c-Type cytochromes (cyt c) are proteins that undergo post-translational modification to covalently bind heme, which allows them to facilitate redox reactions in electron transport chains across all domains of life. Genomic evidence suggests that cyt c are involved in electron transfer processes among the Archaea, especially in members that produce or consume the potent greenhouse gas methane. However, neither the maturation machinery for cyt c in Archaea nor their role in methane metabolism has ever been functionally characterized. Here, we have used CRISPR-Cas9 genome editing tools to map a distinct pathway for cyt c biogenesis in the model methanogenic archaeon Methanosarcina acetivorans, and have also identified substrate-specific functional roles for cyt c during methanogenesis. Although the cyt c maturation machinery from M. acetivorans is universally conserved in the Archaea, our evolutionary analyses indicate that different clades of Archaea acquired this machinery through multiple independent horizontal gene transfer events from different groups of Bacteria. Overall, we demonstrate the convergent evolution of a novel Archaea-specific cyt c maturation machinery and its physiological role during methanogenesis, a process which contributes substantially to global methane emissions.
Keywords: Archaea; Methanosarcina acetivorans; cytochromes; energy conservation; infectious disease; methane; methanogens; microbiology.
Archaea are single-celled organisms that were discovered over half a century ago. Recently, there has been a renewed interest in these microbes because theyplay a key role in climate change by controlling greenhouse gas emissions, like methane. Indeed, methane-producing Archaea generate nearly 70% of the methane gas released into the atmosphere. A group of proteins called c-type cytochromes are essential to energy generation in several methane-producing archaea. However, it is a mystery how Archaea assemble their c-type cytochromes. In fact, genomic studies suggest that Archaea are missing some of the c-type cytochrome assembly machinery that bacteria use. This has led scientists to suspect that Archaea have an alternate mechanism for building these essential components. To solve this mystery, Gupta, Shalvarjian, and Nayak used CRISPR-Cas9 gene-editing tools to characterize which proteins are essential for c-type cytochrome production in Methanosarcina acetivorans, a species of Archaea that produces methane. These experiments showed that M. acetivorans discarded a few parts of the process used by bacteria to generate c-type cytochromes, streamlining the assembly of these proteins. By comparing the genes of different Archaeal species, Gupta, Shalvarjian and Nayak were able to determine that Archaea acquired the genes for producing c-type cytochromes from bacteria via horizontal gene transfer, a process in which genes move directly from one organism into another. The streamlining of the process took place later, as different Archaeal species evolved independently, but losing the same parts of the process. Gupta Shalvajiran and Nayak’s experiments also showed that c-type cytochromes are essential for the growth and fitness of methane-producing Archaea like M. acetivorans. The role of c-type cytochromes in methane production varies in different species of Archaea depending on their growth substrate or where they live. These results provide vital information about how Archaea produce methane, and the tools and techniques developed will aid further investigation of the role of Archaea in climate change.
© 2022, Gupta et al.