How maize leaves build bridges between their veins

Maize leaf veins do more than run alongside one another. Short transverse veins connect neighboring longitudinal veins, creating an interconnected network that carries water, nutrients and products of photosynthesis. How those cross-connections are specified and assembled has remained less understood than the development of the parallel veins themselves.

In a bioRxiv preprint posted on November 2, 2026, Maria Camila Medina and colleagues describe a sequence in which small groups of initial cells mark future connections, neighboring cells join the developing strand, and the resulting tissues integrate with existing veins. The study, which has not been peer reviewed, also identifies two genes important for producing these links.

Building a connection from both sides

The researchers examined developing maize leaves using tissue sections and fluorescent markers. One marker, PIN1a-YFP, makes an auxin-transport protein visible and helps reveal early vascular development.

Transverse vein traces first appeared around a developmental stage called P6, initially near the leaf tip. Their formation then progressed toward the base. Connections in the sheath, the lower leaf region surrounding the stem, began later, at P8.

The earliest visible signs were opposing groups of initial cells beside adjacent longitudinal veins. Cells in the intervening ground tissue initially lacked detectable PIN1a-YFP fluorescence. They subsequently acquired the signal as they became incorporated into the new vascular strand.

Live imaging captured this recruitment within the first 12 hours of observation. Cell divisions followed between 12 and 24 hours, building a thicker, ladder-like strand. By 36 hours, the developing connection contained multiple cells. More cells were recruited where neighboring veins were farther apart, particularly in the sheath.

These observations support the authors’ two-step model: initial cells establish the future connection’s position, followed by recruitment and division of intervening cells. What specifies the initials and coordinates recruitment remains unresolved.

Auxin affects organization without stopping initiation

Auxin is a plant hormone with a major role in vascular development. The team tested its contribution by treating exposed young leaves with the auxin-transport inhibitors NPA and TIBA, or by supplying additional auxin.

Added auxin accelerated recruitment and division, producing two cell layers within 12 hours. Transport inhibitors did not eliminate transverse vein initiation, but they changed cell behavior and strand organization. Higher-dose NPA frequently inhibited subsequent divisions, while TIBA-treated cells often developed abnormal shapes.

The researchers also used DR5, a fluorescent reporter of auxin-responsive gene activity. Detectable DR5 activity appeared later than PIN1a-YFP, first in the initials and then in recruited cells as the strands differentiated.

The authors interpret these findings as evidence that auxin transport helps organize transverse veins, while an earlier developmental pre-pattern may help initiate them. Persistence under inhibitor treatment supports that possibility; it does not establish that auxin has no role in initiation. The signals underlying the proposed pre-pattern remain unknown.

Two genes emerge from the developmental timeline

To identify candidate regulators, the team sequenced RNA from young leaves across developmental stages, using four independent biological replicates. Some leaves were also separated into upper regions containing transverse veins and lower regions without them.

The analysis identified 11 groups of genes with distinct expression patterns. One group became active before the later PIN1a expression increase associated with transverse vein development. It included EREB41 and EREB114, genes encoding transcription factors—proteins that regulate gene activity.

Plants carrying disruptions in both genes produced fewer transverse veins. The researchers observed reductions using both transposon-insertion mutations and CRISPR-generated mutations. In the sheath, reductions reached up to 95% compared with wild-type plants. Approximately a quarter of the remaining transverse veins were abnormal, including incomplete connections and loops returning to the same longitudinal vein.

The double mutants also had narrower leaves. That broader effect leaves an important question: how much of these genes’ activity is specific to vascular patterning rather than leaf development more generally? Their precise position in the regulatory sequence also awaits further genetic work.

Joining the existing network

Anatomical sections showed that mature transverse veins establish continuous xylem, phloem and surrounding bundle-sheath tissue with neighboring longitudinal veins. At insertion points, the supporting sclerenchyma ring was locally interrupted, accommodating the new connection.

Together, the findings provide a cellular and genetic account of how maize adds cross-links to its vascular network. They address a developmental process relevant to leaf organization and photosynthesis research, rather than demonstrating an improvement in crop performance.