TY - JOUR

T1 - Pyrroline-5-carboxylate metabolism protein complex detected in Arabidopsis thaliana leaf mitochondria

AU - Zheng, Yao

AU - Cabassa-Hourton, Cécile

AU - Eubel, Holger

AU - Chevreux, Guillaume

AU - Lignieres, Laurent

AU - Crilat, Emilie

AU - Braun, Hans-Peter

AU - Lebreton, Sandrine

AU - Savouré, Arnould

N1 - Funding Information: This work was supported by the Hubert Curien PROCOPE program between Germany and France (grant no. 46711VJ) funded by the Ministry of Foreign Affairs and by Sorbonne University. This work was also supported by a grant from the China Scholarship Council to YZ. We thank Région Ile de France for financial support of the ProteoSeine@IJM platform.

PY - 2024/2/2

Y1 - 2024/2/2

N2 - Proline dehydrogenase (ProDH) and pyrroline-5-carboxylate (P5C) dehydrogenase (P5CDH) catalyze the oxidation of proline into glutamate via the intermediates P5C and glutamate-semialdehyde (GSA), which spontaneously interconvert. P5C and GSA are also intermediates in the production of glutamate from ornithine and α-ketoglutarate catalyzed by ornithine δ-aminotransferase (OAT). ProDH and P5CDH form a fused bifunctional PutA enzyme in Gram-negative bacteria and are associated in a bifunctional substrate channelling complex in Thermus thermophilus, but the physical proximity of ProDH and P5CDH in eukaryotes has not been described. Here we report evidence of physical proximity and interactions between Arabidopsis ProDH, P5CDH and OAT in the mitochondria of plants during dark-induced leaf senescence when all three enzymes are expressed. Pairwise interactions and localization of the three enzymes were investigated using bimolecular fluorescence complementation (BiFC) with confocal microscopy in tobacco and sub-mitochondrial fractionation in Arabidopsis. Evidence for a complex composed of ProDH, P5CDH, and OAT was revealed by co-migration of the proteins in native conditions upon gel electrophoresis. Co-immunoprecipitation coupled with mass spectrometry analysis confirmed the presence of the P5C metabolism complex in Arabidopsis. Pull-down assays further demonstrated a direct interaction between ProDH1 and P5CDH. P5C metabolism complexes may channel P5C among the constituent enzymes and directly provide electrons to the respiratory electron chain via ProDH.

AB - Proline dehydrogenase (ProDH) and pyrroline-5-carboxylate (P5C) dehydrogenase (P5CDH) catalyze the oxidation of proline into glutamate via the intermediates P5C and glutamate-semialdehyde (GSA), which spontaneously interconvert. P5C and GSA are also intermediates in the production of glutamate from ornithine and α-ketoglutarate catalyzed by ornithine δ-aminotransferase (OAT). ProDH and P5CDH form a fused bifunctional PutA enzyme in Gram-negative bacteria and are associated in a bifunctional substrate channelling complex in Thermus thermophilus, but the physical proximity of ProDH and P5CDH in eukaryotes has not been described. Here we report evidence of physical proximity and interactions between Arabidopsis ProDH, P5CDH and OAT in the mitochondria of plants during dark-induced leaf senescence when all three enzymes are expressed. Pairwise interactions and localization of the three enzymes were investigated using bimolecular fluorescence complementation (BiFC) with confocal microscopy in tobacco and sub-mitochondrial fractionation in Arabidopsis. Evidence for a complex composed of ProDH, P5CDH, and OAT was revealed by co-migration of the proteins in native conditions upon gel electrophoresis. Co-immunoprecipitation coupled with mass spectrometry analysis confirmed the presence of the P5C metabolism complex in Arabidopsis. Pull-down assays further demonstrated a direct interaction between ProDH1 and P5CDH. P5C metabolism complexes may channel P5C among the constituent enzymes and directly provide electrons to the respiratory electron chain via ProDH.

KW - Arabidopsis thaliana

KW - mitochondria

KW - ornithine δ-aminotransferase

KW - proline catabolism

KW - proline dehydrogenase

KW - protein complex

KW - pyrroline-5-carboxylate dehydrogenase

UR - http://www.scopus.com/inward/record.url?scp=85184486048&partnerID=8YFLogxK

U2 - 10.1093/jxb/erad406

DO - 10.1093/jxb/erad406

M3 - Article

C2 - 37843921

VL - 75

SP - 917

EP - 934

JO - Journal of experimental botany

JF - Journal of experimental botany

SN - 0022-0957

IS - 3

ER -