The ABCB family belongs to the larger superfamily of ATP-binding cassette (ABC) transporters, membrane proteins found from bacteria to humans that burn cellular energy in the form of ATP to pump molecules across membranes. In plants, ABC transporters ferry an astonishing variety of cargo: defense compounds, heavy-metal chelators, waxes and, critically, phytohormones. Within this superfamily, the B subgroup—full-length transporters containing both nucleotide-binding domains and transmembrane domains—has attracted particular attention because several of its members in the reference plant Arabidopsis thaliana are proven hormone movers. AtABCB1 and AtABCB19, for instance, are celebrated auxin efflux carriers that help establish the localized auxin gradients sculpting virtually every organ of the plant, and recent structural work has revealed that AtABCB19 also exports brassinosteroids, the steroid hormones that drive stem elongation, vascular development and stress resilience.

What happens to these functions in crop plants with larger, more complex genomes has been far less clear. The Brassica genus, which includes cabbage, broccoli, oilseed rape and choy sum, descended from a common ancestor with Arabidopsis through extensive whole-genome triplication followed by diploidization and rearrangements, meaning that for every Arabidopsis ABCB gene there may be several Brassica relatives whose functions have diverged, specialized or been lost. Floting Chinese cabbage is an economically important vegetable in southern China, and its tall, succulent stalk is the product of vigorous cell elongation intimately tied to brassinosteroid and auxin action—making it an ideal system in which to ask what the ABCB repertoire is doing.

To answer that question, the team carried out a genome-wide survey of the B. rapa var. parachinensis genome, an effort made feasible by a recent high-continuity genome assembly of the species. Using rigorous bioinformatic screening, they identified 36 BraABCB genes and constructed phylogenetic trees that placed them into four evolutionary groups, consistent with the canonical architecture of ABCB families described in other angiosperms. Conserved domain analysis confirmed that the predicted proteins carry the hallmarks expected of functional transporters:Walker A and Walker B motifs and the signature C-loop within the nucleotide-binding domains, together with the membrane-spanning α-helices that form the translocation pathway. When Arabidopsis ABCB proteins were included in the phylogeny, the BraABCB27 and BraABCB28 proteins clustered tightly with AtABCB1 and AtABCB19, immediately flagging them as the closest functional relatives of the best-characterized hormone transporters in any plant.

Evolutionary forensics added context to this inventory. The researchers mapped each BraABCB gene to its position on the ten chromosomes of the species and examined collinearity—the synteny between genomic regions—to trace how the family expanded. Synonymous and nonsynonymous substitution rates (Ka/Ks) calculated for duplicated gene pairs told a story of constraint: with Ka/Ks values well below one, most BraABCB paralogs have been maintained under purifying selection, meaning that the protein-coding sequences have been preserved largely intact since their duplication. In other words, the family did not balloon through sloppy replication but rather through ancient genome duplication events whose products the plant has carefully conserved—an evolutionary signature typically associated with genes that matter.

But conservation at the sequence level is only a hypothesis about function; the real test lies in where the genes are switched on and what the proteins do. The group focused its experimental attention on Group IV, the clade containing the AtABCB1/19 relatives. Mining the promoter regions upstream of these genes revealed a rich catalog of cis-regulatory elements implicated in hormone responsiveness and abiotic stress, including motifs associated with drought, heat, and both auxin and brassinosteroid signaling. To put these predictions to the test, the researchers grew choy sum seedlings under several perturbations—drought stress, elevated temperature, exogenous application of the bioactive brassinosteroid brassinolide, and treatment with the primary natural auxin, indole-3-acetic acid—and quantified the expression of selected Group IV BraABCB genes by reverse-transcription quantitative PCR using the standard 2^-ΔΔCT method. The results were striking in their diversity: individual family members responded differently and often in opposite directions to the same stimulus, indicating that the Brassica expansion of this family is not mere redundancy but a differentiated toolkit, with distinct transporters likely deployed in different tissues, developmental stages and environmental contexts.

Subcellular localization supplied a further piece of the puzzle. Because ABCB transporters must sit in a membrane to move hormones across it, the team fused Group IV BraABCB proteins to fluorescent reporters and expressed the constructs to determine where the fusion proteins accumulated in living cells. The analyses showed predominant localization to the plasma membrane, exactly where an efflux carrier engaged with extracellular signaling and long-distance hormone movement would be expected to reside. This placement also matters for a second reason: the brassinosteroid receptor itself, BRI1 (BRASSINOSTEROID-INSENSITIVE 1), is a leucine-rich repeat receptor kinase embedded in the plasma membrane, discovered in the 1990s as the cell-surface sensor for the steroid hormones. If ABCB transporters share membrane real estate with BRI1, opportunities for physical and functional crosstalk multiply.

That possibility is precisely where the new study makes its most intriguing contribution. In Arabidopsis, a regulatory protein called TWISTED DWARF1—an immunophilin-like co-chaperone—has been shown to associate physically with BRI1 and to be required for the full activity of the ABCB1- and ABCB19-mediated auxin transport machinery; mutations in the corresponding gene produce pleiotropic, hormone-defective growth phenotypes, and the interplay between ABCB transporters and BRI1-related membrane complexes has become a model of how transport and signaling are coordinated at the cell surface. Using bimolecular fluorescence complementation (BiFC), a technique in which two proteins are fused to halves of a fluorescent protein so that a physical interaction brings the halves together and restores fluorescence, together with a split-ubiquitin yeast two-hybrid assay that is better suited to membrane proteins, the Guangzhou University team tested whether the choy sum homologs could engage BRI1-related proteins. The answer was yes: BraABCB27 and BraABCB28, the two closest relatives of AtABCB1 and AtABCB19, showed detectable physical associations with BRI1-related proteins in both systems.

The authors are careful in their claims—and appropriately so. Detecting an association is not the same as demonstrating a transport function, and the study stops short of showing that BraABCB27 and BraABCB28 actually pump brassinosteroids or auxin in planta. What the work provides instead is a complete, experimentally grounded framework: a full inventory of 36 genes with their evolutionary history, a demonstration that Group IV members are stress- and hormone-responsive, confirmation of plasma-membrane targeting, and two strong candidates whose interaction with BRI1-related proteins now invites direct functional interrogation. The logical next steps—CRISPR knockout or overexpression of BraABCB27 and BraABCB28, followed by measurements of stalk elongation, brassinosteroid distribution and auxin gradients—would test whether these transporters are genuinely part of the machinery that regulates the trait for which choy sum is grown.

The broader implications reach beyond a single vegetable. Brassinosteroids have become a major focus of crop engineering because of their capacity to enhance yield, stress tolerance and architectural traits, and recent structural biology has revealed the atomic details of how the Arabidopsis ABCB1 and ABCB19 transporters export these steroids. Whether the same transport-signaling coupling exists in Brassica crops—and how the triplicated genome has diversified it—remains open, but this study supplies the map on which such questions can now be asked. For breeders seeking taller stalks, denser flowering or improved drought resilience in flowering Chinese cabbage, the BraABCB family just moved from anonymous genomic baggage to a shortlist of actionable targets. And for plant biologists more generally, the finding that ABCB-BRI1 associations are conserved in a distantly related crop reinforces an emerging picture: in plants, hormone transport and hormone perception are not separate layers of regulation but physically intertwined systems working at the same membrane, in the same cells, at the same time.

For much of evolutionary history, plants were confined to aquatic environments, growing solely in two dimensions along flat surfaces. This limited their ability to diversify and develop structurally complex organs, confining them to simple forms. Roughly 470 million years ago, however, an extraordinary transformation occurred. Plant cells acquired the machinery to divide and expand in three spatial dimensions, allowing growth upwards and laterally. This developmental breakthrough set the stage for terrestrial ecosystems.

Scientists have long sought to decipher the molecular underpinnings of this critical shift. While previous studies have emphasized the role of gene regulation—specifically how genes controlling growth are switched on and off—the latest research highlights the necessity of metabolic and protein-level control as well. At the center of this discovery is a protein named RAK1, newly identified in the model moss species Physcomitrium patens, a representative of some of the earliest land plants.

RAK1 stands out because it represents an evolutionary fusion of two distinct protein functions—an N-acetyltransferase and a mitogen-activated protein kinase (MAPK). This fusion protein combines signaling capabilities with metabolic regulation, serving as a molecular bridge that enables cells to integrate external signals with internal biochemical processes. This integration is crucial for orchestrating the complex cellular events required for three-dimensional growth and budding.

To probe RAK1’s function, researchers used sophisticated genetic techniques to create moss variants with and without this protein. The absence of RAK1 led to pronounced developmental abnormalities: cells failed to divide properly in multiple orientations and produced malformed buds. This phenotype underscores RAK1’s role as an essential regulatory nexus for developmental reprogramming, without which the moss cannot transition effectively from flat filamentous growth to the formation of complex three-dimensional structures.

This discovery challenges the prevailing paradigm that gene expression changes alone drive plant developmental complexity. Instead, it suggests that precise coordination between signaling pathways and metabolic state is equally vital. RAK1’s dual functionality exemplifies how evolutionary innovation sometimes emerges not by inventing wholly new proteins but by recombining existing domains to create multifunctional molecules that can regulate complex processes more efficiently.

The implications extend beyond moss or even plants. Stem cells across multicellular organisms, including humans, rely on tightly controlled metabolic networks during growth and differentiation. Understanding RAK1’s mechanism offers intriguing parallels that could inform broader biological principles of developmental regulation, potentially shedding light on cellular growth control in diverse systems.

The study also opens new avenues for exploring how ancient molecular fusions contributed to the conquest of land by plants. By timing the emergence of such proteins, scientists may better reconstruct the evolutionary events that shaped terrestrial ecosystems. It also raises fundamental questions about molecular innovation through domain fusion and the evolutionary pressures that select for such multifunctional proteins.

Furthermore, this research reinvigorates interest in Physcomitrium patens as a powerful and accessible model organism for studying plant biology. Its relatively simple body plan, combined with advanced genetic tools, enables high-resolution dissection of developmental pathways. Insights gleaned from moss systems often illuminate fundamental mechanisms conserved through plant evolution.

Overall, the identification and characterization of RAK1 deepen our understanding of how plants evolved the capacity for architectural complexity. This protein exemplifies a crucial molecular switch that enabled early land plants to break free from two-dimensional constraints and establish the verdant landscapes that support life today. It stands as a testament to nature’s ingenuity in repurposing existing components to forge new biological capabilities.

This research was published in the journal New Phytologist and represents an international collaborative effort involving experts from Austria, England, Germany, Japan, and Denmark. It underscores the power of interdisciplinary and cross-border scientific endeavors to unravel the mysteries of life’s grand evolutionary transitions.

As science continues to explore the molecular machinery driving development and evolution, discoveries like RAK1 not only answer longstanding questions but also open new chapters in understanding life’s complexity at the cellular level. The fusion of signaling and metabolic regulation embodied in RAK1 may prove emblematic of a broader principle shaping the evolution of multicellular life.

Subject of Research:
RAK1 protein’s role in enabling three-dimensional growth in moss and its implications for plant evolution.

Article Title:
An N-acetyltransferase-MAPK fusion protein modulates developmental reprogramming in Physcomitrium patens

News Publication Date:
13-May-2026

Web References:
https://nph.onlinelibrary.wiley.com/doi/10.1111/nph.71214

References:
The study published in New Phytologist, DOI: 10.1111/nph.71214

Image Credits:
Photos by Laura Moody

Keywords:
RAK1, moss, three-dimensional growth, plant evolution, Physcomitrium patens, protein fusion, N-acetyltransferase, MAPK, developmental reprogramming, stem cells, metabolism, evolutionary innovation