Is there a signal transduction pathway that links events at the plasma membrane to the phosphorylation state of the mitochondrial pyruvate dehydrogenase complex?

Monoclonal antibodies against the E1α subunit of pyruvate dehydrogenase (PDH) were used to quantify the mitochondrial pyruvate dehydrogenase complex (mtPDC) by enzyme‐linked immunosorbent assay (ELISA). Recombinant Arabidopsis thaliana PDH (E1) was used to calibrate the ELISA. Antibodies against a synthetic phosphopeptide corresponding to phosphorylation site one of E1α were used in an ELISA to quantify phospho‐PDC (P‐PDC). For calibration of the second ELISA, recombinant E1 was phosphorylated in vitro with recombinant A. thaliana E1‐kinase. The two ELISA were used to quantify mitochondrial total‐ and P‐PDC in clarified homogenates from Nicotiana tabacum BY‐2 suspension cells. The level of mtPDC remained constant throughout the 7‐day growth cycle at 25.1 g −1  FW. During the lag (days 0–2) and stationary (day 7) stages of the growth cycle, the mtPDC was completely phosphorylated (inactive), whereas during the log‐growth stage it was completely dephosphorylated (active). Exposure of 3‐ or 7‐day posttransfer suspension cells to osmotic stress significantly decreased proportion of P‐PDC. A series of pharmacological studies were undertaken to gain insight into the signal transduction pathways coupling osmotic stress perception with control of mitochondrial respiration. Results from these studies indicate a signal transduction pathway linking stress perception to control of mitochondrial respiration that includes protein kinases and phosphoprotein phosphatases.