Bi-allelic JAM2 Variants Lead to Early-Onset Recessive Primary Familial Brain Calcification.

Primary familial brain calcification (PFBC) is a rare neurodegenerative disorder characterized by a combination of neurological, psychiatric, and cognitive decline associated with calcium deposition on brain imaging. To date, mutations in five genes have been linked to PFBC. However, more than 50% of individuals affected by PFBC have no molecular diagnosis. We report four unrelated families presenting with initial learning difficulties and seizures and later psychiatric symptoms, cerebellar ataxia, extrapyramidal signs, and extensive calcifications on brain imaging. Through a combination of homozygosity mapping and exome sequencing, we mapped this phenotype to chromosome 21q21.3 and identified bi-allelic variants in JAM2. JAM2 encodes for the junctional-adhesion-molecule-2, a key tight-junction protein in blood-brain-barrier permeability. We show that JAM2 variants lead to reduction of JAM2 mRNA expression and absence of JAM2 protein in patient's fibroblasts, consistent with a loss-of-function mechanism. We show that the human phenotype is replicated in the jam2 complete knockout mouse (jam2 KO). Furthermore, neuropathology of jam2 KO mouse showed prominent vacuolation in the cerebral cortex, thalamus, and cerebellum and particularly widespread vacuolation in the midbrain with reactive astrogliosis and neuronal density reduction. The regions of the human brain affected on neuroimaging are similar to the affected brain areas in the myorg PFBC null mouse. Along with JAM3 and OCLN, JAM2 is the third tight-junction gene in which bi-allelic variants are associated with brain calcification, suggesting that defective cell-to-cell adhesion and dysfunction of the movement of solutes through the paracellular spaces in the neurovascular unit is a key mechanism in CNS calcification

Inhibition of G-protein signalling in cardiac dysfunction of intellectual developmental disorder with cardiac arrhythmia (IDDCA) syndrome

Background: Pathogenic variants of GNB5 encoding the β5 subunit of the guanine nucleotide-binding protein cause IDDCA syndrome, an autosomal recessive neurodevelopmental disorder associated with cognitive disability and cardiac arrhythmia, particularly severe bradycardia. Methods: We used echocardiography and telemetric ECG recordings to investigate consequences of Gnb5 loss in mouse. Results: We delineated a key role of Gnb5 in heart sinus conduction and showed that Gnb5-inhibitory signalling is essential for parasympathetic control of heart rate (HR) and maintenance of the sympathovagal balance. Gnb5-/- mice were smaller and had a smaller heart than Gnb5+/+ and Gnb5+/-, but exhibited better cardiac function. Lower autonomic nervous system modulation through diminished parasympathetic control and greater sympathetic regulation resulted in a higher baseline HR in Gnb5-/- mice. In contrast, Gnb5-/- mice exhibited profound bradycardia on treatment with carbachol, while sympathetic modulation of the cardiac stimulation was not altered. Concordantly, transcriptome study pinpointed altered expression of genes involved in cardiac muscle contractility in atria and ventricles of knocked-out mice. Homozygous Gnb5 loss resulted in significantly higher frequencies of sinus arrhythmias. Moreover, we described 13 affected individuals, increasing the IDDCA cohort to 44 patients. Conclusions: Our data demonstrate that loss of negative regulation of the inhibitory G-protein signalling causes HR perturbations in Gnb5-/- mice, an effect mainly driven by impaired parasympathetic activity. We anticipate that unravelling the mechanism of Gnb5 signalling in the autonomic control of the heart will pave the way for future drug screening

PDXK mutations cause polyneuropathy responsive to pyridoxal 5'-phosphate supplementation.

OBJECTIVE: To identify disease-causing variants in autosomal recessive axonal polyneuropathy with optic atrophy and provide targeted replacement therapy. METHODS: We performed genome-wide sequencing, homozygosity mapping, and segregation analysis for novel disease-causing gene discovery. We used circular dichroism to show secondary structure changes and isothermal titration calorimetry to investigate the impact of variants on adenosine triphosphate (ATP) binding. Pathogenicity was further supported by enzymatic assays and mass spectroscopy on recombinant protein, patient-derived fibroblasts, plasma, and erythrocytes. Response to supplementation was measured with clinical validated rating scales, electrophysiology, and biochemical quantification. RESULTS: We identified biallelic mutations in PDXK in 5 individuals from 2 unrelated families with primary axonal polyneuropathy and optic atrophy. The natural history of this disorder suggests that untreated, affected individuals become wheelchair-bound and blind. We identified conformational rearrangement in the mutant enzyme around the ATP-binding pocket. Low PDXK ATP binding resulted in decreased erythrocyte PDXK activity and low pyridoxal 5'-phosphate (PLP) concentrations. We rescued the clinical and biochemical profile with PLP supplementation in 1 family, improvement in power, pain, and fatigue contributing to patients regaining their ability to walk independently during the first year of PLP normalization. INTERPRETATION: We show that mutations in PDXK cause autosomal recessive axonal peripheral polyneuropathy leading to disease via reduced PDXK enzymatic activity and low PLP. We show that the biochemical profile can be rescued with PLP supplementation associated with clinical improvement. As B6 is a cofactor in diverse essential biological pathways, our findings may have direct implications for neuropathies of unknown etiology characterized by reduced PLP levels. ANN NEUROL 2019;86:225-240

Endocrine and Growth Abnormalities in 4H Leukodystrophy Caused by Variants in POLR3A, POLR3B, and POLR1C.

CONTEXT: 4H or POLR3-related leukodystrophy is an autosomal recessive disorder typically characterized by hypomyelination, hypodontia, and hypogonadotropic hypogonadism, caused by biallelic pathogenic variants in POLR3A, POLR3B, POLR1C, and POLR3K. The endocrine and growth abnormalities associated with this disorder have not been thoroughly investigated to date. OBJECTIVE: To systematically characterize endocrine abnormalities of patients with 4H leukodystrophy. DESIGN: An international cross-sectional study was performed on 150 patients with genetically confirmed 4H leukodystrophy between 2015 and 2016. Endocrine and growth abnormalities were evaluated, and neurological and other non-neurological features were reviewed. Potential genotype/phenotype associations were also investigated. SETTING: This was a multicenter retrospective study using information collected from 3 predominant centers. PATIENTS: A total of 150 patients with 4H leukodystrophy and pathogenic variants in POLR3A, POLR3B, or POLR1C were included. MAIN OUTCOME MEASURES: Variables used to evaluate endocrine and growth abnormalities included pubertal history, hormone levels (estradiol, testosterone, stimulated LH and FSH, stimulated GH, IGF-I, prolactin, ACTH, cortisol, TSH, and T4), and height and head circumference charts. RESULTS: The most common endocrine abnormalities were delayed puberty (57/74; 77% overall, 64% in males, 89% in females) and short stature (57/93; 61%), when evaluated according to physician assessment. Abnormal thyroid function was reported in 22% (13/59) of patients. CONCLUSIONS: Our results confirm pubertal abnormalities and short stature are the most common endocrine features seen in 4H leukodystrophy. However, we noted that endocrine abnormalities are typically underinvestigated in this patient population. A prospective study is required to formulate evidence-based recommendations for management of the endocrine manifestations of this disorder

Self-forming dynamic membrane: a review

This paper presents self-forming dynamic membrane (SFDM) process as a promising alternative to membrane technology. A stable dynamic membrane (DM) is mainly composed of gel layer and cake layer which are responsible for filtration and biodegradation. An understanding of the formation mechanism and impact of operating conditions, support mesh, sludge properties and configuration on the formation, and characteristics of the dynamic membrane is essential to optimize the efficiency and reduce fouling propensity of this technology. The paper thus aims to promote its applicability on wastewater treatment. Review of the influence of mesh support properties, different operating conditions and sludge properties on SFDM performance, and cost comparison between MBR and SFDMBR technologies

Pectic Cell Wall Fragments Regulate Tobacco Thin-Cell-Layer Explant Morphogenesis.

Pectic fragments of cell wall polysaccharides, released from the walls of suspension-cultured sycamore cells by treatment with endopolygalacturonase, were tested for morphogenesis-regulating activity in a modified tobacco thin-cell-layer explant (TCL) bioassay (D. Mohnen, S. Eberhard, V. Marfa, N. Doubrava, P. Toubart, D. J. Gollin, T.A. Gruber, W. Nuri, P. Albersheim, and A. Darvill, manuscript submitted). The pectic fragments inhibited the formation of roots on TCLs grown on a root-inducing medium containing 15 micromolar indole-3-butyric acid and 0.5 micromolar kinetin. Addition of the pectic fragments to a root-inducing medium containing 7 micromolar indole-3-butyric acid and 0.15 micromolar kinetin caused roots to form on the basal end of TCLs. TCLs cultured on this medium in the absence of added pectic fragments formed roots along their entire length. The pectic fragments induced polar tissue enlargement and the formation of flowers on TCLs cultured on transition medium. The flower-inducing activity was stable to heat treatment and proteolytic digestion. Pectic fragments isolated from the walls of suspension-cultured tobacco cells were as effective as those from the walls of sycamore cells in inducing de novo flower formation in the TCLs. These results support the hypothesis that oligosaccharins from plant cell walls regulate morphogenesis

Selection of Cucumis melo L. subsp. melo 'Piel de Sapo' lines with high efficiency in Agrobacterium-mediated genetic transformation

XXVIII International Horticultural Congress on Science and Horticulture for People (IHC2010): International Symposium on Genomics and Genetic Transformation of Horticultural Crops.There are important hindrances for setting up a routine melon transformation protocol, including the strong effect of genotype and the high incidence of polyploid regenerants. Genetic transformation efficiencies for melon lines of Cucumis melo subsp. melo ‘Piel de Sapo’ inodorus type (e.g., ‘T111’), are lower than those obtained with the ‘Védrantais’ line or the accession C. melo subsp. agrestis ‘PI 161375’ (Castelblanque et al., 2008). Four near isogenic lines (NILs), SC 6-6, SC 7-2, SC 7-4, and SC 8-4, selected for their higher regeneration capacity from a population of 32 NILs, and the ‘T111’ line were used to determine their genetic transformation efficiency via A. tumefaciens. These NILs were developed from a doubled haploid line (DHL) population derived from the cross between the Korean accession ‘PI 161375’ and the Spanish ‘T111’. Transgenic melon plants were produced from ‘T111’ and three NILs, using cotyledon explants inoculated with an A. tumefaciens strain harboring a binary vector containing genes encoding β-glucuronidase activity and conferring tolerance to glyphosate, as selection marker. Cotyledonary explants were cultured on the regeneration medium supplemented with 4 mg•L-1 Basta herbicide. Putative transformed plantlets were tested for gene insertion 3-4 months after inoculation. Transformants were selected on the basis of glyphosate tolerance and GUS expression on leaves and roots. Following such protocol, the transformation efficiency has increased from a 3.3%, for the ‘Piel de Sapo’ line ‘T111’ to 23.3% for two of the selected NILs. Out of 105 transgenic plants produced, 11% were diploid and the rest were tetraploid. Higher ploidy levels usually result in sterility, and failure to perpetuate the transgenic line. Another source of explants is under study to facilitate the recovery of diploid transgenic plants.Peer reviewe

Oligogalacturonides inhibit the formation of roots on tobacco explants

α-1,4-Oligogalacturonides with degrees of polymerization (DPs) ranging from 6 to 18 or 2 to 8 were added to tobacco leaf explants and root formation was evaluated after 15 days of incubation. Auxin-induced formation of roots was inhibited by oligogalacturonides with DPs 6–18 but not by the oligogalacturonides with DPs 2–8. The inhibition of root formation by the larger oligogalacturonides was prevented by increasing the amount of auxin present in the medium. Oligogalacturonides (DPs 6–18) also inhibited root formation when added to tobacco thin cell-layer (TCL) explants in a medium that is known to induce the formation of roots. The addition of size-homogeneous oligogalacturonides, to either tobacco leaf explants or TCLs, established that oligogalacturonides with DPs between 10 and 14 were most active in inhibiting the formation of roots. These data suggest that oligogalacturonides of the same size as those known to elicit plant defense responses, and to affect floral development and membrane functions, also inhibit the induction of root morphogenesis in tobacco