RNA recognition by the E2 subunit of the chloroplast pyruvate dehydrogenase complex from Chlamydomonas

Abstract

The dihydrolipoamide acetyltransferase subunit (DLA2) of the chloroplast puruvate dehydrogenase complex (cpPDC) from the green alga Chlamydomonas reinhardtii has previously been shown to possess a moonlighting activity in chloroplast gene expression. Exclusively under mixotrophic growth conditions, DLA2 forms part of an RNP particle with the psbA mRNA that encodes the D1 protein of the photosystem II reaction center. Here, we report on the further characterization of DLA2´s RNA-binding activity. Size-exclusion chromatography and Western analyses revealed that DLA2 is the only cpPDC subunit that shuttles between the metabolic cpPDC and the RNP complex. Microscale thermophoresis-based determination of RNA-binding affinities demonstrated that two DLA2 regions are crucial for RNA recognition, the peripheral E3-binding domain (E3BD) and the C-terminus of the catalytic domain. Specificity for the psbA RNA probe is conferred by the E3BD in vitro, as verified by competitive binding assays in the presence of an excess of the E3 (DLD2) of cpPDC. The data support a model in which an environmental trigger induces release of DLA2 from the cpPDC and its subsequent association with the psbA mRNA.

Upregulation of chloroplastic pyruvate dehydrogenase genes in rice leaf would potentially drive the in planta photorespiratory bypass for higher biomass

“…The PDC constitutes three components, E1 (pyruvate dehydrogenase in heterotetramer state-a2b2), E2 (dihydrolipoyl acetyltransferase, homodimer) and E3 (dihydrolipoyl dehydrogenase, monomer) with copy numbers of these components in the complex is variable 13,14 . Recent study highlights the E2 component's RNA binding activity with psbA mRNA coding for the D1 protein of the PSII reaction center 15 . To understand the expression pattern for genes of photorespiration and the C 4 pathway in leaf and non-leaf (photosynthetic) tissues in rice, transcriptome analysis was performed in the three rice genotypes, with two biological replicates.…”