Deciphering functional redundancy and energetics of malate oxidation in mycobacteria.

Oxidation of malate to oxaloacetate, catalyzed by either malate dehydrogenase (Mdh) or malate quinone oxidoreductase (Mqo), is a critical step of the tricarboxylic acid cycle. Both Mqo and Mdh are found in most bacterial genomes, but the level of functional redundancy between these enzymes remains unclear. A bioinformatic survey revealed that Mqo was not as widespread as Mdh in bacteria but that it was highly conserved in mycobacteria. We therefore used mycobacteria as a model genera to study the functional role(s) of Mqo and its redundancy with Mdh. We deleted mqo from the environmental saprophyte Mycobacterium smegmatis, which lacks Mdh, and found that Mqo was essential for growth on nonfermentable carbon sources. On fermentable carbon sources, the Δmqo mutant exhibited delayed growth and lowered oxygen consumption and secreted malate and fumarate as terminal end products. Furthermore, heterologous expression of Mdh from the pathogenic species Mycobacterium tuberculosis shortened the delayed growth on fermentable carbon sources and restored growth on nonfermentable carbon sources at a reduced growth rate. In M. tuberculosis, CRISPR interference of either mdh or mqo expression resulted in a slower growth rate compared to controls, which was further inhibited when both genes were knocked down simultaneously. These data reveal that exergonic Mqo activity powers mycobacterial growth under nonenergy limiting conditions and that endergonic Mdh activity complements Mqo activity, but at an energetic cost for mycobacterial growth. We propose Mdh is maintained in slow-growing mycobacterial pathogens for use under conditions such as hypoxia that require reductive tricarboxylic acid cycle activity.

CRISPR interference identifies vulnerable cellular pathways with bactericidal phenotypes in Mycobacterium tuberculosis.

Mycobacterium tuberculosis remains a leading cause of death for which new drugs are needed. The identification of drug targets has been advanced by high-throughput and targeted genetic deletion strategies. Each though has limitations including the inability to distinguish between levels of vulnerability, lethality, and scalability as a molecular tool. Using mycobacterial CRISPR interference in combination with phenotypic screening, we have overcome these individual issues to investigate essentiality, vulnerability and lethality for 94 target genes from a diverse array of cellular pathways, many of which are potential antibiotic targets. Essential genes involved in cell wall synthesis and central cellular functions were equally vulnerable and often had bactericidal consequences. Conversely, essential genes involved in metabolism, oxidative phosphorylation, or amino acid synthesis were less vulnerable to inhibition and frequently bacteriostatic. In conclusion, this study provides novel insights into mycobacterial genetics and biology that will help to prioritize potential drug targets.

Three transposable elements exhibiting differential expression in pre-eclampsia overlap with enhancer regions.

Transposable elements (TEs) play a crucial role in placental development and dysfunction. Our study examined TE expression in pre-eclampsia (PE) using RNA-seq datasets. We identified differentially expressed TEs and explored the genomic location of the most significant TEs, investigating their possible regulatory roles. Notably, three TEs overlapped with putative enhancer regions, suggesting a potential regulatory impact on gene expression. These findings highlight the regulatory potential of TEs and their importance in placental development, supporting that TE dysregulation may contribute to PE pathogenesis.

Identification of Chemical Scaffolds That Inhibit the Mycobacterium tuberculosis Respiratory Complex Succinate Dehydrogenase.

Drug-resistant Mycobacterium tuberculosis is a significant cause of infectious disease morbidity and mortality for which new antimicrobials are urgently needed. Inhibitors of mycobacterial respiratory energy metabolism have emerged as promising next-generation antimicrobials, but a number of targets remain unexplored. Succinate dehydrogenase (SDH), a focal point in mycobacterial central carbon metabolism and respiratory energy production, is required for growth and survival in M. tuberculosis under a number of conditions, highlighting the potential of inhibitors targeting mycobacterial SDH enzymes. To advance SDH as a novel drug target in M. tuberculosis, we utilized a combination of biochemical screening and in-silico deep learning technologies to identify multiple chemical scaffolds capable of inhibiting mycobacterial SDH activity. Antimicrobial susceptibility assays show that lead inhibitors are bacteriostatic agents with activity against wild-type and drug-resistant strains of M. tuberculosis. Mode of action studies on lead compounds demonstrate that the specific inhibition of SDH activity dysregulates mycobacterial metabolism and respiration and results in the secretion of intracellular succinate. Interaction assays demonstrate that the chemical inhibition of SDH activity potentiates the activity of other bioenergetic inhibitors and prevents the emergence of resistance to a variety of drugs. Overall, this study shows that SDH inhibitors are promising next-generation antimicrobials against M. tuberculosis.

Unveiling the hidden players: The crucial role of transposable elements in the placenta and their potential contribution to pre-eclampsia.

The human placenta is a vital connection between maternal and fetal tissues, allowing for the exchange of molecules and modulation of immune interactions during pregnancy. Interestingly, some of the placenta's unique functions can be attributed to transposable elements (TEs), which are DNA sequences that have mobilised into the genome. Co-option throughout mammalian evolution has led to the generation of TE-derived regulators and TE-derived genes, some of which are expressed in the placenta but silenced in somatic tissues. TE genes encompass both TE-derived genes with a repeat element in the coding region and TE-derived regulatory regions such as alternative promoters and enhancers. Placental-specific TE genes are known to contribute to the placenta's unique functions, and interestingly, they are also expressed in some cancers and share similar functions. There is evidence to support that aberrant activity of TE genes may contribute to placental pathologies, cancer and autoimmunity. In this review, we highlight the crucial roles of TE genes in placental function, and how their dysregulation may lead to pre-eclampsia, a common and dangerous placental condition. We provide a summary of the functional TE genes in the placenta to offer insight into their significance in normal and abnormal human development. Ultimately, this review highlights an opportunity for future research to investigate the potential dysregulation of TE genes in the development of placental pathologies such as pre-eclampsia. Further understanding of TE genes and their role in the placenta could lead to significant improvements in maternal and fetal health.

Synthesis and Biological Evaluation of Aurachin D Analogues as Inhibitors of Mycobacterium tuberculosis Cytochrome bd Oxidase.

A revised total synthesis of aurachin D (1a), an isoprenoid quinolone alkaloid that targets Mycobacterium tuberculosis (Mtb) cytochrome bd (cyt-bd) oxidase, was accomplished using an oxazoline ring-opening reaction. The ring opening enabled access to a range of electron-poor analogues, while electron-rich analogues could be prepared using the Conrad-Limpach reaction. The aryl-substituted and side-chain-modified aurachin D analogues were screened for inhibition of Mtb cyt-bd oxidase and growth inhibition of Mtb. Nanomolar inhibition of Mtb cyt-bd oxidase was observed for the shorter-chain analogue 1d (citronellyl side chain) and the aryl-substituted analogues 1g/1k (fluoro substituent at C6/C7), 1t/1v (hydroxy substituent at C5/C6) and 1u/1w/1x (methoxy substituent at C5/C6/C7). Aurachin D and the analogues did not inhibit growth of nonpathogenic Mycobacterium smegmatis, but the citronellyl (1d) and 6-fluoro-substituted (1g) inhibitors from the Mtb cyt-bd oxidase assay displayed moderate growth inhibition against pathogenic Mtb (MIC = 4-8 µM).