Among plants, growth forms vary from tiny herbs and woody shrubs to climbing vines and charismatic tall trees. A tremendous amount of research has uncovered the fundamentals of how trees gain their massive stature and thick stems. This process, called “secondary growth,” generates both xylem (wood) and phloem (inner bark) through active cell divisions in the trunk each growing season, which progressively builds the width of a tree. Secondary growth is a critical process in plant growth and development. It provides mechanical support for the growing crown and a continuous vasculature system to transport water and nutrients from the depth of the soil to the most juvenile leaves at the top of a tree. Historically, our understanding of secondary growth was restricted to careful observation of cells and tissues under the microscope, however with advancements in technologies came advancements in science. Today, we have sequencing technologies that harness the power to pinpoint exactly which genes in the DNA of plants control secondary growth. Most research in this area focuses on trees. But trees are not the only plants that undergo secondary growth.
Indeed, climbing woody vines, known as lianas, also generate secondary growth, although in highly modified fashions. In contrast to trees that form a ring of xylem surrounded by phloem, an unusual organization of these vascular tissues can be found in the stems of many lianas—which range from cylindrical stems with assorted distribution of xylem and phloem to asymmetrical and/or lobed stems. Why do vines have such unusual stem architecture? Scientists hypothesize that these alternative anatomies promote increased stem flexibility, mechanical support, and water/sugar translocation, contributing to the vine’s ability to climb up a supporting tree in the forest or a backyard trellis. But how do these complex patterns develop? In our research, we aim to answer this question by investigating the genes controlling the unusual secondary growth of vines.
To answer these research questions, we are studying plants growing at the Arnold Arboretum of Harvard University and across the “Ag Quad” in the College of Agriculture and Life Science at Cornell University. We collect the stems of lianas, with the aim of understanding how their secondary growth differs from trees; therefore, we focus our investigations on the “vascular cambium”—a self-perpetuating group of cells (also called a “meristem”) that creates the complex conduit-like system formed by xylem to transport water and phloem to sugars. Both xylem and phloem are known as complex tissues as they are made up of more than one type of cells. These cells work in a harmonized manner to perform various functions, including water and nutrient transport, storage, and mechanical posture. In woody plants, when produced by the cambium they are called secondary xylem and secondary phloem, which comprise the wood and inner bark, respectively. The cambium is responsible for the bulk of radial growth in the large trunk of an oak tree or the robust stem of a climbing woody vine. To further our understanding of how the vascular cambium generates radial growth, we are employing an integrative approach combining developmental anatomy (morphology) and developmental genetics (molecular biology) through comparative transcriptomes—a method that generates a profile of all active genes during the collection of this biological sample. Therefore, this genetic approach will reveal the genes that are involved in the formation of the vascular cambium, illuminating the potential regulators that uniquely initiate diverse arrangements of vascular tissues that we observe in nature. Such studies are critical not only to comprehend the immense vascular diversity of woody plants, but also to understand how wood is formed, the very tissue that humans depend upon for construction, fuel, fire, and so much more. To study radial growth, many scientists have used model plants such as thale cress (Arabidopsis thaliana), eucalyptus (Eucalyptus grandis), or poplar (Populus spp.) (Tonn & Greb, 2017). Instead of trees and shrubs with the typical mode of radial growth that is observed in most woody plants, we opted for an unconventional approach, studying a group of woody vines with special morphological traits, the wisterias.
Why Wisterias?
“Wisteria” is both the common and scientific name of a small group of plants within the legume family. These twining woody vines are captivating ornamentals grown in gardens all over the United States. Landscapers, artists, and plant lovers value them due to their generous bloom, large flower clusters, various colors, and fragrance. Currently, only four species are recognized in the genus Wisteria, one being native to the US (W. frutescens) (Compton et al., 2019). All other species occur naturally only in temperate zones of China, Japan, and Korea (Compton et al., 2019).
Wisteria species can be identified by vegetative features including the size and number of leaflets—however reproductive characteristics such as type/position of inflorescence, length of flower clusters, or timing of blossom are more critical for identification (Compton et al., 2019). Surprisingly, flower color is not one of the most important features, as different species may have flowers of the same color. Similarly, some wisterias may be identified by looking at the direction that the plants twine. Chinese wisteria (Wisteria sinensis) and American varieties (W. frutescens) normally twine counterclockwise. Japanese wisteria (W. floribunda) grows clockwise.
Another interesting feature of wisterias is the way they achieve radial growth. Most woody plants produce a single solid mass of xylem surrounded by a continuous coating of phloem. However, wisterias climb and grow strong through the formation of multiple increments of xylem and phloem formed in a successive fashion, similar to beetroot or stems of mangrove trees. This phenomenon is called “successive cambia” and originates through the formation of a new vascular cambium that arises in an unusual position (ectopic) repeatedly. These additional vascular increments, which in some cases are concentric and continuous, are not the same as “growth rings”—the seasonal increment of xylem cells (i.e., wood only) with distinctive features (e.g., size, shape) which allow researchers to demonstrate the age of trees. New increments of successive cambia (wood + bark) are usually not annual, and not seasonally dependent; they may be associated with various functional adaptations in plants, including storage in the sugar beets or else an adaptation that maximizes the transport of water, mechanical resistance, and flexibility not only in wisteria but in several tropical woody vines. If you live in temperate regions such as the northeastern US, it is likely that no other plant with successive cambia besides wisteria is present in your garden or in the city park. Most woody plants exhibiting this type of radial growth occur in subtropical and tropical environments, distributed across different botanical families, generally including climbing plants (vines, climbers, twiners).
Because wisterias are such charismatic woody vines that naturally grow in temperate zones, we selected them as a biologically interesting system.
Because wisterias are such charismatic woody vines that naturally grow in temperate zones, we selected these plants as a biologically interesting system to investigate how successive cambia evolved within the seed plants. Over millions of years in the evolutionary history of vascular plants, the vascular cambium has remained conserved in most plant lineages, demonstrating its key biological function (e.g. hydraulic and mechanical properties). Because this vascular cambium is biologically similar in most plant lineages including in plants with and without successive cambia, this research will shed light not only on how wisteria grow by means of multiple vascular increments of successive cambia but will also contribute to the understanding of how large trees with the typical mode of secondary growth expand their large trunks, producing most of the woody biomass in nature.
In Search of the Perfect Wisteria
To investigate these questions on radial growth and the climbing habit, our original plan was to work with the American wisteria (Wisteria frutescens). That plan did not prove practical, as they are rather hard to find in nature and are not as vigorous (and showy) as the Asian species (Wyman, 1949). Slender plants are usually not satisfactory for this investigation since successive cambia in wisterias appear after several years of normal growth (Nejapa et al., 2021), with stem diameter larger than 2 to 3 centimeters. We then discovered the notable collection of wisterias at the Arnold Arboretum of Harvard University in Boston, which includes dozens of wisterias from all different species. This research was proposed to the Arboretum, and we were honored to receive the 2022 Sargent Award, which granted us the opportunity to collect wisterias (and other plants) in the Living Collection.
For botanists in temperate zones, summer is a special time for fieldwork. It was still early in the season when we went to the Arboretum to check on some of the eighteen accessioned American wisterias listed as being in “good” or “fair” condition. Most of these plants are displayed at the Leventritt Shrub & Vine Garden, where wisterias stand along with numerous other spectacular woody vines. These American wisterias were also slender and most likely without successive cambia, similar to some plants we had observed at Cornell University. Still, we examined the branch of one plant (359-2003*C), approximately 2 centimeters wide, growing up one of the trellises. We confirmed our expectations on the absence of successive cambia. Collecting large wisteria stems with successive cambia often involves cutting stems at the base of the plant. Such collecting practices are destructive, as these branches may be responsible for most if not the entire aerial plant body. Given this preliminary observation on Wisteria frutescens, we couldn’t risk collecting more plants at the Leventritt Garden, as collecting must be done in a way that does not impact the general public’s experience of the Arboretum. Our misfortune with American wisteria turned our collection sampling upside-down. Our next steps required some sleuthing—both historical and botanical. We had to turn our investigation towards one of the other Asian species. But which one was the most appropriate? We located a large Japanese wisteria (Wisteria floribunda) at the Arboretum. Michael Dosmann, the Keeper of the Living Collections, allowed us to collect a robust stem from the plant, which is an original W. floribunda brought from Japan after an expedition in 1977 (1894-77*A). The larger branch from this plant was 50 mm wide. We noticed that successive cambia were present in this branch, which gave us the opportunity to collect our first sample to investigate the genetics underlying the formation of successive cambia. This shows the importance of having diverse plant species in the Arboretum collection and the efforts of early Arboretum investigators to acquire them. However, we needed to find other samples of Wisteria floribunda because transcriptomics—the genetic approach used in our study—requires at least three to six separate specimens. Moreover, large stems with secondary growth are critical to investigate and understand stem anatomy of plants with successive cambia. From the plants growing in the Arboretum, we also noticed that, during that summer, only American wisterias had flowers—which may be related to phenology (age of the plant), seasonality (weather conditions) or horticultural practices (pruning). Flowers, however, provide useful information when trying to identify a plant species—and we would need as much information as possible to identify other W. floribunda.
Wisterias in Ithaca and Cornell University
Collecting only one specimen with successive cambia at the Arnold Arboretum made us return to Ithaca in search of more plants. Back in Ithaca, we discovered potential Asian wisterias in three different locations: one plant growing in the Ithaca Commons (downtown), seven to eight plants at A. D. White House (Cornell University), and another population on Cornell’s West Campus. The Cornell Botanic Gardens had one American (Wisteria frutescens) with flowers and two Chinese wisterias (W. sinensis) without flowers, but no Japanese wisteria (W. floribunda). These plants grow over a trellis ornamenting the Nevin Welcome Center. The wisterias growing at A. D. White House and on West Campus were accessible plants that could greatly complement and facilitate our research sampling. However, no one on campus knew the exact species identification of these plants.
How to Identify the Wisterias?
Since there were no flowers on the plants growing in Ithaca, getting a positive identification was our next mission. Examining the direction in which stems twine can narrow the species ID, but is not definitive, as multiple species twine in either given direction. By corresponding with James A. Compton, we also learned that in wisterias this characteristic is not conserved in cultivated species—a reason their paper did not use this feature in the key to identify species (Compton et al., 2019). Among climbing plants in general, it is known that the majority of species (>90%) are right-handed (counterclockwise) (Edwards et al., 2007) which is most likely genetically determined (Smyth, 2016). Notably, Wyman, a former horticulturist at the Arnold Arboretum, reported that “… in the case of Wisteria floribunda two plants of more than a dozen examined were found that twined in the opposite direction from the majority of this species” (Wyman, 1939), which highlighted the problem to identify wisteria species based on their handedness.
During the summer, as we were exploring the identification of wisterias, we also investigated the history of the plants growing on the Cornell Campus. We found that plants growing on West Campus were catalogued neither by Cornell Botanic Gardens nor by the Grounds Department of Facilities and Campus Services, which is responsible for landscaping. Indeed, there were wisterias near the parking lot between University and Stewart Avenue, but no one knew how they ended up there, let alone their species identification. While trying to dig out information about this locality, we simultaneously sought to learn more about the collection of wisteria growing at A. D. White House. Through conversations with numerous people and resources at Cornell, we began to learn not only about wisterias but also gained glimmers of the fascinating history surrounding horticultural plantings on campus.
A.D. White House: the Presidents, the Garden, and the Wisterias
The A. D. White House is a mansion located in the heart of Cornell’s campus. The house is named after Andrew Dickson White, Cornell University’s first president and co-founder. It served as the residence for Cornell University presidents from 1872 to 1949. Today, it is part of the College of Arts & Sciences, housing the Society for the Humanities (Cornell University, 2022). The garden surrounding the house was designed primarily by the fourth president’s wife, Margaret Livingston Farrand—popularly known as Daisy—who had a “serious interest in horticulture” and “the reputation of a highly skilled and distinguished amateur gardener” (Szekely, 2015). Not only an important figure in Cornell’s life, Margaret Farrand was also sister-in-law to Beatrix Farrand—known as one of the first and most celebrated landscape architects in the United States, responsible for many private and institutional gardens in the United States. Notably, Beatrix Farrand had a deep connection with Charles Sprague Sargent, the first director of the Arnold Arboretum (Beatrix Farrand Society, 2020). Unfortunately, no details were found on whether the in-laws communicated to indicate if wisterias were part of their respective gardening projects. Lisa Pearson, Head of the Library and Archives of the Arnold Arboretum, reported that no wisterias were sent from the Arnold Arboretum collection to Cornell University between 1910 and 1970 (only Viburnum and Acer were distributed between the institutions at that time). Since the Farrand family moved out of the A.D. White House 1937, it is unlikely that Margaret Farrand was involved with the wisterias present at Daisy’s Garden today. Edward Cobb, a now-retired researcher of Plant Sciences at Cornell and a fabulous source of institutional knowledge regarding plants, mentioned that wisterias have grown on campus since the ’60s, and they were radically pruned in the ’80s during renovations. As for identification, Kim Klein, who has maintained the garden for several years now, said that some plants have white flowers and others have blue flowers. Still, this did not suffice to identify the wisterias, as there is a wide range of flower colors even within the same species. We also communicated with Nina Bassuk, emeritus professor and program leader of the Urban Horticulture Institute at Cornell, who said that some plants could be Wisteria floribunda. This hint was corroborated by observing a voucher herbarium specimen collected by former staff members of the L. H. Bailey Hortorium Herbarium at Cornell University and by observing that some of these plants twined left to right (clockwise). Although this information was starting to point to one species (Wisteria floribunda), we were still not 100 percent certain. Besides, we still needed at least five more plants to complete our sampling.
The Revelation Through DNA Barcoding
To make sure we would have biological replicates from the same species, we decided to dive deeper into the biology of wisterias and explore potential genetic markers (a gene) that could accurately differentiate them. This method consists of using a short sequence of DNA from a specific gene to identify species, similar to ancestry DNA testing or scanning a barcode in a supermarket—hence the name “DNA barcoding.” In 2019, James A. Compton and collaborators generated an evolutionary (phylogenetic) tree for wisterias and closely related species. To build this tree, they used four different genes. We analyzed this data to decide which of those genes would be most informative to perform the DNA barcoding approach for wisteria. We selected the chloroplast intergenic spacer gene “ndhJ-trnF”. We used leaves to extract DNA from ten plants in Ithaca, including the one plant from the Commons, six plants from A. D. White House and three plants from West Campus. After a couple of days in the lab, our DNA barcoding analysis revealed that the specimen from the Ithaca Commons belonged to Wisteria sinensis while the other two populations were W. floribunda. Because most of the populations belonged to W. floribunda, including large-stemmed specimens with successive cambia, this species was an excellent candidate to understand the odd vascular patterns in the stems of woody vines.
We are currently investigating the first transcriptomes of wisteria stems from different developmental stages, which were paired with transcriptomes of the same developmental stages in common bean, another climbing plant from the legume family (Fabaceae), which does not form successive cambia. By comparing the transcriptomes from different developmental stages of the stem in wisteria and bean, we will be able to make several comparisons within each species and among species, further improving our ability to identify which key genes are turned on and off during the formation of successive cambia. Our hypothesis is that the genes involved in the formation of vascular cambium and secondary growth in trees are also involved in the development of successive cambia, but with distinct patterns of gene expression—that is, they might be turned on and off at different developmental stages, at different tissues and/or at distinct levels. Once we describe the gene expression similarities and differences between the stems of wisteria and bean, other pairs of plants will be included in our analysis to further reveal the genetic underpinnings of successive cambia across seed plants, a trait that has evolved more than a dozen times across seed plants.
From a research perspective, as we constantly visit these plants in the field, we also pay attention to other interesting aspects of climbing plants. For example, a conspicuous feature is the “searchers shoots”—young branches able to move and attach to the support—or the fact that successive cambia are prevailing in basal, contorted, or compressed stems. These apparently subtle characteristics may be clues to understanding how wisterias climb and grow thick and strong, reaching the top of trees, houses, and trellises. Undoubtedly, as we investigate the developmental mechanisms of cambium formation in wisteria, we cannot help but grow curious about how the stems of other woody vines are constructed. Do they have successive cambia too? Do they have similar gene expression patterns to produce secondary growth? Given that eastern North American-Asian distinct species receive high priority with respect to collection development in the Arnold Arboretum, wisteria also has the potential to stimulate the acquisition of new taxa (e.g., Padbruggea) to foster comparative studies of native plants in the US in the future. It seems that incorporating this temperate cousin of wisteria would not only expand the phylogenetic diversity within this clade, but would also bring horticultural benefits, since Padbruggea species are climbers with similarly beautiful flowers. In the end, we hope you continue to appreciate the beauty of wisterias as you wait for winter to pass to admire their blossoms, or you realize how they slowly strangle their support to climb to the top. Yet, if you find yourself wanting to discard a wisteria, we would be happy to help you by cutting some of the large stems to get more data, as we continue unraveling the mysteries of secondary growth in woody plants.
Israel L. Cunha Neto is a postdoctoral associate and Joyce G. Onyenedum is an assistant professor in the School of Integrative Plant Sciences and L.H. Bailey Hortorium at Cornell University.
Acknowledgements
We would like to thank Chelsea D. Specht for suggesting writing a story about wisterias and Jacob B. Landis for insightful discussions. We are grateful to the people involved in the collections at the Arnold Arboretum (Michael S. Dosmann, Devika Jaikumar, and Ellie Mendelson) and Cornell University (Grounds Department [Kim M. Klein], Cornell Botanic Garden, and L. H. Bailey Hortorium Herbarium [Anna M. Stalter]). In addition to people mentioned in the paper, we are also indebted to Corey Earle, Maddie Reynolds, Natasha Bishop, and Tyler Lurie-Spicer for helping us to dig into the A. D. White House history. We also thank Angelique A. Acevedo for helping with DNA extractions and amplification for DNA barcoding.
Works Cited
Arnold Arboretum of Harvard University. 1917. Wisterias. Bulletin of Popular Information. Arnold Arboretum of Harvard University, 3(8), 29–32.
Beatrix Farrand Society. 2020. Beatrix Farrand. <https://www.beatrixfarrandsociety.org/beatrix-farrand/>.
Compton, J. A., Schrire, B. D., Könyves, K., Forest, F., Malakasi, P., Mattapha, S., & Sirichamorn, Y. 2019. The Callerya Group redefined and Tribe Wisterieae (Fabaceae) emended based on morphology and data from nuclear and chloroplast DNA sequences. PhytoKeys, 125, 1–112. https://doi.org/10.3897/phytokeys.125.34877
Cornell University. 2022. A.D. White House History. https://societyhumanities.as.cornell.edu/andrew-dickson-white-house-history
Edwards, W., Moles, A. T., & Franks, P. (2007). The global trend in plant twining direction. Global Ecology and Biogeography, 16(6), 795–800. https://doi.org/10.1111/j.1466-8238.2007.00326.x
Nejapa, R., Cabanillas, P. A., & Pace, M. R. 2021. Cortical origin of the successive cambia in the stems of the charismatic temperate lianescent genus Wisteria (Fabaceae) and its systematic importance. Botanical Journal of the Linnean Society, 1–11.
Smyth, D. R. 2016. Helical growth in plant organs: Mechanisms and significance. Development, 143(18), 3272–3282. https://doi.org/10.1242/dev.134064
Szekely, B. B. 2015. Portrait of a gardener, Daisy Farrand at Cornell. Beatrice Beach Szekely.
Tonn, N., & Greb, T. 2017. Radial plant growth. Current Biology, 27(17), R878–R882. https://doi.org/10.1016/j.cub.2017.03.056
Wyman, D. 1939. Some twining vines. Arnoldia, 7(7), 33–36.
Wyman, D. 1949. The wisterias. Arnoldia, 9(5–6), 17–28.
From “free” to “friend”…
Established in 1911 as the Bulletin of Popular Information, Arnoldia has long been a definitive forum for conversations about temperate woody plants and their landscapes. In 2022, we rolled out a new vision for the magazine as a vigorous forum for tales of plant exploration, behind-the-scenes glimpses of botanical research, and deep dives into the history of gardens, landscapes, and science. The new Arnoldia includes poetry, visual art, and literary essays, following the human imagination wherever it entangles with trees.
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