The underlying research theme in the Stern laboratory is nuclear-cytoplasmic interactions. Within this framework, we study how chloroplast genes and metabolic activities are regulated by the products of nuclear genes, usually acting at the transcriptional or post-transcriptional level. Chloroplasts, as the site of photosynthesis as well as other metabolic pathways, have numerous roles in plant development, and responses to environmental stimuli. Our laboratory focuses on how these roles are modulated through intensive studies of global and individual gene regulation under normal growth or stress conditions. We use different techniques including genetics, plant transformation, and biochemistry to unravel these processes.

feature released -2011

Gene expression is a very complicated process, especially in the chloroplast. For chloroplast genes to be expressed, the DNA sequence is transcribed ("copied") into an RNA sequence that is either functional as an RNA molecule (for example, tRNA and rRNA), or is further translated to a protein counterpart. In addition, many chloroplast RNAs require modification(s) from a precursor form to a functional, mature RNA, such as trimming off extra sequence at the ends and cutting long stretches of RNA into their individual components. The regulation of these steps (DNA > RNA > protein) is known to be orchestrated by numerous proteins originating from nuclear genes, and imported into the chloroplast, such as ribonucleases and polymerases. Recently, the concept of RNA molecules (“noncoding” RNAs) having a regulatory role in both transcription and translation has been proposed for bacteria, plants and humans. David Stern’s laboratory is uncovering the function of these regulatory RNAs in the chloroplast. One such RNA, named AS5 for antisense to the 5S rRNA, was found to have a functional role in the maturation and stabilization of the 5S rRNA. This was determined using two different experimental approaches. First, postdoctoral associate Amber Hotto overexpressed AS5 in tobacco chloroplasts. This resulted in plants that grew slower than the wild-type tobacco, and had reduced accumulation of 5S rRNA. Second, postdoctoral associate Robert Sharwood synthesized numerous variations of AS5 and 5S rRNA precursor transcripts in the test tube, and incubated them together with an extract from spinach chloroplasts that contains the proteins necessary for RNA processing and degradation. Robert observed that adding increased amounts of AS5 to the 5S rRNA precursor with the spinach extract resulted in a decreased accumulation of the mature 5S rRNA. These results are a first step in deciphering the role of chloroplast noncoding RNAs. The lab continues to investigate the function of other noncoding RNAs that have recently been identified.

feature released -2008

Through photosynthesis, plants use sunlight to convert carbon dioxide and water into sugar and then release oxygen into the air. This process is critical to the plant’s production of energy, and to the balance of CO2 and oxygen in the earth’s atmosphere. If the ability of plants to absorb and sequester CO2 could be improved beyond their natural capacity, plants could play an even more important role in helping to mitigate global warming. In one research project, David Stern’s laboratory has concentrated on understanding the molecular regulation of CO2 fixation, which occurs in the chloroplasts of plant cells. It has long been known that plants rely on an enzyme called ribulose bisphosphate carboxylase/oxygenase (Rubisco) to sequester CO2, and that a tremendous amount of the plant’s energy is required to produce the enzyme. It was also known that to conserve energy, plants carefully calibrate how much Rubisco they need at any point in time. What was not known – and recently investigated by the Stern laboratory – is how Rubisco production is regulated in the chloroplast. The Rubisco molecule is composed of eight large subunits (LS) produced in the chloroplast and eight small subunits (SS) produced in the cytoplasm of the cell. The SS molecules migrate into the chloroplast from the cytoplasm where they join with the LS molecules to form Rubisco. But how does the plant regulate Rubisco production when LS and SS are produced by two different genes in two structurally separate parts of the cell? In research with transgenic tobacco plants, Stern and his colleagues showed that when an LS molecule cannot find an SS molecule to combine with, the LS molecule binds to its own messenger RNA,which in turn blocks, or prevents, the RNA from translating into more LS. In effect, the LS molecule shuts down its own production to save energy needed for other processes. This is called auto-regulation, and it controls the amount of Rubisco a plant produces. Other scientists had encountered problems when they attempted to over-produce Rubisco, because their efforts resulted in a Rubisco-production shutdown that was not fully understood. With the lab’s discovery of the regulatory process, scientists may one day be able to modify plants to produce extra Rubisco,which would enable them to sequester more CO2. The result should be increased yields and, perhaps, reduced CO2 in the atmosphere.

feature released -2007

The ribosome is a strange beast. Often likened to a factory, the ribosome can also be seen as a translator, using the four-letter code of RNA to build proteins with 20 kinds of amino acids. To accomplish this, the ribosome needs its many components-that is, dozens of proteins and a few specialized RNA molecules-to work together like a well-oiled machine. The situation is even more complicated for mitochondria and chloroplasts, cellular compartments with their own genes and ribosomes. Of the 54 proteins in a chloroplast's ribosome, 33 come from genes in the nucleus of the cell, while the chloroplast's own genes code for the rest. David Stern's lab studies how the chloroplasts and nuclei of plant cells accomplish ribosome construction and other feats of cooperation. In one experiment, research associate Tom Bollenbach investigated the origin of the four RNA molecules incorporated into chloroplast ribosomes. He knew that all four came from genes adjacent to each other on the chloroplast chromosome, and were copied into RNA in one long stretch. Bollenbach wanted to learn how that stretch of RNA gets cut up into useful form. He and other lab members started by identifying three nuclear plant genes that looked similar to a bacterial gene known to be involved in RNA processing. Using microscopy, they found that only one of the three acted in the chloroplast. The lab next knocked out this gene, RNR1, and got white plants that couldn't photosynthesize: their chloroplasts weren't working. Uncut ribosomal RNAs were building up in these mutant chloroplasts, Bollenbach found, and their ribosomes weren't coming together. So the RNR1 gene must code for an enzyme that processes the ribosomal RNA. Intriguingly, cells sense the buildup of unprocessed RNAs in the absence of RNR1, and scale back production of the chloroplast proteins needed to make the ribosome. The lab is working to determine how plants manage this regulation.

feature released -2006

A long time ago, in a land of primordial goo, two bacteria lent new meaning to the word "teamwork." It didn’t start out that way, most scientists think: One bacterium probably ate the other. But it turned out that the two survived better together than they had independently, and their progeny evolved into a single, more sophisticated organism. This process happened at least twice, yielding the specialized cellular compartments chloroplasts and mitochondria. David Stern's lab studies chloroplasts, light-harvesting factories unique to plants and algae. While structures called nuclei contain most genes in plant cells, chloroplasts hold a few of their own. Most of these genes have to do with harnessing energy from sunlight to make sugars, a process agricultural scientists hope can be made more efficient through genetic engineering. But since the nucleus controls how the chloroplast's genes operate, understanding their interaction—the theme of Stern’s research—is essential. In one project, the Stern lab used a mutant single-celled algae to find how chloroplast gene activity decreases when cells are starved of sulfur. The mutation occurred in a gene in the nucleus that scaled down copying of DNA to RNA under low-sulfur conditions. By comparing the mutant with normal algae, lab members found that the protein affected gene copying not only in the nucleus (as previously thought), but also in the chloroplast. Since plants need sulfur-containing proteins to process light energy and avoid its potentially harmful effects, this system may protect a sulfur-starved cell by cutting how much light it absorbs. This mechanism is just one step in the delicate choreography between nuclei, chloroplasts, and the environment. Future experiments on their relationship will add a novel chapter to our understanding of evolution—one that explains how three organisms became one.

News

• BTI President Participates in Plant Science Summit (September 29, 2011)
• Boyce Thompson Institute President David Stern, PhD, Awarded Fellow of the American Society of Plant Biologists (June 4, 2010)
• The Chlamydomonas Genome Reveals the Evolution of Key Animal and Plant Functions (Nov 6, 2007)
• BTI Postdoc (Feb 2, 2006)
• Stern Featured on MicrobeWorld Radio Program (Dec 13, 2005)
• Lin & Bollenbach win Bogorad Award (Dec 19, 2005)

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