Dietary intake of L-carnitine can promote cardiovascular diseases in humans through microbial production of trimethylamine (TMA) and its subsequent oxidation to trimethylamine N-oxide by hepatic flavin-containing monooxygenases. Although our microbiota are responsible for TMA formation from carnitine, the underpinning molecular and biochemical mechanisms remain unclear. In this study, using bioinformatics approaches, we first identified a two-component Rieske-type oxygenase/reductase (CntAB) and associated gene cluster proposed to be involved in carnitine metabolism in representative genomes of the human microbiota. CntA belongs to a group of previously uncharacterized Rieske-type proteins and has an unusual "bridging" glutamate but not the aspartate residue, which is believed to facilitate intersubunit electron transfer between the Rieske center and the catalytic mononuclear iron center. Using Acinetobacter baumannii as the model, we then demonstrate that cntAB is essential in carnitine degradation to TMA. Heterologous overexpression of cntAB enables Escherichia coli to produce TMA, confirming that these genes are sufficient in TMA formation. Site-directed mutagenesis experiments have confirmed that this unusual "bridging glutamate" residue in CntA is essential in catalysis and neither mutant (E205D, E205A) is able to produce TMA. Taken together, the data in our study reveal the molecular and biochemical mechanisms underpinning carnitine metabolism to TMA in human microbiota and assign the role of this novel group of Rieske-type proteins in microbial carnitine metabolism. methylated amine metabolism | comparative genomics | gut microbiota

A key question in the secondary growth of trees is how differentiation of the vascular cambium cells is directed to concurrently form two different tissues: xylem or phloem. class III homeodomain-leucine zipper (HD-Zip III) genes are known to play critical roles in the initiation, patterning, and differentiation of the vascular system in the process of primary and secondary growth. However, the mechanism of how these genes control secondary vascular differentiation is unknown. Here, we show that a Populus class III HD-Zip gene, PtrHB7, was preferentially expressed in cambial zone. PtrHB7-suppressed plants displayed significant changes in vascular tissues with a reduction in xylem but increase in phloem. Transcriptional analysis revealed that genes regulating xylem differentiation were down-regulated, whereas genes regulating phloem differentiation were up-regulated. Correspondingly, PtrHB7 overexpression enhanced differentiation of cambial cells toward xylem cells but inhibited phloem differentiation. PtrHB7 regulation on cambial cell differentiation was associated with its transcript abundance. Together, the results demonstrated that PtrHB7 plays a critical role in controlling a balanced differentiation between secondary xylem and phloem tissues in the process of Populus secondary growth in a dosage-dependent manner.

Background Plant secondary growth depends on the activity of the vascular cambium, which produces xylem and phloem. Wood derived from xylem is the most abundant form of biomass globally and has played key socio-economic and subsistence roles throughout human history. However, despite intensive study of vascular development, the full diversity of cell types and the gene networks engaged are still poorly understood. Results Here, we have applied an optimized protoplast isolation protocol and RNA sequencing to characterize the high-resolution single-cell transcriptional landscape of highly lignified poplar stems. We identify 20 putative cell clusters with a series of novel cluster-specific marker genes and find that these cells are highly heterogeneous based on the transcriptome. Analysis of these marker genes’ expression dynamics enables reconstruction of the cell differentiation trajectories involved in phloem and xylem development. We find that different cell clusters exhibit distinct patterns of phytohormone responses and emphasize the use of our data to predict potential gene redundancy and identify candidate genes related to vascular development in trees. Conclusions These findings establish the transcriptional landscape of major cell types of poplar stems at single-cell resolution and provide a valuable resource for investigating basic principles of vascular cell specification and differentiation in trees.