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ABSTRACT Rubisco, the most prevalent protein on Earth, catalyzes both a reaction that initiates C₃ carbon fixation, and a reaction that initiates photorespiration, which stimulates protein synthesis. Regulation of the balance between these reactions under atmospheric CO₂ fluctuations remains poorly understood. We have hypothesized that vascular plants maintain organic carbon‐to‐nitrogen homeostasis by adjusting the relative activities of magnesium and manganese in chloroplasts to balance carbon fixation and nitrate assimilation rates. The following examined the influence of magnesium and manganese on carboxylation and oxygenation for rubisco purified from two ecotypes of Plantago lanceolataL.: one adapted to the elevated CO₂ atmospheres that occur near a natural CO₂ spring and the other adapted to more typical CO₂ atmospheres that occur nearby. The plastid DNA coding for the large unit of rubisco was similar in both ecotypes. The kinetics of rubiscos from the two ecotypes differed more when associated with manganese than magnesium. Specificity for CO₂over O₂ (S_c/o) for rubisco from both ecotypes was higher when the enzymes were bound to magnesium than manganese. Differences in the responses of rubisco from P. lanceolata to the metals may account for the adaptation of this species to different CO₂ environments. 

Abstract The behavior of many plant enzymes depends on the metals and other ligands to which they are bound. A previous study demonstrated that tobacco Rubisco binds almost equally to magnesium and manganese and rapidly exchanges one metal for the other. The present study characterizes the kinetics of Rubisco and the plastidial malic enzyme when bound to either metal. When Rubisco purified from five C₃species was bound to magnesium rather than manganese, the specificity for CO₂over O₂, (S_c/o) increased by 25% and the ratio of the maximum velocities of carboxylation / oxygenation (V_cmax/V_omax) increased by 39%. For the recombinant plastidial malic enzyme, the forward reaction (malate decarboxylation) was 30% slower and the reverse reaction (pyruvate carboxylation) was three times faster when bound to manganese rather than magnesium. Adding 6‐phosphoglycerate and NADP⁺inhibited carboxylation and oxygenation when Rubisco was bound to magnesium and stimulated oxygenation when it was bound to manganese. Conditions that favored RuBP oxygenation stimulated Rubisco to convert as much as 15% of the total RuBP consumed into pyruvate. These results are consistent with a stromal biochemical pathway in which (1) Rubisco when associated with manganese converts a substantial amount of RuBP into pyruvate, (2) malic enzyme when associated with manganese carboxylates a substantial portion of this pyruvate into malate, and (3) chloroplasts export additional malate into the cytoplasm where it generates NADH for assimilating nitrate into amino acids. Thus, plants may regulate the activities of magnesium and manganese in leaves to balance organic carbon and organic nitrogen as atmospheric CO₂fluctuates. 

Differences in gene expression within tissues can lead to differences in tissue function. Understanding the transcriptome of a species helps elucidate the molecular mechanisms underlying phenotypic divergence. According to the presence or absence of a reference genome of for a studied species, transcriptome analyses can be divided into reference‑based and reference‑free methods, respectively. Presently, comparisons of complete transcriptome analysis results between those two methods are still rare. In this study, we compared the cochlear transcriptome analysis results of greater horseshoe bats (Rhinolophus ferrumequinum) from three lineages in China with different acoustic phenotypes using reference‑based and reference‑free methods to explore their differences in subsequent analysis. The results gained by reference-based results had lower false-positive rates and were more accurate because differentially expressed genes among the three populations obtained by this method had greater reliability and a higher annotation rate. Some phenotype-related enrichment terms, including those related to inorganic molecules and proton transmembrane channels, were also obtained only by the reference-based method. However, the reference‑based method might have the limitation of incomplete information acquisition. Thus, we believe that a combination of reference‑free and reference‑based methods is ideal for transcriptome analyses. The results of our study provided a reference for the selection of transcriptome analysis methods in the future. 

Photorespiration, or C2 photosynthesis, is generally considered a futile cycle that potentially decreases photosynthetic carbon fixation by more than 25%. Nonetheless, many essential processes, such as nitrogen assimilation, C1 metabolism, and sulfur assimilation, depend on photorespiration. Most studies of photosynthetic and photorespiratory reactions are conducted with magnesium as the sole metal cofactor despite many of the enzymes involved in these reactions readily associating with manganese. Indeed, when manganese is present, the energy efficiency of these reactions may improve. This review summarizes some commonly used methods to quantify photorespiration, outlines the influence of metal cofactors on photorespiratory enzymes, and discusses why photorespiration may not be as wasteful as previously believed.