While lignin geochemistry has been extensively investigated in the Amazon River, little is known about lignin distribution and dynamics within deep, stratified river channels or its transformations within soils prior to delivery to rivers. We characterized lignin phenols in soils, river particulate organic matter (POM), and dissolved organic matter (DOM) across a 4 km elevation gradient in the Madre de Dios River system, Peru, as well as in marine sediments to investigate the source‐to‐sink evolution of lignin. In soils, we found more oxidized lignin in organic horizons relative to mineral horizons. The oxidized lignin signature was maintained during transfer into rivers, and lignin was a relatively constant fraction of bulk organic carbon in soils and riverine POM. Lignin in DOM became increasingly oxidized downstream, indicating active transformation of dissolved lignin during transport, especially in the dry season. In contrast, POM accumulated undegraded lignin downstream during the wet season, suggesting that terrestrial input exceeded in‐river degradation. We discovered high concentrations of relatively undegraded lignin in POM at depth in the lower Madre de Dios River in both seasons, revealing a woody undercurrent for its transfer within these deep rivers. Our study of lignin evolution in the soil‐river‐ocean continuum highlights important seasonal and depth variations of river carbon components and their connection to soil carbon pools, providing new insights into fluvial carbon dynamics associated with the transfer of lignin biomarkers from source to sink.

The history of Earth's carbon cycle reflects trends in atmospheric composition convolved with the evolution of photosynthesis. Fortunately, key parts of the carbon cycle have been recorded in the carbon isotope ratios of sedimentary rocks. The dominant model used to interpret this record as a proxy for ancient atmospheric CO2 is based on carbon isotope fractionations of modern photoautotrophs, and longstanding questions remain about how their evolution might have impacted the record. We interrogated the intersection of environment and evolution by measuring both biomass (ϵp) and enzymatic (ϵRubisco) carbon isotope fractionations of a cyanobacterial strain (Synechococcus elongatus PCC 7942) solely expressing a putative ancestral Form 1B rubisco dating to >>1 Ga. This strain, nicknamed ANC, grows in ambient pCO2 and displays larger ϵp values than WT, despite having a much smaller ϵRubisco (17.23 ± 0.61‰ vs. 25.18 ± 0.31‰, respectively). Measuring both enzymatic and biomass fractionation revealed a surprising result—ANC ϵp exceeded ANC ϵRubisco in all conditions tested, violating prevailing models of cyanobacterial carbon isotope fractionation. However, these models were corrected by accounting for cyanobacterial physiology, notably the CO2 concentrating mechanism (CCM). Our modified model indicated that powered inorganic carbon uptake systems contribute to ϵp, and this effect is exacerbated in ANC. These data suggested that understanding the evolution of both the CCM and rubisco is critical for interpreting the carbon isotope record, and that large fluctuations in the record may reflect the evolving efficiency of carbon fixing metabolisms as well as changes in atmospheric CO2.

Form I rubiscos evolved in Cyanobacteria ≥ 2.5 billion years ago and are enzymatically unique due to the presence of small subunits (RbcS) capping both ends of an octameric large subunit (RbcL) rubisco assembly to form a hexadecameric (L8S8) holoenzyme. Although RbcS was previously thought to be integral to Form I rubisco stability, the recent discovery of a closely related sister clade of octameric rubiscos (Form I’; L8) demonstrates that the L8 complex can assemble without small subunits (Banda et al. 2020). Rubisco also displays a kinetic isotope effect (KIE) where the 3PG product is depleted in 13C relative to 12C. In Cyanobacteria, only two Form I KIE measurements exist, making interpretation of bacterial carbon isotope data difficult. To aid comparison, we measured in vitro the KIEs of Form I’ (Candidatus Promineofilum breve) and Form I (Synechococcus elongatus PCC 6301) rubiscos and found the KIE to be smaller in the L8 rubisco (16.25 ± 1.36‰ vs. 22.42 ± 2.37‰, respectively). Therefore, while small subunits may not be necessary for protein stability, they may affect the KIE. Our findings may provide insight into the function of RbcS and allow more refined interpretation of environmental carbon isotope data.

The history of Earth’s carbon cycle reflects trends in atmospheric composition convolved with the evolution of photosynthesis. Fortunately, key parts of the carbon cycle have been recorded in the carbon isotope ratios of sedimentary rocks. The dominant model used to interpret this record as a proxy for ancient atmospheric CO 2 is based on carbon isotope fractionations of modern photoautotrophs, and longstanding questions remain about how their evolution might have impacted the record. Therefore, we measured both biomass (ε p ) and enzymatic (ε Rubisco ) carbon isotope fractionations of a cyanobacterial strain ( Synechococcus elongatus PCC 7942) solely expressing a putative ancestral Form 1B rubisco dating to ≫1 Ga. This strain, nicknamed ANC, grows in ambient pCO 2 and displays larger ε p values than WT, despite having a much smaller ε Rubisco (17.23 ± 0.61‰ vs. 25.18 ± 0.31‰, respectively). Surprisingly, ANC ε p exceeded ANC ε Rubisco in all conditions tested, contradicting prevailing models of cyanobacterial carbon isotope fractionation. Such models can be rectified by introducing additional isotopic fractionation associated with powered inorganic carbon uptake mechanisms present in Cyanobacteria, but this amendment hinders the ability to accurately estimate historical pCO 2 from geological data. Understanding the evolution of rubisco and the CO 2 concentrating mechanism is therefore critical for interpreting the carbon isotope record, and fluctuations in the record may reflect the evolving efficiency of carbon fixing metabolisms in addition to changes in atmospheric CO 2 .

Plant litter decomposition is a major nutrient input to terrestrial ecosystems that is primarily driven by microorganisms. Litter decomposition results in a flow of dissolved organic carbon (DOC) that links above-ground decomposition to below-ground microbial processes. Litter decomposition is expected to be altered by human-induced global disturbances—specifically nitrogen deposition and altered intensity and frequency of precipitation events—but little is known about impacts on the mobile pool of DOC. This study investigated the effect of simulated nitrogen deposition and increased precipitation events on microbially-driven carbon flow during short-term litter decomposition using a ‘common garden’ experimental design with microcosms containing sterile sand and blue grama grass litter inoculated with different soil microbial communities. Respiration (CO2) was measured throughout the experiment while microbial biomass carbon and nitrogen were quantified at the end. Overall, nitrogen deposition decoupled CO2 and DOC during short-term litter decomposition with respiration increasing and no affect on DOC concentration. Moreover, nitrogen deposition increased microbial biomass and had no effect on carbon use efficiency (CUE). Simulated precipitation events significantly increased DOC concentrations, decreased CUE, increased the microbial metabolic quotient (qCO2), and greatly altered microbial composition and diversity. These findings highlight the complex interactions and responses of surface litter decomposers to the combined effects of climate change and supports the need for more research into how varying microbiomes will respond to different global change scenarios. Furthermore, this study clearly indicates that any increases in soil carbon sequestration from nitrogen deposition are unlikely to arise from a larger supply of DOC.