An autophagy-associated Atg8 protein is involved in the responses of Arabidopsis seedlings to hormonal controls and abiotic stresses

Abstract

Eukaryotes contain a ubiquitous family of autophagy-associated Atg8 proteins. In animal cells, these proteins have multiple functions associated with growth, cancer, and degenerative diseases, but their functions in plants are still largely unknown. To search for novel functions of Atg8 in plants, the present report tested the effect of expression of a recombinant AtAtg8 protein, fused at its N-terminus to green fluorescent protein (GFP) and at its C-terminus to the haemagglutinin epitope tag, on the response of Arabidopsis thaliana plants to the hormones cytokinin and auxin as well as to salt and osmotic stresses. Expression of this AtAtg8 fusion protein modulates the effect of cytokinin on root architecture. Moreover, expression of this fusion protein also reduces shoot anthocyanin accumulation in response to cytokinin feeding to the roots, implying the participation of AtAtg8 in cytokinin-regulated root–shoot communication. External application of cytokinin leads to the formation of novel GFP–AtAtg8-containing structures in cells located in the vicinity of the root vascular system, which are clearly distinct in size and dynamic movement from the GFP–AtAtg8-containing autophagosome-resembling structures that were observed in root epidermis cells. Expression of the AtAtg8 fusion construct also renders the plants more sensitive to a mild salt stress and to a lesser extent to a mild osmotic stress. This sensitivity is also associated with various changes in the root architecture, which are morphologically distinct from those observed in response to cytokinin. The results imply multiple functions for AtAtg8 in different root tissues that may also be regulated by different mechanisms.

Identification and functional analysis of Joka2, a tobacco member of the family of selective autophagy cargo receptors

“…[33][34][35] Similarly to the yeast and metazoan systems the plant ATG8 proteins are critical components of the autophagy pathway, therefore in many studies the GFP-ATG8 fusions have been used as markers of autophagosomes in plants. [36][37][38][39][40][41] The lack of obvious phenotypes of atg mutants grown under nutrient-sufficient conditions suggested that autophagy was not essential for plants. However, more detailed studies reveal that the atg mutants senescent earlier and were hypersensitive to nitrogen starvation and carbon limitation.…”

“…The process is controlled by pathways that interpret the status of cellular energy (AMP-dependent protein kinase) and nutrients (target of rapamycin), and by growth factors such as insulin. 9 The previously considered candidates for regulators of ATG genes in plants include hormones such as cytokinins 37 and ABA. 68 The physiological role of autophagy in plants grown under starvation has been studied before and it has been reported that autophagy-defective mutants are impaired in autophagosome formation and cannot survive certain starvation conditions.…”

Ultrastructure of autophagy in plant cells

“…In animals, ATG8 is involved in autophagosome formation. In plants the fluorescence of GFP-AtATG8 family proteins has been assigned to what were correctly called "autophagic bodies" 16 or less carefully "autophagosome-resembling structures," 15,17 or even erroneously "autophagosomes." [18][19][20] Implication of the presence of autophagosomes is incorrect as we do not yet know the various functions of the plant ATG8 proteins.…”

Role of plant autophagy in stress response

“…The ability of autophagy to scavenge oxidized proteins and regulate the ROS levels suggests that autophagy may also be involved in salt and drought stresses. Some ATG genes, such as AtATG8 in Arabidopsis and OsATG10b in rice, reportedly function in response to salt stress and osmotic stress (Slavikova et al, 2008). The more direct and sufficient proof comes from the research by Bassham et al who found that high salt and osmotic stresses activate autophagy, concomitant with the upregulation of AtATG18a expression.…”