identification of DNA sequences encoding PTS2like motifs and which are embedded in the 5 UTRs. We show that a peroxisomal isoform of U. maydis 6-phosphogluconate dehydrogenase (Pgd1) containing a functional PTS2 signal is translated via initiation at a non-AUG start codon

RESULTS AND DISCUSSION

MATERIALS AND METHODS

DATA AVAILABILITY STATEMENT