Natural Product Screening Reveals Naphthoquinone Complex I Bypass Factors.

Deficiency of mitochondrial complex I is encountered in both rare and common diseases, but we have limited therapeutic options to treat this lesion to the oxidative phosphorylation system (OXPHOS). Idebenone and menadione are redox-active molecules capable of rescuing OXPHOS activity by engaging complex I-independent pathways of entry, often referred to as "complex I bypass." In the present study, we created a cellular model of complex I deficiency by using CRISPR genome editing to knock out Ndufa9 in mouse myoblasts, and utilized this cell line to develop a high-throughput screening platform for novel complex I bypass factors. We screened a library of ~40,000 natural product extracts and performed bioassay-guided fractionation on a subset of the top scoring hits. We isolated four plant-derived 1,4-naphthoquinone complex I bypass factors with structural similarity to menadione: chimaphilin and 3-chloro-chimaphilin from Chimaphila umbellata and dehydro-α-lapachone and dehydroiso-α-lapachone from Stereospermum euphoroides. We also tested a small number of structurally related naphthoquinones from commercial sources and identified two additional compounds with complex I bypass activity: 2-methoxy-1,4-naphthoquinone and 2-methoxy-3-methyl-1,4,-naphthoquinone. The six novel complex I bypass factors reported here expand this class of molecules and will be useful as tool compounds for investigating complex I disease biology.

The Prochlorococcus carbon dioxide-concentrating mechanism: evidence of carboxysome-associated heterogeneity.

The ability of Prochlorococcus to numerically dominate open ocean regions and contribute significantly to global carbon cycles is dependent in large part on its effectiveness in transforming light energy into compounds used in cell growth, maintenance, and division. Integral to these processes is the carbon dioxide-concentrating mechanism (CCM), which enhances photosynthetic CO2 fixation. The CCM involves both active uptake systems that permit intracellular accumulation of inorganic carbon as the pool of bicarbonate and the system of HCO3 (-) conversion into CO2. The latter is located in the carboxysome, a microcompartment designed to promote the carboxylase activity of Rubisco. This study presents a comparative analysis of several facets of the Prochlorococcus CCM. Our analyses indicate that a core set of CCM components is shared, and their genomic organization is relatively well conserved. Moreover, certain elements, including carboxysome shell polypeptides CsoS1 and CsoS4A, exhibit striking conservation. Unexpectedly, our analyses reveal that the carbonic anhydrase (CsoSCA) and CsoS2 shell polypeptide have diversified within the lineage. Differences in csoSCA and csoS2 are consistent with a model of unequal rates of evolution rather than relaxed selection. The csoS2 and csoSCA genes form a cluster in Prochlorococcus genomes, and we identified two conserved motifs directly upstream of this cluster that differ from the motif in marine Synechococcus and could be involved in regulation of gene expression. Although several elements of the CCM remain well conserved in the Prochlorococcus lineage, the evolution of differences in specific carboxysome features could in part reflect optimization of carboxysome-associated processes in dissimilar cellular environments.