Introduction

Photosynthesis provides the organic material required for virtually all life on Earth. Cyanobacteria, algae, and plants also produce oxygen during photosynthesis, which has established and maintained our planet’s oxygen-rich atmosphere over the past few billion years. Cyanobacteria are the only prokaryotes that produce oxygen during photosynthesis and are major contributors to global photosynthesis and oxygen production. They also hold great promise for converting solar power to clean, renewable energy sources.

Our understanding of oxygenic photosynthesis has been primarily advanced by studying cyanobacteria as model organisms, and we have developed a detailed understanding of the fundamentals of photosynthesis. Now, research efforts are focusing on how photosynthetic and other cellular machinery acclimates to changing abiotic parameters, such as light color, intensity, and nutrient availability. These responses and their regulation are important to determine if we are to understand how photosynthesis operates in the natural environment.

Efficient photosynthesis requires effective photon capture by light harvesting antennae, which are called phycobilisomes (PBS) in cyanobacteria (Fig 1). Varying environmental conditions cause these macromolecular structures to change in composition, size, shape, number, and degree of association with photosynthetic reaction centers. Working with several marine and freshwater species, our group investigates how cyanobacteria sense their environment and transform this information into coherent cellular responses, particularly those affecting the phenotypic plasticity of the PBS.

Below are short summaries of the four major research areas of the Kehoe laboratory.

 

Uncovering Mechanisms Regulating Red-Green Light Acclimation

Complementary chromatic acclimation (CCA) is arguably the best studied example of how photosynthetic organisms modify their light harvesting structures and cellular processes to accommodate changes in ambient light color. This process occurs in many freshwater and marine cyanobacteria. The most dazzling consequence of CCA is the massive restructuring of the PBS. The process is most sensitive to red light, which leads to the production PBS containing the brilliant blue blue-pigmented protein phycocyanin 2 (PC2), and green light, which results in synthesis of PBS containing a pink-pigmented protein called phycoerythrin (PE) (Fig 1A). These changes, which are reversed by changing the ambient light color, make the cells are reddish-pink in green light and green-blue in red light (Fig 1B). CCA allows these cells to modify the light absorption characteristics of their PBS to best match the colors of light available for photosynthesis at various depths in lakes and oceans.

The expression of many genes is regulated by this process, including many that encode PBS components. In particular, two distinct sets of genes encoding alternative forms of PBS proteins are diametrically light-color regulated during CCA. Genetic studies have established that CCA regulation occurs through at least two pathways, a transcriptional control pathway called the Rca system and a post-transcriptional regulatory pathway named the Cgi system. The Rca system is a complex phosphorelay consisting of a sensor called RcaE, which was the first discovered member of a superfamily of prokaryotic photoreceptors called phytochromes, and two response regulators called RcaF and RcaC. We have recently shown that RcaC acts as both a transcriptional activator and repressor by binding to specific regulatory elements upstream of both red and green light expressed genes. We are currently identifying components of the Cgi system, which operates through the 5' leader sequence of a gene that is up regulated in green light and may involve components of the core translation initiation machinery.

 

Understanding Blue-Green Light Acclimation in Marine Cyanobacteria

Another important light acclimation response of cyanobacteria is called CA4, which allows marine cyanobacteria of the genus Synechococcus to adjust their light harvesting characteristics to optimized the absorption of blue and green light for photosynthesis (Fig 3, cultures 3b and 3c). This process is extremely important from a global ecological perspective, since marine Synechococcus strains are one of the two dominant cyanobacterial genera in the world’s oceans and a large percentage of the members of this genus are CA4-capable. However, there has been limited progress in understanding CA4 since its discovery 10 years ago because working with CA4 strains in the laboratory is challenging and no genetic system has been developed for a CA4-capable strain.

We collaborate with the laboratory of Dr. Frederic Partensky at the CNRS Station Biologique in Roscoff, France, to study CA4 (Fig 4). We have developed methods for successfully maintaining CA4-capable strains on plates, developed drug resistant markers, and created NimbleGen whole genome tiling arrays, which we have used to demonstrate that the expression of hundreds of genes are differentially regulated during CA4. Among these are genes encoding several putative transcription factors and enzymes that are likely to play roles in the process of CA4. Interestingly, our array results strongly suggest that the red-green light regulated process of CCA and the blue-green light regulate process of CA4 are significantly different in their physiology and regulation and likely arose via convergent evolution. Our collaboration will continue as we develop and use molecular tools for conducting studies of the regulatory mechanisms controlling CA-4 while at the same time carrying out studies to understand the ecological conditions under which CA4 is selectively advantageous. Collectively, these studies will provide a powerful convergence of molecular and ecological information on a globally environmentally important light acclimation process in the marine environment.

Fig 3. Marine Synechococcus strains are colorful and include CA4 types (3b and 3c). (Photo courtesy Dr. Fred Partensky.)

Fig 4. CNRS Station Biologique in Roscoff, France

 

Characterizing a Proteome Remodeling Response To Sulfur Limitation

When an element becomes limiting in the environment, cells may mobilize internally stored forms of the element and simultaneously reduce their requirement for that element. Both of these goals can be accomplished through “proteome remodeling”, whereby a portion of the cell’s proteome that is enriched in that element is degraded and replaced by isozymes that contain much less of the element. Thus, the proteome itself is the storage organ for that element. This nutrient limitation response may be widespread among bacteria in nature. Proteome remodeling was first described as part of response to sulfur limitation in a CCA-capable freshwater cyanobacterium. In addition to light harvesting antennae proteins PC1 (not CCA-controlled) and PC2 (CCA controlled), this organism contains a light harvesting protein called PC3 (Figs 1 and 5) . Soon after red light grown cells are limited for sulfur, the amount of cpc1 and cpc2 RNA decreases and cpc3 RNA increases. The corresponding protein levels also change: PC1/ PC2-containing PBS are rapidly degraded and PBS containing PC3 accumulate (Fig 5). These two PBS forms are virtually equivalent in their photosynthetic light harvesting capabilities, but PC1 and PC2 have relatively large numbers of sulfur-containing amino acids while PC3 contains no methionines and only essential cysteines. PBS make up more than 50% of the total soluble protein in the cell, and this response releases more than a thousand sulfur atoms per PBS, making PBS both light harvesting antennae and sulfur storage organs. This response also leads to the more efficient use of cellular sulfur supplies by producing PC3-containing PBS.

We have uncovered several interesting and complex features of the system regulating this response. The increase in cpc3 RNA is regulated transcriptionally by an activator that binds in the region between -400 and -300 relative to the cpc3 transcription start site, while the loss of cpc1 and cpc2 RNA during sulfur limitation occurs post-transcriptionally. We are now focusing on the control of the interactions between cpc3, cpc1, and cpc2 RNA levels during sulfur limitation and the identity of the transcriptional regulatory system controlling cpc3 induction.

Fig 5. Change in PBS structure in response to changes in external sulfur availability.

 

Improving Light Harvesting for Efficient Biosolar Energy Production

Our reliance on fossil fuels is clearly not sustainable in the long term and is creating serious problems for our environment. We believe that using solar energy capture by cyanobacteria will help alleviate these problems. One major problem that must be overcome is the fact that photosynthetic microorganisms are typically capable of using only 10% of the photons available in full summer sunlight. In addition, traditional attempts to use such organisms in batch cultures leads to high levels of self-shading, which further reduces the overall solar conversion efficiency down to only a few percent- too low to be economically viable. We are working to develop cyanobacteria with modified light harvesting antennae that efficiently use all intensities and wavelengths of light for solar-based energy production. Our research is being carried out in collaboration with Dr. Roger Ely’s group at Oregon State University and Dr. Catherine Page’s group at the University of Oregon.