A key piece emerges in the noncanonical gibberellin signaling puzzle: PLANT UBX DOMAIN-CONTAINING PROTEIN1

Gibberelins (GAs) function in a myriad of plant developmental processes, such as germination, tissue elongation, and the transition from vegetative growth to reproduction. The action of GA relies on transcriptional regulation through DELLA proteins, which interact with and modify the activity of several transcription factors. In the presence of GA, the receptor GA-INSENSITIVE DWARF1 (GID1) interacts with DELLA, which allows the recognition of DELLA by the F-box protein SLEEPY1 (SLY1), part of an ubiquitin E3 ligase, targeting DELLA for destruction (Blázquez et al., 2020).

Although the GA–GID1–DELLA link represents the canonical GA core signaling, some observations suggest the involvement of additional pathways within GA responses. sly1 knockout lines, which cannot degrade DELLA, display the typical phenotype of GA insensitivity, such as increased seed dormancy, dwarfism, and delayed flowering (Steber et al., 1998; McGinnis et al., 2003). Overexpression of GID1 can partially rescue sly1 phenotypes (Ariizumi et al., 2008; Ariizumi et al., 2013) without inducing DELLA degradation, suggesting that some aspects of GA signaling can still occur independently of DELLA destruction, possibly through the interaction of GID1 with other proteins (Ariizumi et al., 2013).

In this issue of The Plant Physiology, Hauvermale et al. (2022) found that the PLANT UBX DOMAIN-CONTAINING PROTEIN 1 (PUX1), which has been previously implicated in growth regulation (Rancour et al., 2004), functions in GA signaling. First, the authors performed a screen for GID1 interactors, and PUX1 emerged as one of the strongest candidates. PUX1 was able to bind to the three GID1 homologs in Arabidopsis (Arabidopsis thaliana), and the interaction required the UBX domain of PUX1 and was independent of GA.

UBX proteins associate with CELL DIVISION CYCLE 48 (CDC48), a segregase that unfolds and extracts polyubiquitinated proteins from membranes and complexes, acting as a major component of the ubiquitin-dependent protein degradation pathway (Bègue et al., 2019). CDC48 acts in a hexameric complex, and binding of PUX1 induces disassembly of the complex, thus modulating CDC48 functions (Rancour et al., 2004).

To investigate if the ability of PUX1 to associate with CDC48 could affect GID1 functions, the authors performed a velocity sedimentation analysis with extracts of suspension-cultured cells. Velocity sedimentation measures the movement of molecules through a solution (in this case a 15%–40% glycerol gradient) under a centrifugal force. Bigger and more voluminous particles sediment faster and are assigned a higher Sverberg (S) value. After centrifugation, fractions of the solution were analyzed through immunoblot to determine the presence of the different proteins. CDC48 was found in fractions of around 17S, which corresponds with its conformation as part of a hexamer. In extracts from cells overexpressing PUX1, CDC48 also appeared in fractions related to proteins of about 5–8S, which supports a role of PUX1 in promoting the disassembly of CDC48 complexes. In the extracts from control cells, GID1 co-fractionated with PUX1 but not with hexameric CDC48, whereas in the extracts of PUX1 overexpressors, a shift in GID1 migration was detected, co-fractioning with PUX1 and CDC48 subunits at 5–8S. Altogether, these results suggest the possibility of a GID1–PUX1–CDC48 complex acting in vivo.

Finally, to assess the physiological role of PUX1 in GA responses, the authors characterized the phenotype of pux1 knockout lines. In normal conditions, pux1 plants had longer stems and roots, as previously reported in Rancour et al. (2004), and flowered earlier than the wild-type (Figure 1), a phenotype consistent with increased GA signaling. Furthermore, the pux1 plants were less sensitive to the GA inhibitor, paclobutrazol (PAC), as the PAC effects on repressing germination and root elongation were less severe in the pux1 lines than in the wild-type. Altogether, these results suggest that PUX1 acts as a negative regulator of GA signaling.

Figure 1.

pux1 knockout lines display an early flowering phenotype. A, Both pux1-1 and pux1-2 begin to bolt 20-day after germination (DAG), ∼8–12 days earlier than the Wassilewskija (Ws) wild-type. B, Comparison between Ws and the pux1 plants at 32 DAG. Adapted from Hauvermale et al. (2022).

In summary, the work of Hauvermale et al. pinpoints PUX1 as a molecular player in GA signaling, possibly through its ability to interact with the GA receptor GID1 and CDC48, a key member of the protein degradation pathway. Future research may focus on the mechanism downstream of the GID1–PUX–CDC48 interaction and the possible role of PUX1 or CDC48 in regulating GID1 activity or stability.

Conflict of interest statement. None declared.