Convergence of mitochondrial and chloroplastic ANAC017/PAP-dependent retrograde signalling pathways and suppression of programmed cell death

Abstract

The energy-converting organelles mitochondria and chloroplasts are tightly embedded in cellular metabolism and stress response. To appropriately control organelle function, extensive regulatory mechanisms are at play that involve two-way exchange between the nucleus and mitochondria/chloroplasts. In recent years, our understanding of how mitochondria and chloroplasts provide ‘retrograde’ feedback to the nucleus, resulting in targeted transcriptional changes, has greatly increased. Nevertheless, mitochondrial and chloroplast retrograde signalling have largely been studied independently, and only few points of interaction have been found or proposed. Through reassessment of recent publications, this perspective proposes that two of the most well-studied retrograde signalling pathways in plants, those mediated by ANAC017 and those mediated by phosphoadenosine phosphate (PAP), are most likely convergent and can direct overlap** genes. Furthermore, at least part of this common retrograde response appears targeted towards suppression of programmed cell death (PCD) triggered by organellar defects. The identified target genes are discussed in light of their roles in PCD suppression and amplifying the signalling cascade via positive-feedback loops. Finally, a mechanism is proposed that may explain why the convergence of PAP/ANAC017-dependent signalling appears capable of suppressing some types of PCD lesions, but not others, based on the subcellular location of the initial PCD-inducing dysfunction.

Main

The complex endosymbiotic origin of eukaryotic cells and the inherent separation of intracellular compartments necessitates efficient communication between the organelles and the nucleus. The feedback provided by organelles to the nucleus – retrograde signalling – has been described in all eukaryotic kingdoms (animals, fungi and plants), but each kingdom seems to have its proprietary systems.^{1, 2, 3} Plants also evolved intricate retrograde pathways between chloroplasts and the nucleus.⁴ Mitochondria and chloroplasts have a key role during normal life in energy metabolism and biosynthesis of important compounds for the cell. However, they can also have key roles in the execution of programmed cell death (PCD) to remove unwanted cells that may produce toxic levels of reactive molecules or are infected by pathogens.^{5, 6, 7, 8} Therefore, many systems are in place that control organelle numbers, composition and quality, and keep track of their suicidal tendencies.³ This perspective paper proposes that mitochondrial and chloroplast retrograde signalling – or at least some types – are convergent on overlap** target genes. Furthermore, this common response mechanism may help prevent PCD initiation and steer the balance towards cell survival, most likely by suppressing excessive oxidative stress and repairing organelle damage.

The ANAC017 Pathway and PCD

The most clearly understood pathway for mitochondrial retrograde signalling in plants involves activation of the transcription factor ANAC017.^{9, 10, 11, 12} This activation can be triggered by acute inhibition of mitochondrial function, for example, by antimycin A (complex III).¹⁰ Recently it was shown that the ANAC017-dependent signalling pathway is also active when mitochondrial biogenesis is disturbed by genetic defects, for instance when the mitochondrial prohibitin AtPHB3 scaffolding complex is defective, or when mitochondrial/plastid RNA polymerase RpoTmp is impaired.⁹ ANAC017 has a C-terminal transmembrane domain, which probably anchors it into the endoplasmic reticulum (ER).^{10, 11} Currently, we have little understanding of how signals from dysfunctional mitochondria reach the ER and activate ANAC017. Most evidence points towards mitochondrial reactive oxygen species (ROS) production, with H₂O₂ as the most likely mobile signal.^{10, 13} Inhibitor studies suggested that rhomboid proteases might be involved in release of ANAC017 from the ER.¹⁰

Although we know little about the activation of ANAC017, we have a good overview of its downstream target genes, at least 200 in Arabidopsis.⁹ Many of these encode mitochondrial proteins such as alternative oxidase (AOX1a), alternative NADH dehydrogenases, OPA3 potentially involved in mitochondrial fission and a range of oxidative stress inducible genes with less-defined roles.^{14, 15, 16} On the basis of recent findings, it appears that one of the functions of the ANAC017 pathway may be to suppress cell death. First, when ANAC017 function is abolished in mitochondrial RNA polymerase rpotmp anac017 double mutants, the plants develop spontaneous lesions.^{9, 17} This is likely caused at least in part by the lack of ANAC017-dependent induction of AOX1a in rpotmp anac017 double mutants, as rpotmp aox1a double mutants also display similar lesions.¹⁸ Although the exact reasons for lesion formation in rpotmp aox1a or rpotmp anac017 mutants are unknown, they are likely caused by spontaneous PCD. In agreement, AOX has been shown extensively to suppress PCD in plants during inhibition of mitochondrial function.^{Figure 1). It is tempting to speculate that they are in fact directly related or operate in parallel, although a possible mechanism behind this interaction is not clear yet. Many downstream genes appear unique to either ANAC017 or PAP pathways (although the mutants were not compared directly under the same conditions), so their overlap is most likely partial.12 Whether ANAC017/PAP target genes are induced in vivo depends on the circumstances of the cellular dysfunction, most probably the location and type of ROS produced (Figure 2). Evidence to date implicates superoxide and H₂O₂ produced in chloroplasts and mitochondria, but changes in chloroplastic redox poise per seare sufficient to inactivate SAL1 and increase PAP. It is intriguing that the induction of the PAP pathway can rescue some PCD-causing defects, but plants apparently do not always activate this mechanism: for instance, in the mips1 mutants. Perhaps the needed cross-talk between signalling pathways is not present (yet) to activate the PAP pathway in those cases. Alternatively, overactivation of the PAP pathway may overall be more detrimental than beneficial, resulting in negative selection pressure. Finally and critically, the function of mitochondrial-targeted SAL1 remains to be determined.}