Biologists Engineer Larger, Tougher Crops for Fuel, Bioproducts

Disruption of plant gene boosts lignin production, growth, and immunity

September 3, 2025

UPTON, N.Y. — Cell walls don’t just provide support and protection for plants — they’re also packed with energy-rich biomaterials that could open new pathways for additional fuel, chemical, and material sources in the U.S. That’s why biologists at the U.S. Department of Energy’s (DOE) Brookhaven National Laboratory are untangling the complex genetic mechanisms that regulate these useful plant materials, known as biomass.

In a study just published in The Plant Biotechnology Journal, the research team identified a plant protein that plays a key role in three important biological processes in poplar plants — iron deficiency responses, cell wall biosynthesis, and the synthesis of disease-fighting molecules.

“The protein is called PtrbHLH011, and it first caught our attention several years ago when we were identifying genes and proteins that influence how poplar plants respond to nutrition stresses,” said Meng Xie, a Brookhaven biologist and the lead author on the new paper. “We found that expression of the PtrbHLH011 gene was largely reduced in stressed plants growing in an iron-deficient medium.”

During photosynthesis, plants need iron to convert sunlight into chemical energy that powers growth. With a deeper understanding of how plant genes and proteins like PtrbHLH011 work, biologists are working to develop bioenergy crops that can hyperaccumulate this important mineral and thrive even on iron-deficient, marginal land.

Traditionally, researchers have worked to increase cell wall sugars that can be converted into biofuels. But in recent years, a rigid cell wall component called lignin has caught their attention because it can be used to produce valuable bioproducts with industrial applications, like cement and adhesives.

“Different environmental factors can affect not just cell wall biosynthesis but also the ratio of cell wall components, like sugars and lignin,” Xie explained. “We set out to study the molecular mechanism underlying this so-called ‘environmental plasticity.’”

Gene “knock out” with a big payoff

Because some proteins have overlapping roles — or several, seemingly unrelated roles — it can be difficult to untangle the function of one from another. So, biologists often “knock out,” or deactivate, a gene to better understand the function of the protein it codes for.

In this case, collaborators at the University of Maryland developed poplar plants lacking PtrbHLH011.

The knockout plants simultaneously produced twice as much lignin and exhibited enhanced growth for the first time ever. This was especially surprising because prior studies show that increasing lignin content — and consequently, stiffening cell walls — typically diverts energy from growth and limits the overall biomass yield.

The modified plants also accumulated three times more iron in their leaves and increased production of flavonoids, which are compounds that can help plants fight disease.

Consistent with these observations, plants engineered by Brookhaven biologists to overexpress the PtrbHLH011 gene exhibited the opposite traits: stunted growth, weaker cell walls, increased sensitivity to disease, and yellow leaves characteristic of nutrient stress.

“PtrbHLH011 is a special type of protein called a transcription factor, meaning it binds to specific sequences of plant DNA and regulates the expression of several target genes,” explained Yuqiu Dai, a postdoctoral fellow at Brookhaven Lab and a first author on the new paper. “So, we expected that disrupting the PtrbHLH011 gene would affect several biological processes associated with its target genes.”

However, the researchers were surprised to find that knocking out the PtrbHLH011 protein increased several processes that require significant amounts of energy, which would normally impose a significant metabolic burden for the plants.

“We suspect the three-fold increase in leaf iron content boosted photosynthesis in the plants, ultimately generating more energy to support plant growth and the synthesis of lignin and flavonoids,” said Xie.

The surge in flavonoid synthesis is especially compelling as biologists from Brookhaven and beyond ramp up bio-preparedness efforts to protect U.S. bioenergy plants from disease. Through future studies examining how plants respond to infection and disease, researchers aim to uncover underlying mechanisms that could be leveraged to strengthen crops’ resistance to pathogens that reduce biomass yield.

With the identification of a gene regulatory mechanism that is modulated by PtrbHLH011, the Brookhaven researchers are also working to fine tune the expression of its specific target genes.

“If we can modulate the individual ‘downstream’ target genes, rather than a transcription factor that regulates all of them, we’ll be able to more precisely control one biological process at a time,” Dai said.

Xie added, “The foundational understanding we established during this study will enable our biotechnology efforts to advance the production of bioenergy and bioproduct feedstocks.

“These findings were the result of successful integration of multiple DOE Office of Science facilities and capabilities,” said Xie. For example, collaborators at the Joint Genome Institute measured the levels of gene expression in the engineered plants, and a collaborator at the Molecular Foundry provided insights into how the newly discovered regulatory mechanism adapted as land plants, like poplar, evolved. The Brookhaven researchers used confocal microscopy at the Center for Functional Nanomaterials (CFN) to visualize where PtrbHLH011 was expressed in plant cells. They measured biomass lignin content in Brookhaven’s Biology Department. And with collaborators from the National Synchrotron Light Source II (NSLS-II), the researchers conducted X-ray bio-imaging experiments at the Submicron Resolution X-ray Spectroscopy (SRX) and Life Science X-ray Scattering (LiX) beamlines to study iron accumulation and cell wall structure in the poplar plants.

The Joint Genome Institute and the Molecular Foundry are DOE Office of Science user facilities at DOE’s Lawrence Berkeley National Laboratory. CFN and NSLS-II are DOE Office of Science user facilities at Brookhaven Lab.

This research was supported by the DOE Office of Science.

Brookhaven National Laboratory is supported by the Office of Science of the U.S. Department of Energy. The Office of Science is the single largest supporter of basic research in the physical sciences in the United States and is working to address some of the most pressing challenges of our time. For more information, visit science.energy.gov.

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