Periodic Reporting for period 3 - VirFix (Virus inhibition of oceanic CO2 fixation)
Okres sprawozdawczy: 2024-01-01 do 2025-06-30
Oceans are the ‘beating heart’ of Earth’s carbon cycle: with <1 billion tonnes of phytoplankton alive in the ocean at any one time but with 45 billion tonnes of new phytoplankton produced each year, phytoplankton have to reproduce themselves entirely, on average, 45 times a year, or roughly once a week. In contrast, the world’s land plants have a total biomass of 500 billion tonnes, most of it wood, but with much slower growth compared to phytoplankton plants reproduce themselves entirely only once every ten years. As a result, the functioning of this ‘fast’ marine phytoplankton carbon pump (i.e. the fixation of atmospheric CO2 into biomass and the subsequent sinking and burial of this biomass into the deep ocean) is critical for absorbing and sequestering the increased levels of CO2 currently entering the atmosphere from anthropogenic input. Cyanobacteria of the genera Prochlorococcus and Synechococcus are numerically the most abundant phytoplankton components in marine systems contributing 25% of global oceanic primary production but with this figure increasing to as much as 90% of the total in nutrient poor regions. We have discovered that viruses infecting these cyanobacteria (so called cyanophages), as well as directly controlling cell abundance through cell lysis, also shutdown CO2 fixation during the infection process. This has profound implications for surface ocean biogeochemistry, including carbon dynamics as well as the planet's climate. Since cyanophage outnumber their hosts 10-fold causing an estimated 10 to the power 25 infections a day, we have estimated that around 5 billion tonnes of carbon is lost to viral-induced inhibition of CO2 fixation per year (around 3-fold greater production than the sum of all coral reefs, salt marshes, estuaries, and marine macrophytes on Earth!).
Thus, Virfix had the following broad objectives: i) to obtain a mechanistic understanding of how viruses switch off CO2 fixation in these organisms as well as ii) model how this inhibition affects global estimates of marine primary production. We also sought to iii) understand much more broadly how viruses subvert the metabolism of their phytoplankton hosts particularly under different light and nutrient regimes.
Thus, Virfix had the following broad objectives: i) to obtain a mechanistic understanding of how viruses switch off CO2 fixation in these organisms as well as ii) model how this inhibition affects global estimates of marine primary production. We also sought to iii) understand much more broadly how viruses subvert the metabolism of their phytoplankton hosts particularly under different light and nutrient regimes.
We have identified a viral (cyanophage) protein that plays a role in inhibiting cyanobacterial CO2 fixation. This viral protein has been purified and shown to inhibit the key CO2 fixing enzyme RuBisCO purified from the host. However, inhibition of RuBisCO activity was not complete which leads us to believe there is another protein component(s) required for complete inhibition. This work is on-going. We have also expressed the cyanophage protein in a heterologous cyanobacterial host and shown that CO2 fixation is inhibited by 90%. In this heterologous host the cyanophage protein co-localises with an organelle where RuBisCo is housed.
Given that some cyanophage genomes also encode proteins that are directly involved in photosynthesis we have begun to study the function of these cyanophage proteins. To enable this, we are developing tools to make cyanophage gene knockouts. We have recently developed a random chemical mutagenesis system for constructing cyanophage mutants and are currently developing a positive selection strategy to knock out specific cyanophage genes. In addition, we are also using proteomics to elucidate the location of cyanophage proteins during the infection cycle so as to target proteins with interesting intracellular locations.
To understand how nutrient stress affects cyanophage infection dynamics we have used transcriptomics to identify genes specifically induced under stress conditions. We have identified a novel suite of genes specifically induced under a specific nutrient stress and moreover have found that this suite of genes are controlled by the hosts regulatory system. This highlights how cyanophage have evolved to exploit their host’s environmental sensing mechanisms to coordinate their own gene expression in response to resource limitation.
To determine how the carriage of host-like genes in cyanophage genomes is affected by environmental parameters we have successfully isolated several hundred new cyanophage from a set of cruise samples spanning the length of the Atlantic Ocean. These cyanophage are currently being purified and their genomes sequenced so that we can directly determine their gene content and how this correlates with the environmental parameters from which these phage were isolated.
Given that some cyanophage genomes also encode proteins that are directly involved in photosynthesis we have begun to study the function of these cyanophage proteins. To enable this, we are developing tools to make cyanophage gene knockouts. We have recently developed a random chemical mutagenesis system for constructing cyanophage mutants and are currently developing a positive selection strategy to knock out specific cyanophage genes. In addition, we are also using proteomics to elucidate the location of cyanophage proteins during the infection cycle so as to target proteins with interesting intracellular locations.
To understand how nutrient stress affects cyanophage infection dynamics we have used transcriptomics to identify genes specifically induced under stress conditions. We have identified a novel suite of genes specifically induced under a specific nutrient stress and moreover have found that this suite of genes are controlled by the hosts regulatory system. This highlights how cyanophage have evolved to exploit their host’s environmental sensing mechanisms to coordinate their own gene expression in response to resource limitation.
To determine how the carriage of host-like genes in cyanophage genomes is affected by environmental parameters we have successfully isolated several hundred new cyanophage from a set of cruise samples spanning the length of the Atlantic Ocean. These cyanophage are currently being purified and their genomes sequenced so that we can directly determine their gene content and how this correlates with the environmental parameters from which these phage were isolated.
We have already obtained significant new information on the function of cyanophage genes and how they affect photosynthesis in their cyanobacterial host. This has required the development of mutagenesis protocols as well as the use of state of the art ‘omics approaches. Our research is already producing high quality publications and is training a new cohort of young researchers in cyanophage biology specifically with respect to how these viruses modulate photosynthesis. The project is also benefitting from key national and international collaborations.