A novel role for kynurenine 3-monooxygenase in mitochondrial dynamics.

The enzyme kynurenine 3-monooxygenase (KMO) operates at a critical branch-point in the kynurenine pathway (KP), the major route of tryptophan metabolism. As the KP has been implicated in the pathogenesis of several human diseases, KMO and other enzymes that control metabolic flux through the pathway are potential therapeutic targets for these disorders. While KMO is localized to the outer mitochondrial membrane in eukaryotic organisms, no mitochondrial role for KMO has been described. In this study, KMO deficient Drosophila melanogaster were investigated for mitochondrial phenotypes in vitro and in vivo. We find that a loss of function allele or RNAi knockdown of the Drosophila KMO ortholog (cinnabar) causes a range of morphological and functional alterations to mitochondria, which are independent of changes to levels of KP metabolites. Notably, cinnabar genetically interacts with the Parkinson’s disease associated genes Pink1 and parkin, as well as the mitochondrial fission gene Drp1, implicating KMO in mitochondrial dynamics and mitophagy, mechanisms which govern the maintenance of a healthy mitochondrial network. Overexpression of human KMO in mammalian cells finds that KMO plays a role in the post-translational regulation of DRP1. These findings reveal a novel mitochondrial role for KMO, independent from its enzymatic role in the kynurenine pathway

BRAF^V600E-mutated serrated colorectal neoplasia drives transcriptional activation of cholesterol metabolism

Abstract BRAF mutations occur early in serrated colorectal cancers, but their long-term influence on tissue homeostasis is poorly characterized. We investigated the impact of short-term (3 days) and long-term (6 months) expression of Braf V600E in the intestinal tissue of an inducible mouse model. We show that Braf V600E perturbs the homeostasis of intestinal epithelial cells, with impaired differentiation of enterocytes emerging after prolonged expression of the oncogene. Moreover, Braf V600E leads to a persistent transcriptional reprogramming with enrichment of numerous gene signatures indicative of proliferation and tumorigenesis, and signatures suggestive of metabolic rewiring. We focused on the top-ranking cholesterol biosynthesis signature and confirmed its increased expression in human serrated lesions. Functionally, the cholesterol lowering drug atorvastatin prevents the establishment of intestinal crypt hyperplasia in Braf V600E -mutant mice. Overall, our work unveils the long-term impact of Braf V600E expression in intestinal tissue and suggests that colorectal cancers with mutations in BRAF might be prevented by statins

Mutations in SLC29A3, Encoding an Equilibrative Nucleoside Transporter ENT3, Cause a Familial Histiocytosis Syndrome (Faisalabad Histiocytosis) and Familial Rosai-Dorfman Disease

The histiocytoses are a heterogeneous group of disorders characterised by an excessive number of histiocytes. In most cases the pathophysiology is unclear and treatment is nonspecific. Faisalabad histiocytosis (FHC) (MIM 602782) has been classed as an autosomal recessively inherited form of histiocytosis with similarities to Rosai-Dorfman disease (RDD) (also known as sinus histiocytosis with massive lymphadenopathy (SHML)). To elucidate the molecular basis of FHC, we performed autozygosity mapping studies in a large consanguineous family and identified a novel locus at chromosome 10q22.1. Mutation analysis of candidate genes within the target interval identified biallelic germline mutations in SLC29A3 in the FHC kindred and in two families reported to have familial RDD. Analysis of SLC29A3 expression during mouse embryogenesis revealed widespread expression by e14.5 with prominent expression in the central nervous system, eye, inner ear, and epithelial tissues including the gastrointestinal tract. SLC29A3 encodes an intracellular equilibrative nucleoside transporter (hENT3) with affinity for adenosine. Recently germline mutations in SLC29A3 were also described in two rare autosomal recessive disorders with overlapping phenotypes: (a) H syndrome (MIM 612391) that is characterised by cutaneous hyperpigmentation and hypertrichosis, hepatomegaly, heart anomalies, hearing loss, and hypogonadism; and (b) PHID (pigmented hypertrichosis with insulin-dependent diabetes mellitus) syndrome. Our findings suggest that a variety of clinical diagnoses (H and PHID syndromes, FHC, and familial RDD) can be included in a new diagnostic category of SLC29A3 spectrum disorder

Enteropathogenic E.coli type III effectors EspG and espG2 disrupt the microtubule network of intestinal epithelial cells

Enteropathogenic Escherichia coli infection of intestinal epithelial cells leads to localized depletion of the microtubule cytoskeleton, an effect that is dependent on delivery of type III translocated effector proteins EspG and Orf3 (designated EspG2) to the site of depletion. Microtubule depletion involved disruption rather than displacement of microtubules

Glomerular involvement in the arthrogryposis, renal dysfunction and cholestasis syndrome

BACKGROUND: Arthrogryposis, renal dysfunction and cholestasis (ARC) syndrome is a multisystem autosomal-recessive disorder caused by defects in the VPS33B and VIPAR genes, involved in localization of apical membrane proteins. Affected children usually die by 1 year of age, often secondary to infective complications. The classic renal manifestation previously described in ARC syndrome is proximal-tubular dysfunction. The aim of this study is to gain further insight into the renal manifestations of this syndrome. METHODS: Clinical review of three cases of ARC syndrome presenting to a tertiary centre. Together with measurement of VPS33B and VIPAR protein expression in the human glomerulus. RESULTS: The cases demonstrated severe failure to thrive and in addition to commonly described features profound proteinuria and albuminuria, together with hypoalbuminaemia, suggesting glomerular involvement of this syndrome. Western blotting of conditionally immortalized human glomerular cells and ex vivo immunofluorescent analysis of the human glomerulus revealed that VPS33B and VIPAR were highly expressed in glomerular endothelium, and podocytes, but not in the mesangium. CONCLUSIONS: ARC syndrome affects the glomerulus as well as the proximal tubule in the kidney. Our molecular studies suggest that both cell types that constitute the glomerular filtration barrier are affected in this condition, providing an explanation for the albuminuria that we have observed in our cases

Bypassing mitochondrial defects rescues Huntington's phenotypes in Drosophila

Huntington's disease (HD) is a fatal neurodegenerative disease with limited treatment options. Human and animal studies have suggested that metabolic and mitochondrial dysfunctions contribute to HD pathogenesis. Here, we use high-resolution respirometry to uncover defective mitochondrial oxidative phosphorylation and electron transfer capacity when a mutant huntingtin fragment is targeted to neurons or muscles in Drosophila and find that enhancing mitochondrial function can ameliorate these defects. In particular, we find that co-expression of parkin, an E3 ubiquitin ligase critical for mitochondrial dynamics and homeostasis, produces significant enhancement of mitochondrial respiration when expressed either in neurons or muscles, resulting in significant rescue of neurodegeneration, viability and longevity in HD model flies. Targeting mutant HTT to muscles results in larger mitochondria and higher mitochondrial mass, while co-expression of parkin increases mitochondrial fission and decreases mass. Furthermore, directly addressing HD-mediated defects in the fly's mitochondrial electron transport system, by rerouting electrons to either bypass mitochondrial complex I or complexes III-IV, significantly increases mitochondrial respiration and results in a striking rescue of all phenotypes arising from neuronal mutant huntingtin expression. These observations suggest that bypassing impaired mitochondrial respiratory complexes in HD may have therapeutic potential for the treatment of this devastating disorder

A novel role for kynurenine 3-monooxygenase in mitochondrial dynamics.

The enzyme kynurenine 3-monooxygenase (KMO) operates at a critical branch-point in the kynurenine pathway (KP), the major route of tryptophan metabolism. As the KP has been implicated in the pathogenesis of several human diseases, KMO and other enzymes that control metabolic flux through the pathway are potential therapeutic targets for these disorders. While KMO is localized to the outer mitochondrial membrane in eukaryotic organisms, no mitochondrial role for KMO has been described. In this study, KMO deficient Drosophila melanogaster were investigated for mitochondrial phenotypes in vitro and in vivo. We find that a loss of function allele or RNAi knockdown of the Drosophila KMO ortholog (cinnabar) causes a range of morphological and functional alterations to mitochondria, which are independent of changes to levels of KP metabolites. Notably, cinnabar genetically interacts with the Parkinson's disease associated genes Pink1 and parkin, as well as the mitochondrial fission gene Drp1, implicating KMO in mitochondrial dynamics and mitophagy, mechanisms which govern the maintenance of a healthy mitochondrial network. Overexpression of human KMO in mammalian cells finds that KMO plays a role in the post-translational regulation of DRP1. These findings reveal a novel mitochondrial role for KMO, independent from its enzymatic role in the kynurenine pathway

Generation of Distinct Differentially Culturable Forms of Burkholderia following Starvation at Low Temperature

 Bacteria have developed unique mechanisms to adapt to environmental stresses and challenges of the immune system. Here, we report that Burkholderia pseudomallei, the causative agent of melioidosis, and its laboratory surrogate, Burkholderia thailandensis, utilize distinct mechanisms for surviving starvation at different incubation temperatures. At 21°C, Burkholderia are present as short rods which can rapidly reactivate and form colonies on solid media. At 4°C, Burkholderia convert into coccoid forms that cannot be cultured on solid agar but can be resuscitated in liquid media supplemented with supernatant obtained from logarithmic phase cultures of B. thailandensis, or catalase and Tween 80, thus displaying characteristics of differentially culturable bacteria (DCB). These DCB have low intensity fluorescence when stained with SYTO 9, have an intact cell membrane (propidium iodide negative), and contain 16S rRNA at levels comparable with growing cells. We also present evidence that lytic transglycosylases, a family of peptidoglycan-remodeling enzymes, are involved in the generation of coccoid forms and their resuscitation to actively growing cells. A B. pseudomallei ΔltgGCFD mutant with four ltg genes deleted did not produce coccoid forms at 4°C and could not be resuscitated in the liquid media evaluated. Our findings provide insights into the adaptation of Burkholderia to nutrient limitation and the generation of differentially culturable bacteria.  Bacteria have developed unique mechanisms to adapt to environmental stresses and challenges of the immune system. Here, we report that Burkholderia pseudomallei, the causative agent of melioidosis, and its laboratory surrogate, Burkholderia thailandensis, utilize distinct mechanisms for surviving starvation at different incubation temperatures. At 21°C, Burkholderia are present as short rods which can rapidly reactivate and form colonies on solid media. At 4°C, Burkholderia convert into coccoid forms that cannot be cultured on solid agar but can be resuscitated in liquid media supplemented with supernatant obtained from logarithmic phase cultures of B. thailandensis, or catalase and Tween 80, thus displaying characteristics of differentially culturable bacteria (DCB). These DCB have low intensity fluorescence when stained with SYTO 9, have an intact cell membrane (propidium iodide negative), and contain 16S rRNA at levels comparable with growing cells. We also present evidence that lytic transglycosylases, a family of peptidoglycan-remodeling enzymes, are involved in the generation of coccoid forms and their resuscitation to actively growing cells. A B. pseudomallei ΔltgGCFD mutant with four ltg genes deleted did not produce coccoid forms at 4°C and could not be resuscitated in the liquid media evaluated. Our findings provide insights into the adaptation of Burkholderia to nutrient limitation and the generation of differentially culturable bacteria. </p