Their finding, published in the July 11, 2003, issue of the journal Cell, is a major step forward in understanding plant fertility. The discovery could also help researchers understand similar biological processes, such as now nerve cells find each other and make appropriate connections. It may even provide clues about repairing spinal cord injuries.
"Since agriculture, which supplies nearly 80 percent of the world's food supply, depends so profoundly on plant fertility, understanding this process is fundamentally important," said Daphne Preuss, Ph.D., professor of molecular and cell biology and an investigator in the Howard Hughes Medical Institute at the University of Chicago.
When a pollen grain is deposited on the surface of a flower, it somehow has to grow a tube from the stigma of the flower, past several different cell types to where the eggs are, digesting tissue as it grows to burrow all the way inside. "While a few molecules involved in this process have been identified over the years," said Preuss, "we really still don't understand how this tube gets from start to finish."
Working with Arabidopsis, a popular model plant, Preuss and colleagues from her lab found that plants produce a carefully controlled gradient of gamma-amino butyric acid (GABA), a molecule best known for its role in the mammalian nervous system, to lure a pollen tube toward the egg cells. GABA acts like a light at the end of a tunnel, stimulating the initial growth of the pollen tube and shining ever brighter as the tube gets closer to its goal.
The researchers found that the key to regulating GABA levels is an enzyme they named POP2 that degrades GABA. Arabidopsis flowers produce high levels of GABA then eliminate varying amounts of it from different structures, so that a small amount is present at the surface of the pistil, where it stimulates pollen tube growth. Higher concentrations are found closer to the eggs, leading the tubes toward the target.
The study grew out of the team's chance finding of abnormal pollen tubes on plants that were later found to lack POP2.
"We saw the pollen tubes just winding around and totally missing their targets on one particular mutant," said Preuss. Co-author Laura Brass, a former Ph.D. student in the Preuss lab, analyzed the mutant strain and pinpointed the gene that caused the defect, which the researchers named POP2.
By comparing the sequence of the defective protein produced by POP2 to other known proteins, lead author Ravishankar Palanivelu, Ph.D., a post-doctoral fellow in Preuss's laboratory, concluded that it was an enzyme called an aminotransferase. It was not until the researchers found that the mutant plant contained a hundred-fold elevation in GABA, however, that they learned which molecule the enzyme degraded.
Further studies confirmed that the chemical normally concentrates near the egg-containing ovule. In contrast, in the mutant plants, GABA is diffused throughout the tissues. In these mutants, the pollen tubes are "just overwhelmed with signal," said Preuss. Instead of a light at the end of the tunnel it was "like staring at the sun."
Finding a reproductive role for GABA in plants is a good example of nature's ability to make the most of what's available, said Preuss. GABA is small, comparatively simple molecule. Many plants and animals use it as a source of carbon or nitrogen or to send signals from cell to cell. Animals use it to regulate hormone secretion, inhibit certain signals between nerve cells, and perhaps even to guide embryonic neurons to their destination.
GABA is only one of several substances involved in pollen tube guidance, however, said Preuss. The researchers are analyzing other mutants with altered pollen tube growth.
The research was supported by the Department of Energy, the Searle Scholars Program and the University of Chicago.
Journal
Cell